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Sample records for caput epididymal spermatozoa

  1. Synthesis and secretion of proteins by perifused caput epididymal tubules, and association of secreted proteins with spermatozoa

    International Nuclear Information System (INIS)

    Klinefelter, G.R.; Hamilton, D.W.

    1985-01-01

    We have used perifusion organ culture of proximal and distal caput epididymal tubules of the rat to study the secretion of proteins by epididymal epithelium and uptake of the luminal radioactive proteins by sperm. The amount of incorporation of L-[35S]methionine into luminal fluid proteins was time dependent and completely inhibited by cycloheximide. The association of labeled proteins with cultured sperm was also dependent on time and continuous, with sperm still acquiring labeled luminal proteins after protein synthesis was arrested. A Mr = 46,000 molecule was found to be heavily labeled in luminal fluid and sperm extracts. Fluorograms of all L-[35S]methionine extracts immunoprecipitated using an antiepididymal alpha-lactalbumin antibody (Klinefelter and Hamilton, 1984) showed labeling of an Mr = 18,000 molecule and, in addition, the Mr = 46,000 molecule, but immunostaining was specific only for the Mr = 18,000 molecule and the heavy chain of the immunoglobulin. We suggest that the Mr = 46,000 molecule may be galactosyltransferase. Galactose oxidase-NaB[3H]4 labeling of the cultured caput sperm cell surface revealed a Mr = 23,000 molecule that was able to be immunoprecipitated with antiepididymal alpha-lactalbumin antibody. Our data suggest that this cell surface molecule is similar to one component of the fluid epididymal alpha-lactalbumin-like complex and, in addition, show that glycosylation of the sperm surface can occur in the caput epididymidis

  2. Glycoproteins of bovine epididymal spermatozoa--a cytochemical study.

    Science.gov (United States)

    Sinowatz, F; Friess, A E; Wrobel, K H

    1984-01-01

    Modifications in bull sperm plasmamembrane during epididymal passage were investigated by the use of four different lectins: Concanavalin A (Con A); Ricinus communis I (RCA1); Wheat germ agglutinin (WGA); Ulex europaeus agglutinin I (UEA1). During sperm passage from caput to cauda epididymidis agglutination by RCA1 and WGA distinctly increased. Similar but somewhat less pronounced difference in the agglutinability was found for Con A. No agglutination was observed with UEA1. Ultrastructural examination of Con A binding sites on sperm plasma membrane with a Con A-horseradish peroxidase-gold technique (Con A-HRP-G) revealed a significant increase in the number of gold granules on the sperm tails during the epididymal passage of spermatozoa. No change in WGA-binding sites was observed between caput and cauda spermatozoa using a WGA-peroxidase method.

  3. Sodium coupled bicarbonate influx regulates intracellular and apical pH in cultured rat caput epididymal epithelium.

    Science.gov (United States)

    Zuo, Wu-Lin; Li, Sheng; Huang, Jie-Hong; Yang, Deng-Liang; Zhang, Geng; Chen, Si-Liang; Ruan, Ye-Chun; Ye, Ke-Nan; Cheng, Christopher H K; Zhou, Wen-Liang

    2011-01-01

    The epithelium lining the epididymis provides an optimal acidic fluid microenvironment in the epididymal tract that enable spermatozoa to complete the maturation process. The present study aims to investigate the functional role of Na(+)/HCO(3)(-) cotransporter in the pH regulation in rat epididymis. Immunofluorescence staining of pan cytokeratin in the primary culture of rat caput epididymal epithelium showed that the system was a suitable model for investigating the function of epididymal epithelium. Intracellular and apical pH were measured using the fluorescent pH sensitive probe carboxy-seminaphthorhodafluor-4F acetoxymethyl ester (SNARF-4F) and sparklet pH electrode respectively to explore the functional role of rat epididymal epithelium. In the HEPES buffered Krebs-Henseleit (KH) solution, the intracellular pH (pHi) recovery from NH(4)Cl induced acidification in the cultured caput epididymal epithelium was completely inhibited by amiloride, the inhibitor of Na(+)/H(+) exchanger (NHE). Immediately changing of the KH solution from HEPES buffered to HCO(3)(-) buffered would cause another pHi recovery. The pHi recovery in HCO(3)(-) buffered KH solution was inhibited by 4, 4diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), the inhibitor of HCO(3)(-) transporter or by removal of extracellular Na(+). The extracellular pH measurement showed that the apical pH would increase when adding DIDS to the apical side of epididymal epithelial monolayer, however adding DIDS to the basolateral side had no effect on apical pH. The present study shows that sodium coupled bicarbonate influx regulates intracellular and apical pH in cultured caput epididymal epithelium.

  4. Sodium coupled bicarbonate influx regulates intracellular and apical pH in cultured rat caput epididymal epithelium.

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    Wu-Lin Zuo

    Full Text Available The epithelium lining the epididymis provides an optimal acidic fluid microenvironment in the epididymal tract that enable spermatozoa to complete the maturation process. The present study aims to investigate the functional role of Na(+/HCO(3(- cotransporter in the pH regulation in rat epididymis.Immunofluorescence staining of pan cytokeratin in the primary culture of rat caput epididymal epithelium showed that the system was a suitable model for investigating the function of epididymal epithelium. Intracellular and apical pH were measured using the fluorescent pH sensitive probe carboxy-seminaphthorhodafluor-4F acetoxymethyl ester (SNARF-4F and sparklet pH electrode respectively to explore the functional role of rat epididymal epithelium. In the HEPES buffered Krebs-Henseleit (KH solution, the intracellular pH (pHi recovery from NH(4Cl induced acidification in the cultured caput epididymal epithelium was completely inhibited by amiloride, the inhibitor of Na(+/H(+ exchanger (NHE. Immediately changing of the KH solution from HEPES buffered to HCO(3(- buffered would cause another pHi recovery. The pHi recovery in HCO(3(- buffered KH solution was inhibited by 4, 4diisothiocyanatostilbene-2,2-disulfonic acid (DIDS, the inhibitor of HCO(3(- transporter or by removal of extracellular Na(+. The extracellular pH measurement showed that the apical pH would increase when adding DIDS to the apical side of epididymal epithelial monolayer, however adding DIDS to the basolateral side had no effect on apical pH.The present study shows that sodium coupled bicarbonate influx regulates intracellular and apical pH in cultured caput epididymal epithelium.

  5. Evaluation of epididymal function through specific protein on spermatozoa.

    Science.gov (United States)

    Del Río, A G; De Sánchez, L Z; Sirena, A

    1984-01-01

    Investigations were focused on the characterization of specific epididymal proteins on the human spermatozoa as a representative parameter for epididymal function. An easy and attainable method, suitable for investigators and clinical use, is proposed in this article.

  6. Low-density Lipoprotein Improves Motility and Plasma Membrane Integrity of Cryopreserved Canine Epididymal Spermatozoa.

    Science.gov (United States)

    Prapaiwan, N; Tharasanit, T; Punjachaipornpol, S; Yamtang, D; Roongsitthichai, A; Moonarmart, W; Kaeoket, K; Manee-In, S

    2016-05-01

    Cryopreservation of caudal epididymal spermatozoa is an effective technique to conserve genetic potentials of superior dogs when it is not possible to collect ejaculated spermatozoa. Although hen egg yolk is commonly supplemented into the semen extender, active substances within the egg yolk which protect sperm against cryoinjury remain to be discovered. Among its compositions, low-density lipoprotein (LDL) has been reported to have a cryoprotective property for sperm cryopreservation. However, the effects of LDL on dog epididymal spermatozoa during cryopreservation have not yet been investigated. This study aimed to investigate the effects of LDL on epididymal spermatozoa quality following cryopreservation and thawing. After routine castration of 12 dogs, caudal epididymides from individuals were separated from the testes and cut into a few pieces in a Tris-buffer. Spermatozoa recovered from each sample were examined at once for sperm quality and divided into six groups of extender: no LDL, 20% egg yolk, 4%, 8%, 16%, and 24% LDL, before cryopreservation. The sperm aliquots were then equilibrated and conventionally frozen. After thawing, sperm motility, morphology, plasma membrane integrity, and acrosome integrity were evaluated. The results revealed that 4% LDL and 20% egg yolk yielded significantly higher sperm motility (57.69% and 52.69%, respectively, p<0.05) than other LDLs. In addition, 4% LDL yielded the significantly highest plasma membrane integrity (70.54%, p<0.05). In conclusion, the supplementation of 4% LDL in Tris-glucose extender could be applied for cryopreservation of canine epididymal spermatozoa.

  7. Low-density Lipoprotein Improves Motility and Plasma Membrane Integrity of Cryopreserved Canine Epididymal Spermatozoa

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    N. Prapaiwan

    2016-05-01

    Full Text Available Cryopreservation of caudal epididymal spermatozoa is an effective technique to conserve genetic potentials of superior dogs when it is not possible to collect ejaculated spermatozoa. Although hen egg yolk is commonly supplemented into the semen extender, active substances within the egg yolk which protect sperm against cryoinjury remain to be discovered. Among its compositions, low-density lipoprotein (LDL has been reported to have a cryoprotective property for sperm cryopreservation. However, the effects of LDL on dog epididymal spermatozoa during cryopreservation have not yet been investigated. This study aimed to investigate the effects of LDL on epididymal spermatozoa quality following cryopreservation and thawing. After routine castration of 12 dogs, caudal epididymides from individuals were separated from the testes and cut into a few pieces in a Tris-buffer. Spermatozoa recovered from each sample were examined at once for sperm quality and divided into six groups of extender: no LDL, 20% egg yolk, 4%, 8%, 16%, and 24% LDL, before cryopreservation. The sperm aliquots were then equilibrated and conventionally frozen. After thawing, sperm motility, morphology, plasma membrane integrity, and acrosome integrity were evaluated. The results revealed that 4% LDL and 20% egg yolk yielded significantly higher sperm motility (57.69% and 52.69%, respectively, p<0.05 than other LDLs. In addition, 4% LDL yielded the significantly highest plasma membrane integrity (70.54%, p<0.05. In conclusion, the supplementation of 4% LDL in Tris-glucose extender could be applied for cryopreservation of canine epididymal spermatozoa.

  8. Role of WNT signaling in epididymal sperm maturation.

    Science.gov (United States)

    Cheng, Jin-Mei; Tang, Ji-Xin; Li, Jian; Wang, Yu-Qian; Wang, Xiu-Xia; Zhang, Yan; Chen, Su-Ren; Liu, Yi-Xun

    2018-02-01

    Spermatozoa maturation, a process required for spermatozoa to acquire progressive motility and the ability to fertilize ova, primarily occurs in the caput and corpus of the epididymis. Despite considerable efforts, the factor(s) promoting epididymal sperm maturation remains unclear. Recently, WNT signaling has been implicated in epididymal sperm maturation. To further investigate WNT signaling function in epididymal sperm maturation, we generated Wntless conditional knockout mice (Wls cKO), Wls flox/flox ; Lcn5-Cre. In these mice, WNTLESS (WLS), a conserved membrane protein required for all WNT protein secretion, was specifically disrupted in the principal cells of the caput epididymidis. Immunoblot analysis showed that WLS was significantly reduced in the caput epididymidis of Wls cKO mice. In the caput epididymidis of Wls cKO mice, WNT 10A and WNT 2b, which are typically secreted by the principal cells of the caput epididymis, were not secreted. Interestingly, sperm motility analysis showed that the WLS deficiency in the caput epididymidis had no effect on sperm motility. Moreover, fertility tests showed that Wls cKO male mice had normal fertility. These results indicate that the disruption of WLS in principal cells of the caput epididymidis inhibits WNT protein secretion but has no effect on sperm motility and male fertility, suggesting that WNT signaling in the caput epididymidis may be dispensable for epididymal sperm maturation in mice.

  9. Low-density Lipoprotein Improves Motility and Plasma Membrane Integrity of Cryopreserved Canine Epididymal Spermatozoa

    OpenAIRE

    N. Prapaiwan; T. Tharasanit; S. Punjachaipornpol; D. Yamtang; A. Roongsitthichai; W. Moonarmart; K. Kaeoket; S. Manee-in

    2016-01-01

    Cryopreservation of caudal epididymal spermatozoa is an effective technique to conserve genetic potentials of superior dogs when it is not possible to collect ejaculated spermatozoa. Although hen egg yolk is commonly supplemented into the semen extender, active substances within the egg yolk which protect sperm against cryoinjury remain to be discovered. Among its compositions, low-density lipoprotein (LDL) has been reported to have a cryoprotective property for sperm cryopreservation. Howeve...

  10. Acetylcarnitine metabolism and the partial purification and characterization of an acetylcarnitine hydrolase from bovine caudal epididymal spermatozoa

    International Nuclear Information System (INIS)

    Bruns, K.A.

    1987-01-01

    Epididymal spermatozoa are capable of utilizing extracellular substrates for energy, but carbohydrates and free or esterified fatty acids are present in only very low concentrations in epididymal fluid. Acetyl-L-carnitine has been identified in epididymal fluid in low mM concentrations in several mammalian species and could possibly be an energy substrate for epididymal spermatozoa. Evidence that extracellular acetyl-L-carnitine can be used by intact caudal epididymal spermatozoa for energy, and a model for the metabolism of acetyl-L-carnitine by epididymal spermatozoa are presented here. Intact bovine and hamster caudal epididymal spermatozoa oxidized [1- 14 C] acetyl-L-carnitine to 14 CO 2 in a time-, cell number-, and substrate concentration-dependent manner. No concomitant uptake of acetyl-D,L-[N-methyl- 3 H] carnitine was observed by cells from the same preparations. Half-maximal rates of oxidation were observed at 8 mM and 4.5 mM acetyl-L-carnitine for the two species, respectively; the rates of oxidation at these concentrations were 15.3 nmol/10 8 cells·h and 2.9 nmol/10 7 cells·h. Intact spermatozoa in incubation with [ 3 H] acetyl-L-carnitine were observed to produce [ 3 H] acetate in the medium, and addition of sodium acetate competed for the uptake of radioactive acetate by these cells

  11. Laboratory handling of epididymal and testicular spermatozoa: What can be done to improve sperm injections outcome.

    Science.gov (United States)

    Esteves, Sandro C; Varghese, Alex C

    2012-09-01

    Spermatozoa from azoospermic males can be retrieved from either the epididymis or the testis, depending on the type of azoospermia, using different surgical methods such as percutaneous epididymal sperm aspiration (PESA), testicular sperm aspiration (TESA), testicular sperm extraction (TESE), and microsurgical testicular sperm extraction (micro- TESE). After collecting the epididymal fluid or testicular tissue, laboratory techniques are used to remove contaminants, cellular debris, noxious microorganisms, and red blood cells. Processed spermatozoa may be used for intracytoplasmic sperm injection or eventually be cryopreserved. However, spermatozoa collected from either the epididymis or the testis are often compromised and more fragile than ejaculated ones. Therefore, sperm processing techniques should be used with great caution to avoid jeopardizing the sperm fertilizing potential in treatment cycles. In this review, we describe the current methods for processing surgically-retrieved specimens, either fresh or frozen- thawed, and provide the tips and pitfalls for facilitating the handling of such specimens. In addition, we present the available laboratory tools to aid in the identification of viable immotile spermatozoa to be used in conjunction with assisted reproductive techniques. Review of the literature was carried out using PubMed and Science Direct search engines.

  12. Laboratory handling of epididymal and testicular spermatozoa: What can be done to improve sperm injections outcome

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    Sandro C Esteves

    2012-01-01

    Full Text Available Spermatozoa from azoospermic males can be retrieved from either the epididymis or the testis, depending on the type of azoospermia, using different surgical methods such as percutaneous epididymal sperm aspiration (PESA, testicular sperm aspiration (TESA, testicular sperm extraction (TESE, and microsurgical testicular sperm extraction (micro- TESE. After collecting the epididymal fluid or testicular tissue, laboratory techniques are used to remove contaminants, cellular debris, noxious microorganisms, and red blood cells. Processed spermatozoa may be used for intracytoplasmic sperm injection or eventually be cryopreserved. However, spermatozoa collected from either the epididymis or the testis are often compromised and more fragile than ejaculated ones. Therefore, sperm processing techniques should be used with great caution to avoid jeopardizing the sperm fertilizing potential in treatment cycles. In this review, we describe the current methods for processing surgically-retrieved specimens, either fresh or frozen- thawed, and provide the tips and pitfalls for facilitating the handling of such specimens. In addition, we present the available laboratory tools to aid in the identification of viable immotile spermatozoa to be used in conjunction with assisted reproductive techniques. Review of the literature was carried out using PubMed and Science Direct search engines.

  13. Epididymitis

    Science.gov (United States)

    ... infection, including sexually transmitted infections (STIs), such as gonorrhea or chlamydia. Sometimes, a testicle also becomes inflamed — ... you urinate. Causes Causes of epididymitis include: STIs. Gonorrhea and chlamydia are the most common causes of ...

  14. On the proluminal movement of 3H-androgens across the rat epididymal epithelium

    International Nuclear Information System (INIS)

    Turner, T.T.; Jones, C.E.; Roddy, M.S.

    1989-01-01

    3H-Androgens in rat epididymal interstitium have previously been shown to move into the epididymal lumen against a concentration gradient. This is true especially in the caput epididymidis. The present investigation used the technique of in vivo epididymal perifusion and tubule micropuncture to demonstrate that the proluminal movement of 3H-androgens is subject to competitive inhibition (unlabeled testosterone in the perifusion fluid at 10 times and 100 times the concentration of 3H-testosterone significantly reduced proluminal movement of isotope) and is not energy-dependent (1 mM 2,4-dinitrophenol in perifusion fluid did not reduce the proluminal movement of isotope). Additionally, dry-mount autoradiography demonstrated high intraluminal concentrations of isotope relative to interstitial concentrations after caput tubule incubation in 3H-dihydrotestosterone (3H-DHT), and showed that the high intraluminal concentrations of isotope were not dependent on the presence of spermatozoa, i.e. proluminal movement of 3H-androgens was not due to binding to intraluminal spermatozoa. Isolation of caput epididymidal sperm on filters followed by 3H-DHT binding experiments also failed to demonstrate the presence of specific binding of this androgen to spermatozoa. Finally, it was confirmed that electrophoresed epididymal lumen fluid contains a single 3H-DHT binding peak that is at its highest concentration in the caput epididymal fluid. These data are consistent with the conclusion that intraluminal androgen-binding protein is an important factor in transepithelial androgen movement

  15. PERCENTAGE OF VIABLE SPERMATOZOA COLLECTED FROM THE EPIDIDYMES OF DEATH LOCAL DOG

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    I Nyoman Sulabda

    2012-11-01

    Full Text Available The purpose of this study to determine the effectof post mortem time on percentage of lifeepididymessperm from postmortem dog caudae epididymides. A total of 9 dog were usedand divided into three group. T0 was control group, T1, 3 hours postmortem and T2, 6hours postmortem. This way, samples were obtained at different times postmortem. Spermwere extracted from the caudae epididymes by means of cuts.The result showed that the percentage of life sperm were 67,16 ± 5.67(T0, 46.33 ± 5.60(T1 and 24.00 ± 4.35 respectively. We could appreciate that percentage of life wasaffected by postmortem time. There was significant decrease life sperm recovered fromepididymes postmortem (P<0.01. In conclusion, epididymes sperm from dog undergodecrease of percentage of life, but it could stay acceptable within many hours postmortem.We intepreted these data to indicate that it may still be possible to obtain viablespermatozoa many hours later.

  16. VIABILITY AND PLASMA MEMBRANE INTEGRITY OF THE SPOTTED BUFFALO EPIDIDYMAL SPERMATOZOA AFTER THAWING WITH THE ADDITION OF DEXTROSE INTO THE EXTENDER

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    M. RIZAL

    2009-01-01

    Full Text Available h e objective of this study was to obtain the viability and plasma membrane integrity of the spotted buff alo epididymal sperm after addition of dextrose into Andromed® extender. Spermatozoa that have been collected from cauda epididymis were diluted with Andromed® extender as control (K and Andromed® + 0.2% dextrose (P1 and Andromed® + 0.4% dextrose (P2 as treatments. h e results showed that the quality of epididymal spermatozoa decreased during cryopreservation process. h e percentage of motility after thawing in P1 (46% and P2 (46.67% were signifi cantly higher (P<0.05 compared to K (41% as well as the percentage of live sperm in P1 (58.8% and P2 (60% compared to K (52.2%. h e percentage of membrane integrity in P1, P2 and K were 67.4; 66.8 and 68 %, respectively. In conclusion, the addition of 0.2 and 0.4% of dextrose into Andromed® acted as an extra cellular cryoprotectant and could maintain the viability and membrane integrity of the spotted buff alo epididymal spermatozoa after thawing.

  17. Ultrastructural and hormonal changes in rat cauda epididymal spermatozoa induced by Boswellia papyrifera and Boswellia carterii.

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    Ahmed, Mukhtar; Ali, Daoud; Harrath, Abdel Halim; Hussain, Tajamul; Al-Daghri, Nasser; Alokail, Majed S; Aladakatti, Ravindranath H; Ghodesawar, Mukhtar Ahmed G

    2014-04-01

    Boswellia papyrifera and Boswellia carterii diffuses smoke polluting air that adversely affects indoor environment that certainly harm human health. Therefore, this study aims at ascertaining the effect of these plants on gonadal hormones and molecular changes in rat spermatozoa. The animals were exposed to 4 g/kg body weight of B. papyrifera and B. carterii daily for 120 days along with suitable controls. Significant decreases in FSH, LH and testosterone levels were evidenced, along with a reduction of protein, sialic acid, and carnitine levels. In sperm physiology, sperm count, motility, speed decrease, whereas sperm anomalies increase. TEM observation indicates morphological changes in plasma and acrosomal membranes, cytoplasmic droplet in the tail region, vacuolated, and disorganization of the mitochondrial sheath. These findings demonstrate that B. papyrifera and B. carterii smoke affects the process of sperm formation and maturation, which indicates the detrimental effects of these plants on the reproductive system. Copyright © 2014 Académie des sciences. Published by Elsevier SAS. All rights reserved.

  18. The expression of the new epididymal luminal protein of PDZ domain containing 1 is decreased in asthenozoospermia

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    A-Juan Liang

    2018-01-01

    Full Text Available Spermatozoa are not mature until they transit the epididymis where they acquire motility and the ability to fertilize an egg through sequential modifications. The epididymis has three functional regions, caput, corpus, and cauda, and the luminal proteins of the epididymis play important roles in the above modifications. However, the proteins with differential enrichment between the caput and cauda are still largely unknown. To reveal the functions of the caput and cauda during sperm maturation, luminal proteins from caput and cauda of mice were analyzed by isobaric tag for relative and absolute quantitation (iTRAQ. Overall, 128 differentially enriched proteins were found, of which 46 were caput enriched and 82 were cauda enriched. Bioinformatic analysis showed that lipid metabolism was active in the caput; while anion- and cation-binding activity and phosphorus and organophosphate metabolism were active in the cauda. A new epididymal luminal protein, the caput-enriched PDZ domain containing 1 (Pdzk1, also named Na+/H+ exchange regulatory cofactor 3 (NHERF3, which plays a critical role in cholesterol metabolism and carnitine transport, was found in the lipid metabolism. Western blotting and immunofluorescence analyses showed that Pdzk1 was expressed in the epididymis but not in the testis, and localized at the middle piece of the sperm tail. Pdzk1 protein level was also reduced in the spermatozoa in case of asthenozoospermic patients compared with that in normozoospermic men, suggesting that Pdzk1 may participate in sperm maturation regulation and may be associated with male infertility. These results may provide new insights into the mechanisms of sperm maturation and male infertility.

  19. Variations of motility and survival with storage time at 4°C of epididymal spermatozoa Ouled-Djellal breed rams in Eastern Algeria.

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    Safsaf, B; Belkadi, S; Belkacem, L; Mamache, B; Tlidjane, M

    2015-03-01

    The aim of this study was to evaluate some reproduction performances in Ouled-Djellal rams. This study involved genital organs removed after slaughter from 54 rams at the municipal slaughterhouse of Batna (East Algeria). The measurements of survival and mobility of epididymal sperm followed at 0, 24, 48 and 72 h after collection, revealed significant (p0.001) according to time. Thus, concerning the sperm motility the values were 91.00±2.40%, 89.20±2.40%, 77.00±6.20% and 62.60±1.20% at 0, 24, 48 and 72 h, respectively. Indeed, in live sperm, the viability rates were 82.15±1.48%, 77.67±1.74%, 66.56±1.95% and 52.30±1.46% at 0, 24, 48 and 72 h, respectively. This study revealed that epididymal spermatozoa stored at 04°C for 72 h kept their mobility and vitality at nearly a half of their the original parameters.

  20. Variations of motility and survival with storage time at 4°C of epididymal spermatozoa Ouled-Djellal breed rams in Eastern Algeria

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    B. Safsaf

    2015-03-01

    Full Text Available Aim: The aim of this study was to evaluate some reproduction performances in Ouled-Djellal rams. Materials and Methods: This study involved genital organs removed after slaughter from 54 rams at the municipal slaughterhouse of Batna (East Algeria. Results: The measurements of survival and mobility of epididymal sperm followed at 0, 24, 48 and 72 h after collection, revealed significant (p0.001 according to time. Thus, concerning the sperm motility the values were 91.00±2.40%, 89.20±2.40%, 77.00±6.20% and 62.60±1.20% at 0, 24, 48 and 72 h, respectively. Indeed, in live sperm, the viability rates were 82.15±1.48%, 77.67±1.74%, 66.56±1.95% and 52.30±1.46 at 0, 24, 48 and 72 h, respectively. Conclusion: This study revealed that epididymal spermatozoa stored at 04°C for 72 h kept their mobility and vitality at nearly a half of their the original parameters.

  1. Lactate dehydrogenase activity of rat epididymis and spermatozoa: Effect of constant light

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    RH Ponce

    2009-12-01

    Full Text Available During its passage through the epididymis, the gamete undergoes a process of “maturation” leading to the acquisition of its fertilizing ability. The epididymis displays regional variations in the morphology and metabolic properties of its epithelium which are relevant for the progressive development of mature sperm characteristics. The epididymis has spontaneous peristaltic contractions and receives sympathetic innervation that is modulated by melatonin, a hormone synthesized and released by the pineal gland. Constant lighting disrupts melatonin synthesis and secretion. We have studied the effect of constant light on lactate dehydrogenase (LDH; EC 1.1.1.27 and its isozyme C4 activities and protein content in whole epididymis, epididymal tissue and in spermatozoa from caput and cauda segments. Animals were exposed from birth to an illumination schedule of 14 h light: 10 h dark (group L:D. At 60 days of age one group of animals was submitted to constant light over 50 days (group L:L. In order to test the fertilizing ability, the rats of each group were mated with soliciting estrous females. The percentage of pregnancies in females mated with males maintained in L:L was remarkably lower than those in females mated with males maintained in the L:D photoperiod (44% and 88% respectively. Constant light increased protein concentration and LDH activity in caput as well as in cauda of total epididymis. On the contrary, in epididymal tissue, the protein content decreased in both epididymal sections compared with controls. When enzymatic activity was expressed in Units per spermatozoa, constant light induced a significant reduction of total LDH and LDHC4 in caput and cauda spermatozoa while LDH activity of epididymal tissue was not affected. In spite of the decrease in LDH per sperm cell when rats were exposed to constant light, in total epididymis (epididymis tissue plus sperm cells content and in spermatozoa, values of enzyme activities expressed per

  2. LCN6, a novel human epididymal lipocalin

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    Soundararajan Rama

    2003-11-01

    Full Text Available Abstract Background The lipocalin (LCN family of structurally conserved hydrophobic ligand binding proteins is represented in all major taxonomic groups from prokaryotes to primates. The importance of lipocalins in reproduction and the similarity to known epididymal lipocalins prompted us to characterize the novel human epididymal LCN6. Methods and Results LCN6 cDNA was identified by database analysis in a comprehensive human library sequencing program. Macaca mulatta (rhesus monkey cDNA was obtained from an epididymis cDNA library and is 93% homologous to the human. The gene is located on chromosome 9q34 adjacent LCN8 and LCN5. LCN6 amino acid sequence is most closely related to LCN5, but the LCN6 beta-barrel structure is best modeled on mouse major urinary protein 1, a pheromone binding protein. Northern blot analysis of RNAs isolated from 25 human tissues revealed predominant expression of a 1.0 kb mRNA in the epididymis. No other transcript was detected except for weak expression of a larger hybridizing mRNA in urinary bladder. Northern hybridization analysis of LCN6 mRNA expression in sham-operated, castrated and testosterone replaced rhesus monkeys suggests mRNA levels are little affected 6 days after castration. Immunohistochemical staining revealed that LCN6 protein is abundant in the caput epithelium and lumen. Immunofluorescent staining of human spermatozoa shows LCN6 located on the head and tail of spermatozoa with the highest concentration of LCN6 on the post-acrosomal region of the head, where it appeared aggregated into large patches. Conclusions LCN6 is a novel lipocalin closely related to Lcn5 and Lcn8 and these three genes are likely products of gene duplication events that predate rodent-primate divergence. Predominant expression in the epididymis and location on sperm surface are consistent with a role for LCN6 in male fertility.

  3. Down-regulation of CatSper1 channel in epididymal spermatozoa contributes to the pathogenesis of asthenozoospermia, whereas up-regulation of the channel by Sheng-Jing-San treatment improves the sperm motility of asthenozoospermia in rats.

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    Wang, Ya-Nan; Wang, Bo; Liang, Ming; Han, Cai-Yan; Zhang, Bin; Cai, Jie; Sun, Wei; Xing, Guo-Gang

    2013-02-01

    To determine the expression of CatSper1 channel in epididymal spermatozoa in a rat model of asthenozoospermia, induced by cyclophosphamide (CP), and further examine the effects of soluble granules of Sheng-Jing-San (SJS), a traditional Chinese medicine recipe, on CatSper1 expression and sperm motility in the CP-induced asthenozoospermic rats. Placebo-controlled, randomized trial. Neuroscience Research Institute, Peking University, China. Sexually mature male Sprague-Dawley rats (n = 60). In the CP group, CP at the dose of 35 mg/kg intraperitoneally injected into rats once a day for 7 days; in the normal saline (NS) group, 0.9% saline solution was injected as control. Sperm motility and count were evaluated by computer-assisted sperm assay (CASA); protein and mRNA expression of CatSper1 channel in epididymal spermatozoa was determined by Western blotting and quantitative real-time RT-PCR, respectively. The rats were randomly divided into five groups with 12 rats in each group: CP, normal saline (NS), CP + SJS, CP + NS, and treatment naïve. In the CP + SJS group, after the last injection of CP, SJS at a dose of 30 mg/kg was intragastrically administrated to rats once a day for 14 days; in CP + NS group, saline solution instead of SJS was administrated as control. In the treatment naïve group, rats were normally fed for 21 days as controls. We found a statistically significant reduction of the CatSper1 channel, which is associated with an impairment of sperm motility in the epididymal spermatozoa of CP-induced asthenozoospermic rats. Soluble granules of SJS could dramatically restore the CP-induced down-regulation of CatSper1 in epididymal spermatozoa, which greatly improved the sperm motility in the asthenozoospermic rats. Down-regulation of the CatSper1 channel in epididymal spermatozoa likely contributes to the pathogenesis of asthenozoospermia, whereas up-regulation of the channel by SJS improves sperm motility and thus can be used as an effective therapeutic

  4. Study of the role of epididymal alpha-glucosidase in the fertility of male rats by the administration of the enzyme inhibitor castanospermine.

    Science.gov (United States)

    Yeung, C H; Cooper, T G

    1994-11-01

    The activity of epididymal alpha-glucosidase in adult rats was rapidly suppressed to histochemically undetectable levels within 2 days by the continuous release of the enzyme inhibitor castanospermine via a peritoneal osmotic pump at a rate of 100-200 nmol h-1. It was established that mating activities overnight depleted 72% of the spermatozoa in the distal cauda, which was replenished in 2 days, and that fertility began to decline 3 weeks after efferent duct ligation. Male rats of proven mating proficiency and fertility were treated with castanospermine, or buffered saline as control, for up to 30 days and enzyme inhibition was confirmed at the end of treatment by histochemistry. Fertility was normal at the first mating test on day 7, significantly decreased at the second mating on day 9, but recovered in a stepwise manner at subsequent matings on days 12 and 14. Delaying the third mating until day 25 did not sustain the transient subfertility. However, prolonging sperm storage in the distal cauda epididymides and preventing replenishment with freshly matured spermatozoa, by efferent duct ligation for 14 days performed on day 15 during castanospermine administration, caused a decrease in fertility and a change in the kinematics of epididymal spermatozoa of the castanospermine-treated group. In control rats, binding of epididymal spermatozoa to Vicia faba, a lectin specific for glucose and glucosamine, and mannose and mannosamine residues, decreased from the proximal caput to the distal corpus coincident with the increase in alpha-glucosidase activity on the epithelial brush border. Lectin binding then increased in the cauda where enzyme activity was absent. However, castanospermine treatment did not significantly alter this binding profile. The findings suggest that epididymal alpha-glucosidase does not play a crucial role in the development of sperm fertilizing capacity, but may be involved in the preparation of spermatozoa for storage.

  5. Effects of exposure of epididymal boar spermatozoa to seminal plasma on the binding of zona pellucida proteins during in vitro capacitation

    NARCIS (Netherlands)

    Harkema, W.; Colenbrander, B.; Engel, B.; Woelders, H.

    2004-01-01

    The purpose of the investigation was to determine whether seminal plasma plays a role in the increase during in vitro capacitation of the number of boar spermatozoa with enhanced binding of zona pellucida proteins. Ejaculated spermatozoa and spermatozoa collected from the caudae epididymides of

  6. CT Demonstration of Caput Medusae

    Science.gov (United States)

    Weber, Edward C.; Vilensky, Joel A.

    2009-01-01

    Maximum intensity and volume rendered CT displays of caput medusae are provided to demonstrate both the anatomy and physiology of this portosystemic shunt associated with portal hypertension. (Contains 2 figures.)

  7. Novel phenotype of mouse spermatozoa following deletion of nine β-defensin genes.

    Science.gov (United States)

    Dorin, Julia R

    2015-01-01

    β-defensin peptides are a large family of antimicrobial peptides. Although they kill microbes in vitro and interact with immune cells, the precise role of these genes in vivo remains uncertain. Despite their inducible presence at mucosal surfaces, their main site of expression is the epididymis. Recent evidence suggests that a major function of these peptides is in sperm maturation. In addition to previous work suggesting this, work at the MRC Human Genetics Unit, Edinburgh, has shown that homozygous deletion of a cluster of nine β-defensin genes in the mouse results in profound male sterility. The spermatozoa derived from the mutants had reduced motility and increased fragility. Epididymal spermatozoa isolated from the cauda region of the homozygous mutants demonstrated precocious capacitation and increased spontaneous acrosome reactions compared with those from wild-types. Despite this, these mutant spermatozoa had reduced ability to bind to the zona pellucida of oocytes. Ultrastructural examination revealed a disintegration of the microtubule structure of mutant-derived spermatozoa isolated from the epididymal cauda region, but not from the caput. Consistent with premature acrosome reaction and hyperactivation, spermatozoa from mutant animals had significantly increased intracellular calcium content. This work demonstrates that in vivo β-defensins are essential for successful sperm maturation, and that their disruption alters intracellular calcium levels, which most likely leads to premature activation and spontaneous acrosome reactions that result in hyperactivation and loss of microtubule structure of the axoneme. Determining which of the nine genes are responsible for the phenotype and the relevance to human sperm function is important for future work on male infertility.

  8. Novel phenotype of mouse spermatozoa following deletion of nine β-defensin genes

    Directory of Open Access Journals (Sweden)

    Julia R Dorin

    2015-01-01

    Full Text Available β-defensin peptides are a large family of antimicrobial peptides. Although they kill microbes in vitro and interact with immune cells, the precise role of these genes in vivo remains uncertain. Despite their inducible presence at mucosal surfaces, their main site of expression is the epididymis. Recent evidence suggests that a major function of these peptides is in sperm maturation. In addition to previous work suggesting this, work at the MRC Human Genetics Unit, Edinburgh, has shown that homozygous deletion of a cluster of nine β-defensin genes in the mouse results in profound male sterility. The spermatozoa derived from the mutants had reduced motility and increased fragility. Epididymal spermatozoa isolated from the cauda region of the homozygous mutants demonstrated precocious capacitation and increased spontaneous acrosome reactions compared with those from wild-types. Despite this, these mutant spermatozoa had reduced ability to bind to the zona pellucida of oocytes. Ultrastructural examination revealed a disintegration of the microtubule structure of mutant-derived spermatozoa isolated from the epididymal cauda region, but not from the caput. Consistent with premature acrosome reaction and hyperactivation, spermatozoa from mutant animals had significantly increased intracellular calcium content. This work demonstrates that in vivo β-defensins are essential for successful sperm maturation, and that their disruption alters intracellular calcium levels, which most likely leads to premature activation and spontaneous acrosome reactions that result in hyperactivation and loss of microtubule structure of the axoneme. Determining which of the nine genes are responsible for the phenotype and the relevance to human sperm function is important for future work on male infertility.

  9. Pentoxifylline effects on capacitation and fertility of stallion epididymal sperm

    NARCIS (Netherlands)

    Guasti, P. N.; Monteiro, G. A.; Maziero, R. R.D.; Carmo, M. T.; Dell'Aqua, J. A.; Crespilho, A. M.; Rifai, E. A.; Papa, F. O.

    2017-01-01

    The aims of this study were to determinate whether pentoxifylline (PTX) increases the motion parameters of fresh and frozen-thawed equine epididymal spermatozoa, to evaluate the tyrosine phosphorylation of frozen-thawed epididymal sperm in the presence of PTX and to determine whether the

  10. Epididymal research: more warp than weft?

    Directory of Open Access Journals (Sweden)

    Trevor G Cooper

    2015-01-01

    Full Text Available From a review of some aspects of epididymal structure, function and research done largely in my research area over the last 50 years, I conclude that more is known than is understood of sperm maturation and storage in the epididymis. Highly qualified technicians have not always applied sophisticated modern techniques in well-considered experiments to physiologically relevant and properly-prepared samples, so that our understanding of the biological problem of the nature of the epididymal epithelial influence on maturing epididymal spermatozoa has not kept pace with the outpouring of data generated, much of which is difficult to interpret. We stand at a crossroads of where to aim our limited resources and personnel: should we continue new technology-led studies in many directions, backtrack to test hypotheses and fill in gaps in our knowledge, or consider more biological directions to our research?

  11. Immunohistochemical localization of androgen receptor in rat caput epididymis during postnatal development

    Directory of Open Access Journals (Sweden)

    Sema Timurkaan

    2011-09-01

    Full Text Available Objectives: The aim of this study was to investigate the developmental pattern of androgen receptor (AR in caput epididymis.Materials and methods: In this study three randomly selected rats were sacrificed at ages 21, 56, 90 and 120 days old. All male rats were anesthetized with ethyl ether before killing. Then, the caput epididymides were removed and fixed in Bouin’s fixative at +4°C for 36 hour. Afterwards the tissue samples were embedded in paraffin for routine histological methods. Later the tissues were sectioned at 5μm and mounted on poly-L-lysin-coated slides. To solve the antigen masking problem, we performed microwave stimulated antigen retrieval technique before the immunohistochemical staining. Avidin-Biotin-Peroxidase Complex (ABC method was applied for immunohistochemical staining.Results: In all age groups of rats studied, positive immunohistochemical staining for the AR appeared in nuclei of epididymal cells. The staining intensity of AR positive cells did not change depending on age. In caput epididymis, immunostainable AR was found in tubular epithelial cells (principal cells, basal cells and apical cells and peritubular smooth muscle cells. The AR staining in the epithelial cells appeared to be stronger than in the peritubular smooth muscle cells. In the epithelial cells; staining intensity was stronger in principal cells than in basal cells and apical cells.Conclusion: Staining intensity of AR positive epididymal cells irrespective of age indicated the necessity of androgens for postnatal differentiation and maintaining the structure of the epididymis. Stronger staining intensity in principal cells suggested that principal cells are more sensitive to androgen stimulation. J Clin Exp Invest 2011; 2 (3: 260-266.

  12. Ethane dimethanesulfonate (EDS) perturbs epididymal epithelial cell function in vitro

    International Nuclear Information System (INIS)

    Klinefelter, G.

    1990-01-01

    The formation of sperm granulomas in the epididymis following exposure to EDS, a Leydig cell toxicant, was reported by Cooper and Jackson in 1970. Recent work suggests that EDS may effect the epididymis directly. An in vitro system was developed to determine the nature of any direct effect. The caput epididymis from adult rats was dissected free of connective tissue and small pieces of the tissue were enzymatically digested until plaques of epididymal epithelial cells were obtained. Plaques were cultured on an extracellular matrix gelled on top of a semipermeable filter creating dual-compartment environments. The epithelial cells maintained typical morphology and protein secretion in this culture system for several days. Beginning on day 3, EDS (1 mM) was added to the basal compartment, with or without 35 S-methionine. After 24 hours, 35 S-labelled culture medium was taken from the apical compartment and analyzed by SDS-PAGE and fluorography. EDS caused decreased secretion of several proteins, including a 39 Kd molecule. Interestingly, a 39 Kd protein was also shown to disappear from sperm taken from the caput epididymidis following in vivo exposure to EDS. Unlabelled cultures were fixed and processed for light microscopy. No alterations in morphological integrity were observed. Thus, epididymal epithelial cell function is directly altered by EDS exposure

  13. Epididymal lithiasis in Gallus gallus domesticus

    Directory of Open Access Journals (Sweden)

    JM Rocha Jr.

    2009-03-01

    Full Text Available A study on the occurrence of epididymal lithiasis (EL in Gallus gallus domesticus of different breeds and from different geographical regions of Brazil was carried out. Forty breeding roosters were collected in the states Santa Catarina, São Paulo, Minas Gerais, Espírito Santo and Goiás, representing two specific pathogen free Leghorn lines (SPF, one commercial Leghorn layer breeder (Hy-Line line, two commercial broiler breeder lines (Ross and Cobb, and two backyard chicken flocks. In commercial poultry breeders, EL was observed mainly after 55 weeks of age, and lithus resulted in lower fertility caused by inadequate maturation of spermatozoa and testicular atrophy, which evolved to more severe testicular dysfunction at 100 weeks of age. One hundred percent of the sampled roosters of the evaluated SPF lines presented epididymal lithiasis, which was also observed in all commercial genetic lines. The analysis of EL occurrence revealed no differences among regions and breeds: all regions and genetic lines presented epididymal calculi and the occurrence seemed higher in the more intensively-reared chickens. As for the backyard roosters, 50% presented calculi. Considering the occurrence of EL in the SPF flocks, its primary association with infectious bronchitis virus (IBV infection was discarded, despite its contribution to fertility loss.

  14. Epididymal expression of the forkhead transcription factor Foxi1 is required for male fertility

    OpenAIRE

    Blomqvist, Sandra Rodrigo; Vidarsson, Hilmar; Söder, Olle; Enerbäck, Sven

    2006-01-01

    An essential aspect of male reproductive capacity is the immediate availability of fertilization-ready spermatozoa. To ensure this, most mammals rely on post-testicular sperm maturation. In epididymis, germ cells are matured and stored in a quiescent state that readily can be altered to produce active spermatozoa. This depends on active proton secretion into the epididymal lumen. We have identified Foxi1 as an important regulator of gene expression in narrow and clear cells—the major proton s...

  15. Blood epididymal barrier to [3H]-inulin in intact and vasectomized hamsters

    International Nuclear Information System (INIS)

    Turner, T.T.; D'Addario, D.A.; Howards, S.S.

    1981-01-01

    The net transport of [ 3 H]-inulin into the fluids of the hamster seminiferous and caput, corpus, and cauda epididymal tubules was examined in both intact animals and those vasectomized 10 months previously. Mean isotope concentrations in reproductive tract tubule fluids did not exceeded 10 per cent of blood plasma isotope concentrations during the experiment. There were no significant differences in net transport of [ 3 H]-inulin into any of the tubule fluids sampled. Ten months after vasectomy, the seminiferous tubule, and all regions of the epididymal tubule retain the capacity to exclude [ 3 H]-insulin. Thus in the hamster 10 months after vasectomy, the blood testis and blood epididymal barriers to inulin are intact

  16. Influence of rete testis fluid deprivation on the kinetic parameters of goat epididymal 5 alpha-reductase.

    Science.gov (United States)

    Kelce, W R; Lubis, A M; Braun, W F; Youngquist, R S; Ganjam, V K

    1990-01-01

    A surgical technique to cannulate the rete testis of the goat was utilized to examine the effects of rete testis fluid (RTF) deprivation on the enzymatic activity of epididymal 5 alpha-reductase. Kinetic techniques were used to determine whether the regional enzymatic effect of RTF deprivation is to decrease the apparent number of 5 alpha-reductase active sites or the catalytic activity of each active site within the epididymal epithelium. Paired comparisons of (Vmax)app and (Km)app values between control and RTF-deprived epididymides indicated that RTF deprivation affected the value of (Vmax)app with no apparent change in the values of (Km)app in caput, corpus, and cauda epididymal regions. We conclude that RTF deprivation in the goat epididymis for 7 days results in a decreased number of apparent 5 alpha-reductase active sites within the epididymal epithelium.

  17. The Immunoexpression of FSH-R in the Ductuli Efferentes and the Epididymis of Men and Rat: Effect of FSH on the Morphology and Steroidogenic Activity of Rat Epididymal Epithelial Cells In Vitro

    Directory of Open Access Journals (Sweden)

    Małgorzata Świder-Al-Amawi

    2010-01-01

    Full Text Available The Sertoli cells were regarded as the only target for FSH in male reproductive system. The expression of FSH receptor (FSH-R was detected also in epithelial cells of the caput epididymis of rat and monkey. We showed in the immunohistochemistry study the expression of FSH-R in rat and human ductuli efferentes and the caput, corpus, and cauda epididymis, moreover, by Western blot analysis in the caput and cauda epididymis of rat. Additionally, we presented that the morphology of rat epididymal epithelial cells in vitro was affected by FSH, and FSH stimulation resulted in the increase of 17β-estradiol synthesis by rat caput epididymal cells in dose-depended manner. In conclusion, the identification of FSH receptors in human and rat epididymides supports our results that the epididymis is a target organ not only for LH but additionally for FSH. On the basis of the results we showed for the first time that morphology of epididymal epithelial cells and epididymal steroidogenesis can be regulated by FSH.

  18. Caput medusae in alcoholic liver disease | Hari Kumar | Nigerian ...

    African Journals Online (AJOL)

    Caput medusae and palmar erythema are cardinal signs in cirrhosis of liver with portal hypertension. Palmar erythema is described more often as a marker for alcoholic etiology of chronic liver disease. The peripheral stigmata of chronic liver disease are not routinely seen now a days due to early diagnosis and better ...

  19. Histomorphological changes by epididymal lithiasis in roosters

    Directory of Open Access Journals (Sweden)

    I Geraldo

    2014-03-01

    Full Text Available Epididymal lithiasis (EL histopathology is described using light and electronic microscopy in roosters (Gallus gallus domesticus naturally affected by EL in Minas Gerais, Brazil. The histologic and morphological changes by EL in roosters was performed regarding cellular and subcellular details through light and electron microscopy. Efferent ductules epithelium lysosomal increase in size and numbers, membrane rupture, cellular vacuolation, ciliary loss, basal membrane degeneration, inflammatory reaction with mononuclear infiltrations, edema, epithelial and vascular endothelium losses were described. All industrial and freerange chickens showed EL in varying degrees in the efferent ductules (ED. However, ED altered areas did not correlate with the presence of luminal stones. Non-ciliated ED epithelium cells presented several atypically large lysosomes. Plicae loss and basal vacuoles were observed in the epithelium of dilated regions. Cellular cilia loss and apical cytoplasmic membrane rupture resulted in leakage of the cytoplasmic contents to the ED lumen, and ED epithelium desquamation occurred with or without lesion to the basal membrane. Basal membrane alterations were associated with profound sub-epithelial connective tissue damage. Aggregations of desquamated epithelium and spermatozoa were seen in the lumen of ED and compact aggregates were considered the basis for calculi formation. The widespread occurrence and high severity of EL lesions are indicative of the importance of EL as a cause of infertility in male chickens.

  20. Sclerotherapy for hydrocoele and epididymal cysts.

    Science.gov (United States)

    Nash, J R

    1979-04-01

    A prospective study was carried out on the efficacy of sclerotherapy for the treatment of hydrocoeles and epididymal cysts. Thirty-six hydrocoeles and 13 epididymal cysts were treated and followed up for between 1 and 2 years. Thirty-four hydrocoeles were cured, 1 failed to respond to treatment and 1 recurred after treatment. All 13 epididymal cysts were cured.

  1. Effect of nonylphenol on male reproduction: Analysis of rat epididymal biochemical markers and antioxidant defense enzymes

    Energy Technology Data Exchange (ETDEWEB)

    Aly, Hamdy A.A., E-mail: hamdyaali@yahoo.com [Department of Pharmacology and Toxicology, Faculty of Pharmacy, King Abdulaziz University, Jeddah (Saudi Arabia); Domènech, Òscar [Department of Physical Chemistry, Faculty of Pharmacy, Barcelona University (Spain); Banjar, Zainy M. [Department of Medical Biology, School of Medicine, King Abdulaziz University, Jeddah (Saudi Arabia)

    2012-06-01

    The mechanism by which nonylphenol (NP) interferes with male reproduction is not fully elucidated. Therefore, the present study was conducted to evaluate the effect of NP on male reproductive organ's weight, sperm characteristics, and to elucidate the nature and mechanism of action of NP on the epididymis. Adult male Wistar rats were gavaged with NP, dissolved in corn oil, at 0, 100, 200 or 300 mg/kg/day for 30 consecutive days. Control rats were gavaged with vehicle (corn oil) alone. Body weight did not show any significant change while, absolute testes and epididymides weights were significantly decreased. Sperm count in cauda and caput/corpus epididymides, and sperm motility was significantly decreased. Daily sperm production was significantly decreased in a dose-related manner. Sperm transit time in cauda epididymis was significantly decreased by 300 mg/kg, while in the caput/corpus epididymis it was significantly decreased by 200 and 300 mg/kg of NP. Plasma LDH was significantly increased while; plasma testosterone was significantly decreased in a dose-related pattern. In the epididymal sperm, NP decreased acrosome integrity, Δψm and 5′-nucleotidase activity. Hydrogen peroxide (H{sub 2}O{sub 2}) production and LPO were significantly increased in a dose-related pattern. The activities of SOD, CAT and GPx were significantly decreased in the epididymal sperm. In conclusion, this study revealed that NP treatment impairs spermatogenesis and has a cytotoxic effect on epididymal sperm. It disrupts the prooxidant and antioxidant balance. This leads oxidative stress in epididymal sperms of rat. Moreover, the reduction in sperm transit time may affect sperm quality and fertility potential. -- Highlights: ► The nature and mechanism of action of NP on rat epididymis were elucidated. ► NP decreased sperm count, motility, daily sperm production and sperm transit time. ► NP decreased sperm acrosome integrity, Δψm and 5′-nucleotidase activity. ► Plasma

  2. Effect of nonylphenol on male reproduction: Analysis of rat epididymal biochemical markers and antioxidant defense enzymes

    International Nuclear Information System (INIS)

    Aly, Hamdy A.A.; Domènech, Òscar; Banjar, Zainy M.

    2012-01-01

    The mechanism by which nonylphenol (NP) interferes with male reproduction is not fully elucidated. Therefore, the present study was conducted to evaluate the effect of NP on male reproductive organ's weight, sperm characteristics, and to elucidate the nature and mechanism of action of NP on the epididymis. Adult male Wistar rats were gavaged with NP, dissolved in corn oil, at 0, 100, 200 or 300 mg/kg/day for 30 consecutive days. Control rats were gavaged with vehicle (corn oil) alone. Body weight did not show any significant change while, absolute testes and epididymides weights were significantly decreased. Sperm count in cauda and caput/corpus epididymides, and sperm motility was significantly decreased. Daily sperm production was significantly decreased in a dose-related manner. Sperm transit time in cauda epididymis was significantly decreased by 300 mg/kg, while in the caput/corpus epididymis it was significantly decreased by 200 and 300 mg/kg of NP. Plasma LDH was significantly increased while; plasma testosterone was significantly decreased in a dose-related pattern. In the epididymal sperm, NP decreased acrosome integrity, Δψm and 5′-nucleotidase activity. Hydrogen peroxide (H 2 O 2 ) production and LPO were significantly increased in a dose-related pattern. The activities of SOD, CAT and GPx were significantly decreased in the epididymal sperm. In conclusion, this study revealed that NP treatment impairs spermatogenesis and has a cytotoxic effect on epididymal sperm. It disrupts the prooxidant and antioxidant balance. This leads oxidative stress in epididymal sperms of rat. Moreover, the reduction in sperm transit time may affect sperm quality and fertility potential. -- Highlights: ► The nature and mechanism of action of NP on rat epididymis were elucidated. ► NP decreased sperm count, motility, daily sperm production and sperm transit time. ► NP decreased sperm acrosome integrity, Δψm and 5′-nucleotidase activity. ► Plasma LDH was

  3. Identification of New Epididymal Luminal Fluid Proteins Involved in Sperm Maturation in Infertile Rats Treated by Dutasteride Using iTRAQ

    Directory of Open Access Journals (Sweden)

    Shu-Wu Xie

    2016-05-01

    Full Text Available Background: Spermatozoa become mature and acquire fertilizing capacity during their passage through the epididymal lumen. In this study, we identified new epididymal luminal fluid proteins involved in sperm maturation in infertile rats by dutasteride, a dual 5α-reductase inhibitor, in order to provide potential epididymal targets for new contraceptives and infertility treatment. Methods: Male rats were treated with dutasteride for 28 consecutive days. We observed the protein expression profiles in the epididymal luminal fluids in infertile and normal rats using isobaric tags for relative and absolute quantitation (iTRAQ technique. The confidence of proteome data was validated by enzyme-linked immunosorbent assays. Results: 1045 proteins were tested, and 23 of them presented different expression profiling in the infertile and normal rats. The seven proteins were down-regulated, and 16 proteins were up-regulated. Among the seven proteins which were significantly down-regulated by dutasteride in the epididymal luminal fluids, there were three β-defensins (Defb2, Defb18 and Defb39, which maybe the key proteins involved in epididymal sperm maturation and male fertility. Conclusions: We report for the first time that dutasteride influences the protein expression profiling in the epididymal luminal fluids of rats, and this result provides some new epididymal targets for male contraception and infertility therapy.

  4. Why is Taeniatherum caput-medusae (medusahead) Invasive in North America and not in its Native Eurasia?

    Science.gov (United States)

    Taeniatherum caput-medusae (medusahead) is an exotic annual grass introduced to North America in 1887 that has since invaded an estimated four million ha of rangelands. Contrary, in its native ranges of Eurasia, T. caput-medusae is not considered to be invasive. Why is it that T. caput-medusae expre...

  5. Epididymitis

    Science.gov (United States)

    ... to treat the infection. Sexually transmitted infections need antibiotics. Your sexual partners should also be treated. You may need pain medicines and anti-inflammatory medicines. If you are taking amiodarone, you may ...

  6. Unique regional distribution of delta 4-3-ketosteroid-5 alpha-oxidoreductase and associated epididymal morphology in the marsupial, Didelphis virginiana.

    Science.gov (United States)

    Kelce, W R; Krause, W J; Ganjam, V K

    1987-09-01

    The epididymal epithelial ultrastructure has been described in the adult male North American opossum, Didelphis virginiana. Morphological results have suggested that absorptive activity is prominent in the proximal epididymal region by virtue of numerous microvilli, an endocytotic complex, dense granules, and multivesicular bodies in the apical cytoplasm. In contrast, the middle and distal epididymal regions exhibit ultrastructural features indicative of protein synthesis such as large invaginated euchromatic nuclei, large nucleoli, and increased amounts of granular endoplasmic reticulum. It is in the middle and distal epididymal regions where sperm head rotation and sperm pairing take place. Epididymal delta 4-3-ketosteroid-5 alpha-oxidoreductase (5 alpha-reductase) activity also has been measured. It has been found that the level of enzyme activity differs significantly (p less than 0.01) between the proximal, middle, and distal epididymal regions. Enzyme-specific activity has been found to be highest in the middle region (47.6 +/- 5.4 picomoles 5 alpha-reduced androgens formed/b/mg protein), lower in the distal region (18.3 +/- 0.7 picomoles 5 alpha-reduced androgens formed/b/mg protein), with little activity (2.4 +/- 1.2 picomoles 5 alpha-reduced androgens formed/h/mg protein) found in the proximal epididymal region. This regional distribution of enzyme activity differs markedly from that reported for eutherian mammals. Both the suggested epididymal protein synthetic and secretory activity and the level of epididymal 5 alpha-reductase activity appear to correlate regionally with the morphological changes that occur in the opossum spermatozoa as they transit the epididymis.

  7. [Effects of L-carnitine on the apoptosis of spermatogenic cells and epididymal sperm count and motility in rats with diabetes mellitus].

    Science.gov (United States)

    Kang, Ning; Ma, Jie-hua; Zhou, Xin; Fan, Xiao-bo; Shang, Xue-jun; Huang, Yu-feng

    2011-05-01

    To explore the effects of L-carnitine (LC) on the apoptosis of spermatogenic cells and on the count and motility of epididymal sperm in rats with diabetes mellitus (DM). Twenty-four SD rats (200-230 g) were randomly divided into a control group, a DM model group and an LC group. After the establishment of DM models in the latter two groups by injection of streptozotocin (STZ) at 65 mg/kg, the controls and DM models were treated intragastrically with physiological saline, while the rats in the LC group with LC at 300 mg/kg, all for 6 consecutive weeks. Twenty-four hours after the last administration, all the rats were killed for the detection of the count and motility of epididymal sperm and the apoptosis of spermatogenic cells. The motilities of caput and cauda epididymal sperm were (53.7 +/- 1.8)% and (60.3 +/- 1.6)% in the LC group, significantly higher than in the DM model group ([32.2 +/- 2.0]% and [40.5 +/- 1.4]%, P count of cauda epididymal sperm was (25.5 +/- 1.1) x 10(6)/100 mg in the DM models, and was increased to (32.0 +/- 1.5) x 10(6)/100 mg after LC treatment (P sperm count, improved sperm motility, and reduced the apoptosis of spermatogenic cells in rats with DM.

  8. Circulating aldosterone induces the apical accumulation of the proton pumping V-ATPase and increases proton secretion in clear cells in the caput epididymis.

    Science.gov (United States)

    Roy, Jeremy W; Hill, Eric; Ruan, Ye Chun; Vedovelli, Luca; Păunescu, Teodor G; Brown, Dennis; Breton, Sylvie

    2013-08-15

    Clear cells express the vacuolar proton-pumping H(+)-ATPase (V-ATPase) and acidify the lumen of the epididymis, a process that is essential for male fertility. The renin-angiotensin-aldosterone system (RAAS) regulates fluid and electrolyte balance in the epididymis, and a previous study showed binding of aldosterone exclusively to epididymal clear cells (Hinton BT, Keefer DA. Steroid Biochem 23: 231-233, 1985). We examined here the role of aldosterone in the regulation of V-ATPase in the epididymis. RT-PCR showed expression of the mineralocorticoid receptor [MR; nuclear receptor subfamily 3, group C member 2 (NR3C2)] and 11-β-dehydrogenase isozyme 2 (HSD11β2) mRNAs specifically in clear cells, isolated by fluorescence-activated cell sorting from B1-enhanced green fluorescent protein (EGFP) mice. Tail vein injection of adult rats with aldosterone, 1,2-dioctanoyl-sn-glycerol (DOG), or 8-(4-chlorophenylthio)-cAMP (cpt-cAMP) induced V-ATPase apical membrane accumulation and extension of V-ATPase-labeled microvilli in clear cells in the caput epididymis but not in the cauda. V-ATPase activity was measured in EGFP-expressing clear cells using the intracellular pH (pHi)-sensing dye seminaphthorhodafluor-5F-5-(and 6)-carboxylic acid, acetoxymethyl ester acetate (SNARF-5F). Aldosterone induced a rapid increase in the rate of Na(+)- and bicarbonate-independent pHi recovery following an NH4Cl-induced acid load in clear cells isolated from the caput but not the cauda. This effect was abolished by concanamycin A, spironolactone, and chelerythrine but not myristoylated-protein kinase inhibitor (mPKI) or mifepristone. Thus aldosterone increases V-ATPase-dependent proton secretion in clear cells in the caput epididymis via MR/NR3C2 and PKC activation. This study, therefore, identifies aldosterone as an active member of the RAAS for the regulation of luminal acidification in the proximal epididymis.

  9. DNA fragmentation in spermatozoa

    DEFF Research Database (Denmark)

    Rex, A S; Aagaard, J.; Fedder, J

    2017-01-01

    Sperm DNA Fragmentation has been extensively studied for more than a decade. In the 1940s the uniqueness of the spermatozoa protein complex which stabilizes the DNA was discovered. In the fifties and sixties, the association between unstable chromatin structure and subfertility was investigated....... In the seventies, the impact of induced DNA damage was investigated. In the 1980s the concept of sperm DNA fragmentation as related to infertility was introduced as well as the first DNA fragmentation test: the Sperm Chromatin Structure Assay (SCSA). The terminal deoxynucleotidyl transferase nick end labelling...... (TUNEL) test followed by others was introduced in the nineties. The association between DNA fragmentation in spermatozoa and pregnancy loss has been extensively investigated spurring the need for a therapeutic tool for these patients. This gave rise to an increased interest in the aetiology of DNA damage...

  10. Isoindolinone-containing meroterpenoids with α-glucosidase inhibitory activity from mushroom Hericium caput-medusae.

    Science.gov (United States)

    Chen, Lin; Li, Zheng-Hui; Yao, Jian-Neng; Peng, Yue-Ling; Huang, Rong; Feng, Tao; Liu, Ji-Kai

    2017-10-01

    Hericium caput-medusae is an edible and medicinal mushroom closely relative to H. erinaceus. According to our detailed chemical investigation, two novel isoindolinone-containing meroterpene dimers, caputmedusins A (1) and B (2), as well as nine analogues, caputmedusins C-K (3-11), were isolated from the fermentation broth of H. caput-medusae. Their structures were elucidated by analyses of 1D and 2D NMR spectroscopic methods. The absolute configurations of 1-4 were speculated based on the specific optical rotation and biogenetic consideration. The absolute configurations of 10 and 11 were rationalized by the calculation of 1 H NMR chemical shifts. Caputmedusins A-C (1-3) showed moderate inhibitory activity against α-glucosidase with the IC 50 values of 39.2, 36.2 and 40.8μM, respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Impact of the Processes of Total Testicular Regression and Recrudescence on the Epididymal Physiology of the Bat Myotis nigricans (Chiroptera: Vespertilionidae.

    Directory of Open Access Journals (Sweden)

    Mateus R Beguelini

    Full Text Available Myotis nigricans is a species of vespertilionid bat, whose males show two periods of total testicular regression within the same annual reproductive cycle in the northwest São Paulo State, Brazil. Studies have demonstrated that its epididymis has an elongation of the caudal portion, which stores spermatozoa during the period of testicular regression in July, but that they had no sperm during the regression in November. Thus, the aim of this study was to analyze the impact of the total testicular regression in the epididymal morphophysiology and patterns of its hormonal regulation. The results demonstrate a continuous activity of the epididymis from the Active to the Regressing periods; a morphofunctional regression of the epididymis in the Regressed period; and a slow recrudescence process. Thus, we concluded that the processes of total testicular regression and posterior recrudescence suffered by M. nigricans also impact the physiology of the epididymis, but with a delay in epididymal response. Epididymal physiology is regulated by testosterone and estrogen, through the production and secretion of testosterone by the testes, its conduction to the epididymis (mainly through luminal fluid, conversion of testosterone to dihydrotestosterone by the 5α-reductase enzyme (mainly in epithelial cells and to estrogen by aromatase; and through the activation/deactivation of the androgen receptor and estrogen receptor α in epithelial cells, which regulate the epithelial cell morphophysiology, prevents cell death and regulates their protein expression and secretion, which ensures the maturation and storage of the spermatozoa.

  12. Hydrodynamics of insect spermatozoa

    Science.gov (United States)

    Pak, On Shun; Lauga, Eric

    2010-11-01

    Microorganism motility plays important roles in many biological processes including reproduction. Many microorganisms propel themselves by propagating traveling waves along their flagella. Depending on the species, propagation of planar waves (e.g. Ceratium) and helical waves (e.g. Trichomonas) were observed in eukaryotic flagellar motion, and hydrodynamic models for both were proposed in the past. However, the motility of insect spermatozoa remains largely unexplored. An interesting morphological feature of such cells, first observed in Tenebrio molitor and Bacillus rossius, is the double helical deformation pattern along the flagella, which is characterized by the presence of two superimposed helical flagellar waves (one with a large amplitude and low frequency, and the other with a small amplitude and high frequency). Here we present the first hydrodynamic investigation of the locomotion of insect spermatozoa. The swimming kinematics, trajectories and hydrodynamic efficiency of the swimmer are computed based on the prescribed double helical deformation pattern. We then compare our theoretical predictions with experimental measurements, and explore the dependence of the swimming performance on the geometric and dynamical parameters.

  13. Optimization of a protocol for cryopreservation of mouse spermatozoa using cryotubes.

    Science.gov (United States)

    Hasegawa, Ayumi; Yonezawa, Kazuya; Ohta, Akihiko; Mochida, Keiji; Ogura, Atsuo

    2012-01-01

    The rapid increase in the number of genetically modified mouse strains has produced a high demand for their frozen spermatozoa from laboratories and mouse banking facilities. Historically, plastic straws have been used preferentially as containers for frozen mammalian spermatozoa because spermatozoa frozen in plastic straws have a high survival rate after thawing. However, plastic straws are more fragile and are used less often than the cryotubes used for conventional cell freezing. In this study, we sought to develop a new protocol for sperm freezing using cryotubes as the container to increase the accessibility of mouse sperm cryopreservation. Epididymal spermatozoa were collected from mature ICR or C57BL/6J (B6) males and were suspended in 18% raffinose and 3% skim milk solution. We then optimized the following conditions using the sperm survival rate as an index: 1) distance of cryotubes from the surface of the liquid nitrogen at freezing, 2) volume of the sperm suspension in the cryotube and 3) temperature of warming sperm during thawing. The best result was obtained when cryotubes containing 10 µl of sperm suspension were immersed 1 cm below the surface of the liquid nitrogen and then thawed at 50 C. The fertilization rates using spermatozoa frozen and thawed using this method were 63.1% in ICR mice and 28.2% in B6 mice. The latter rate was increased to 62.3% by adding reduced glutathione to the fertilization medium. After embryo transfer, 68% and 62% of the fertilized oocytes developed into normal offspring in the ICR and B6 strains, respectively. These results show that cryotubes can be used for cryopreservation of mouse spermatozoa under optimized conditions. This protocol is easy and reproducible, and it may be used in laboratories that do not specialize in sperm cryopreservation.

  14. Epididymis response partly compensates for spermatozoa oxidative defects in snGPx4 and GPx5 double mutant mice.

    Directory of Open Access Journals (Sweden)

    Anaïs Noblanc

    Full Text Available We report here that spermatozoa of mice lacking both the sperm nucleus glutathione peroxidase 4 (snGPx4 and the epididymal glutathione peroxidase 5 (GPx5 activities display sperm nucleus structural abnormalities including delayed and defective nuclear compaction, nuclear instability and DNA damage. We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2O(2-scavenger activity especially in the cauda epididymis. Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity. Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation. Nevertheless, the enzymatic epididymal salvage response is sufficient to maintain full fertility of double KO males whatever their age, crossed with young WT female mice.

  15. Steroid metabolism by monkey and human spermatozoa

    International Nuclear Information System (INIS)

    Rajalakshmi, M.; Sehgal, A.; Pruthi, J.S.; Anand-Kumar, T.C.

    1983-01-01

    Freshly ejaculated spermatozoa from monkey and human were washed and incubated with tritium labelled androgens or estradiol to study the pattern of spermatozoa steroid metabolism. When equal concentrations of steroid substrates were used for incubation, monkey and human spermatozoa showed very similar pattern of steroid conversion. Spermatozoa from both species converted testosterone mainly to androstenedione, but reverse conversion of androstenedione to testosterone was negligible. Estradiol-17 beta was converted mainly to estrone. The close similarity between the spermatozoa of monkey and men in their steroid metabolic pattern indicates that the rhesus monkey could be an useful animal model to study the effect of drugs on the metabolic pattern of human spermatozoa

  16. Acute epididymitis in Greek children: a 3-year retrospective study.

    Science.gov (United States)

    Sakellaris, George S; Charissis, Giorgos C

    2008-07-01

    The aim of the study was to compare historical features, physical examination findings, and testicular color Doppler ultrasound in pediatric patients with epididymitis compared to testicular torsion and torsion of the appendix testes. A retrospective review of the medical records of 66 boys presenting with clinical aspects of acute scrotum over a 3-year period was performed. Sixty-six patients were included in the study (29 with epididymitis, 8 with testicular torsion and 12 with torsion of the appendix testis, 4 with scrotal abscesses, 5 with scrotal swelling, and 1 with inflamed epididymal cyst). The duration of symptoms ranged from 6 h to 4 days with a peak on the second day. Urine cultures and viral testes were negative in all patients. Color Doppler ultrasound was diagnostic for epididymitis in 28 patients (96.6%). Systemic intravenous antibiotics were given in all 29 patients with epididymitis. No patient showed signs of testicular atrophy in the follow-up. The increasing incidence of epididymitis should question the policy of routine exploration of the acute scrotum in children. The history and physical examination cannot reliably identify those boys who can be managed conservatively. Color Doppler ultrasound is a useful adjunct in the evaluation of the acute scrotum when physical findings are equivocal but it can also be misleading.

  17. Protective effect of combined pumpkin seed and ginger extracts on sperm characteristics, biochemical parameters and epididymal histology in adult male rats treated with cyclophosphamide.

    Science.gov (United States)

    Aghaie, Somaieh; Nikzad, Hossein; Mahabadi, Javad Amini; Taghizadeh, Mohsen; Azami-Tameh, Abolfazl; Taherian, Aliakbar; Sajjadian, Seyyed Mohammad Sajjad; Kamani, Mehran

    2016-09-01

    Reproductive toxicity is one of the side effects of cyclophosphamide (CP) in cancer treatment. Pumpkin seeds and Zingiber officinale are natural sources of antioxidants. We investigated the possible protective effect of combined pumpkin seed and Zingiber officinale extracts on sperm characteristics, epididymal histology and biochemical parameters of CP-treated rats. Male adult Wistar rats were divided randomly into six groups. Group 1, as a control, received an isotonic saline solution injection intraperitoneally (IP). Group 2 were injected IP with a single dose of CP (100 mg/kg) once. Groups 3 and 4 received CP plus 300 and 600 mg/kg combined pumpkin seed and Zingiber officinale extract (50:50). Groups 5 and 6 received only 300 and 600 mg/kg combined pumpkin seed and Zingiber officinale extract. Six weeks after treatment, sperm characteristics, histopathological changes and biochemical parameters were assessed. In CP-treated rats, motile spermatozoa were decreased, and abnormal or dead spermatozoa increased significantly (P < 0.001) but administration of the mixed extract improved sperm parameters. Epididymal epithelium and fibromascular thickness were also improved in extract-treated rats compared to control or CP groups. Biochemical analysis showed that the administration of combined extracts could increase the total antioxidant capacity (TAC) level significantly in groups 3, 4, 5 and 6. Interestingly, the mixed extract could decrease most of the side effects of CP such as vacuolization and separation of epididymal tissue. Our findings indicated that the combined extracts might be used as a protective agent against CP-induced reproductive toxicity.

  18. Confocal Microscopy and Image Analysis Indicates a Region-Specific Relation between Active Caspases and Cytoplasm in Ejaculated and Epididymal Sperm

    Science.gov (United States)

    García Vazquez, Susana; Aragón Martínez, Andrés; Flores-Alonso, Juan Carlos

    2012-01-01

    Previously, it was suggested a relation between the presence of apoptosis markers with cytoplasm in mammalian sperm. In this work, flow cytometry, confocal microscopy and image analysis were used to analyze the relationship between active caspase-3 and -7 and intracellular esterases expression in ejaculated or epididymal ram sperm. Sperm obtained from ejaculates from the caput, corpus, or cauda of the epididymis were treated with an inhibitor of active caspase-3 and -7 and a marker of cytoplasmic esterases. Additionally, ejaculated sperm were incubated for one, two, or three hours before evaluation for active caspases. Sperm subpopulations positive for active caspases and/or intracellular esterases were detected by flow cytometry and confocal microscopy; however, image analysis of confocal images showed that the correlation between active caspases and cytoplasmic esterases in sperm is region-specific. Lower values of Spearman correlation coefficients were found when whole sperm or head sperm was analyzed; however, a high correlation was observed for midpiece sperm. Incubation of sperm for two or three hours promoted the autoactivation of caspases. It has been suggested that the presence of apoptotic markers in sperm are related with a process of abortive apoptosis and with errors during spermiogenesis. Our results permit us suggest that the origin of the relationship between active caspases and cytoplasmic esterases is due to differentiation errors occurring during spermiogenesis because the percentages of sperm with active caspases are not different in the caput, corpus, or cauda of the epididymis. In this study we demonstrate that existing sperm subpopulations can express active caspases and intracellular esterases and that the correlation between these molecules is high in midpiece sperm. PMID:22530029

  19. Management of acute epididymitis: are European guidelines being followed?

    Science.gov (United States)

    Drury, Nigel E; Dyer, Jonathan P; Breitenfeldt, Nicole; Adamson, Andrew S; Harrison, G S M

    2004-10-01

    Acute epididymitis is increasing in men aged 35 years or under due to sexually-transmitted Chlamydia trachomatis. This study examines whether Urological surgeons are following European guidelines for the management of acute epididymitis in these patients. A postal questionnaire survey was conducted of specialists in Urology in two regions of the UK. Of 79 completed replies, 41 (52%) take a detailed sexual history but only 34 (43%) refer patients to a Genitourinary medicine clinic. Quinolones are the most commonly prescribed first-line antibiotic by 56 (71%) respondents, principally ciprofloxacin. The current management of acute epididymitis in young men must be improved. Ciprofloxacin is not the optimal antimicrobial for the treatment of urogenital chlamydial infection. We recommend that all such patients be referred to local Genitourinary medicine services for contact-tracing and treatment of their sexual partners.

  20. Neonatal outcome and congenital malformations in children born after ICSI with testicular or epididymal sperm

    DEFF Research Database (Denmark)

    Fedder, Jens; Loft, A; Parner, Erik Thorlund

    2013-01-01

    STUDY QUESTION: Does neonatal outcome including congenital malformations in children born after ICSI with epididymal and testicular sperm [testicular sperm extraction (TESE)/percutaneous epididymal sperm aspiration (PESA)/testicular sperm aspiration (TESA) (TPT)] differ from neonatal outcome in c...

  1. Mycoplasma genitalium attaches to human spermatozoa

    DEFF Research Database (Denmark)

    Svenstrup, Helle Friis; Fedder, Jens; Abraham-Peskir, Joanna

    2003-01-01

    BACKGROUND: Mycoplasma genitalium causes urogenital diseases in men and women and is presumed to be sexually transmitted. We wanted to investigate whether spermatozoa could serve as vectors for M.genitalium in order to cause upper genital diseases in women. METHODS: By use of Nomarski light...... microscopy and transmission X-ray microscopy, the attachment of M.genitalium to spermatozoa was studied. Semen was incubated in vitro with M.genitalium. Purified, motile spermatozoa were examined for attachment of M.genitalium by immunofluorescence microscopy. RESULTS: Mycoplasma genitalium was shown...... to adhere to the head, midpiece and tail of the spermatozoa. The spermatozoa became immotile when many M.genitalium were attached. However, the motile spermatozoa were demonstrated to carry M.genitalium and in this case the mycoplasmas were seen to attach mostly to the midpiece or neck region. Occasionally...

  2. Assessment of azadirachtin-A, a neem tetranortritarpinoid, on rat spermatozoa during in vitro capacitation.

    Science.gov (United States)

    Katte, Teesta V; Rajyalakshmi, Malempati; Aladakatti, Ravindranath H

    2018-05-05

    The exploration of the biological assessment of technical azadirachtin, a tetranortritarpinoid from the neem seed kernel, was reviewed. The present study was, therefore, designed to evaluate the dose-dependent in vitro effects of azadirachtin-A, particularly on the functional studies and determination of molecular events, which are critical in the process of sperm capacitation. To assess the effects of the azadirachtin-A on the functional studies, sperm capacitation, the total sperm adenosine triphosphate levels, acrosome reaction (AR), the sperm-egg interaction and the determination of molecular events like cyclic adenosine-3',5'-monophosphate and calcium levels, the appropriate volumes of the sperm suspension were added to the medium to a final concentration of 1×106 sperm/mL and incubated in a humidified atmosphere of 5% CO2 in air at 37°C. The increasing quantities 0.5-2.0 mM/mL and the equivalent volumes of 50% dimethyl sulfoxide were added to the control dishes prior to the addition of spermatozoa and then observed at various time-points for motility and other analyses. Results revealed the dose- and time-dependent decrease in the functional consequence of capacitation, i.e. the percentage of motile spermatozoa, motility score and sperm motility index, levels of molecular events in spermatozoa, followed by declined spontaneous AR leading to lesser binding of the cauda epididymal sperm to the Zona pellucida. The findings confirm the inhibition of rat sperm motility by blocking some biochemical pathways like energy utilization. They also demonstrate that sperm capacitation is associated with the decrease in AR and that the levels of molecular events in spermatozoa can guide us towards the development of a new male contraceptive constituent.

  3. A Patient Presenting with Concurrent Testis Torsion and Epididymal Leiomyoma

    Directory of Open Access Journals (Sweden)

    E. Arpali

    2013-01-01

    Full Text Available Leiomyomas are the second most common tumors of epididymis. Patients with leiomyomas are sometimes misdiagnosed with testicular tumors. A Case of a patient with a scrotal mass presenting with testicular torsion is reported. Concurrent occurrence of testicular torsion and epididymal leiomyoma is an extremely rare condition.

  4. Effects of concomitant diabetes mellitus and hyperthyroidism on testicular and epididymal histoarchitecture and steroidogenesis in male animals*

    Science.gov (United States)

    Korejo, Nazar Ali; Wei, Quan-wei; Shah, Atta Hussain; Shi, Fang-xiong

    2016-01-01

    This study evaluated the effects of comorbid disorders of diabetes and hyperthyroidism in the adult male mice. In total, 32 ICR strain mice were equally distributed into four groups: control (C), diabetic (D), diabetic-plus-hyperthyroid (DH), and hyperthyroid (H). Mice allocated for diabetes received a single intraperitoneal injection of streptozotocin (STZ) at 200 mg/kg body weight. At the onset of diabetes, one group of mice was concomitantly injected levothyroxine (LT4; 0.3 mg/kg body weight) and the other set of animals received the same treatment independently on a daily basis. The body weight, as well as the testicular and epididymal weights, was reduced markedly in D and DH mice. Higher trends of blood glucose levels were seen in the DH group, in comparison to euthyroid diabetic mice. Thyroid hormones could exert a transient effect on blood glucose homeostasis by altering the serum blood glucose level in diabetic patients. Histomorphometric analysis showed increased luminal sizes of seminiferous tubules, along with decreased epithelial height and atrophic changes in germinal stem cells in the testis of DH and H mice. Caput epididymis of DH mice showed extensive compaction of principal cells, loss of stereocilia, lipid vacuolization, and inflammatory infiltrations; however, damaged tubular integrity, packed clear cells, exfoliated cells, and round spermatids were profoundly noticed in the cauda epididymis. Hyperthyroidism elevated the serum testosterone levels in H and DH mice and produced critical damages to the histoarchitecture of the epididymis. Collectively, this experiment endeavored to mimic the polyglandular autoimmune syndrome, which will be helpful to better understand the reasons for male infertility in diabetic-cum-hyperthyroid patients. PMID:27819132

  5. Effects of concomitant diabetes mellitus and hyperthyroidism on testicular and epididymal histoarchitecture and steroidogenesis in male animals.

    Science.gov (United States)

    Korejo, Nazar Ali; Wei, Quan-Wei; Shah, Atta Hussain; Shi, Fang-Xiong

    This study evaluated the effects of comorbid disorders of diabetes and hyperthyroidism in the adult male mice. In total, 32 ICR strain mice were equally distributed into four groups: control (C), diabetic (D), diabetic-plus-hyperthyroid (DH), and hyperthyroid (H). Mice allocated for diabetes received a single intraperitoneal injection of streptozotocin (STZ) at 200 mg/kg body weight. At the onset of diabetes, one group of mice was concomitantly injected levothyroxine (LT4; 0.3 mg/kg body weight) and the other set of animals received the same treatment independently on a daily basis. The body weight, as well as the testicular and epididymal weights, was reduced markedly in D and DH mice. Higher trends of blood glucose levels were seen in the DH group, in comparison to euthyroid diabetic mice. Thyroid hormones could exert a transient effect on blood glucose homeostasis by altering the serum blood glucose level in diabetic patients. Histomorphometric analysis showed increased luminal sizes of seminiferous tubules, along with decreased epithelial height and atrophic changes in germinal stem cells in the testis of DH and H mice. Caput epididymis of DH mice showed extensive compaction of principal cells, loss of stereocilia, lipid vacuolization, and inflammatory infiltrations; however, damaged tubular integrity, packed clear cells, exfoliated cells, and round spermatids were profoundly noticed in the cauda epididymis. Hyperthyroidism elevated the serum testosterone levels in H and DH mice and produced critical damages to the histoarchitecture of the epididymis. Collectively, this experiment endeavored to mimic the polyglandular autoimmune syndrome, which will be helpful to better understand the reasons for male infertility in diabetic-cum-hyperthyroid patients.

  6. Painless inter epididymal testicular torsion of the spermatic cord

    Directory of Open Access Journals (Sweden)

    Salomon V. Romano

    2007-02-01

    Full Text Available Inter epididymal testicular torsion of the spermatic cord is extremely rare and usually diagnosed at surgery. We present an unusual case of spermatic cord torsion in a 14-year-old male patient. It is important to highlight that the torsion occurred only on the distal half of the epididymis leaving the head untwisted and edematous. In addition, the fact that this condition was painless made this case extremely rare and motivated our presentation.

  7. Painless inter epididymal testicular torsion of the spermatic cord

    OpenAIRE

    Salomon V. Romano; Haime S. Hernan; Norberto Fredotovich

    2007-01-01

    Inter epididymal testicular torsion of the spermatic cord is extremely rare and usually diagnosed at surgery. We present an unusual case of spermatic cord torsion in a 14-year-old male patient. It is important to highlight that the torsion occurred only on the distal half of the epididymis leaving the head untwisted and edematous. In addition, the fact that this condition was painless made this case extremely rare and motivated our presentation.

  8. Biochemical and binding characteristics of boar epididymal fluid proteins

    Czech Academy of Sciences Publication Activity Database

    Maňásková-Postlerová, Pavla; Davidová, Nina; Jonáková, Věra

    2011-01-01

    Roč. 879, č. 1 (2011), s. 100-106 ISSN 1570-0232 R&D Projects: GA ČR(CZ) GA303/09/1285; GA MŠk(CZ) 1M06011; GA ČR(CZ) GD523/08/H064 Institutional research plan: CEZ:AV0Z50520701 Keywords : epididymal proteins * RP HPLC * biotin-labeled ligands Subject RIV: CE - Biochemistry Impact factor: 2.888, year: 2011

  9. Normal development of hamster and rabbit eggs fertilized by spermatozoa labelled with the fluorescent thiol alkylating agent, monobromobimane

    International Nuclear Information System (INIS)

    Fleming, A.D.; Cummins, J.M.; Kuehl, T.J.; Seidel, G.E. Jr.; Yanagimachi, R.

    1986-01-01

    Cauda epididymal spermatozoa of the golden hamster were labelled with the thiol-alkylating reagent, monobromobimane (MB). Female hamsters underwent uterine insemination with labelled spermatozoa at laparotomy under metofane anesthesia. All 12 females examined between 5 and 54 h postinsemination yielded a total of 83/100 (83%) eggs in the process of fertilization or embryos. Under ultraviolet (UV) exposure all exhibited a fluorescent tail which, in the 4- and 8-cell embryos, could be seen to be fraying or disintegrating. As cleavage progressed, labelled tail components came to be restricted among the blastomeres such that at the 4- and 8-cell stage the tail could be seen in only one to three blastomeres. To study complete development and pregnancy another 12 females received uterine insemination. After recovery from anesthesia (approximately equal to 4 h) these females were mated with a vasectomized male bearing a dominant genetic marker (black eyes) to allow unequivocal determination of paternity in the fetuses and young produced. Seven became pregnant with one female losing her pregnancy about Day 7 of gestation. Two females sacrificed on Day 13 produced 17 normal fetuses and one resorption. Four females delivered 16 young, all normal at birth and in subsequent growth and fertility. In addition, insemination of female rabbits with MB-labelled spermatozoa yielded normal embryos (50/52 96%) from 3 does on Day 2 and (38/64 60%) from 4 does on Days 4 or 5. Two normal litters (9 bunnies) have delivered from 3 does allowed to carry to term

  10. Sperm flagellum volume determines freezability in red deer spermatozoa.

    Directory of Open Access Journals (Sweden)

    José Luis Ros-Santaella

    Full Text Available The factors affecting the inter-individual differences in sperm freezability is a major line of research in spermatology. Poor sperm freezability is mainly characterised by a low sperm velocity, which in turn is associated with low fertility rates in most animal species. Studies concerning the implications of sperm morphometry on freezability are quite limited, and most of them are based on sperm head size regardless of the structural parts of the flagellum, which provides sperm motility. Here, for the first time, we determined the volumes of the flagellum structures in fresh epididymal red deer spermatozoa using a stereological method under phase contrast microscopy. Sperm samples from thirty-three stags were frozen and classified as good freezers (GF or bad freezers (BF at two hours post-thawing using three sperm kinetic parameters which are strongly correlated with fertility in this species. Fourteen stags were clearly identified as GF, whereas nineteen were BF. No significant difference in sperm head size between the two groups was found. On the contrary, the GF exhibited a lower principal piece volume than the BF (6.13 µm3 vs 6.61 µm3, respectively, p = 0.006. The volume of the flagellum structures showed a strong negative relationship with post-thawing sperm velocity. For instance, the volume of the sperm principal piece was negatively correlated with sperm velocity at two hours post-thawing (r = -0.60; p<0.001. Our results clearly show that a higher volume of the sperm principal piece results in poor freezability, and highlights the key role of flagellum size in sperm cryopreservation success.

  11. Incorporation of nanoparticles within mammalian spermatozoa using in vitro capacitation

    Science.gov (United States)

    There is still much unknown about the journey of spermatozoa within the female genital tract. Recent studies have investigated mammalian spermatozoa labeling with fluorescent quantum dot nanoparticles (QD) for non-invasive imaging. Furthermore, the incorporation of these QD within the spermatozoa ma...

  12. Percutaneous medial hemi-epiphysiodesis using a transphyseal screw for caput valgum associated with developmental dysplasia of the hip.

    Science.gov (United States)

    Shin, Chang Ho; Hong, Wan Kee; Lee, Doo Jae; Yoo, Won Joon; Choi, In Ho; Cho, Tae-Joon

    2017-11-14

    The purpose of this study was to evaluate the radiologic outcome of percutaneous medial hemi-epiphysiodesis using a transphyseal screw for the management of caput valgum associated with developmental dysplasia of the hip (DDH). Eighteen hips (18 patients) having caput valgum treated with screw hemi-epiphysiodesis were followed for more than 2 years, and were included in this study. The mean age at the time of the index operation was 8.3 years (range, 4.3 to 10.7 years) and age at the latest follow-up was 12.2 years (range, 9.4 to 16.4 years). The screw in 5 hips was changed into a longer one at postoperative 21.8 months (range, 14 to 29 months) because the proximal femur outgrew the screw. The screws in 11 hips were removed at the mean age of 10.9 years (range, 8.0 to 14.5 years). We retrospectively analyzed the change in various radiologic parameters over time. The mean Hilgenreiner-epiphyseal angle (HEA) of the operated side was 5.1 ± 11.3° preoperatively, and increased to 20.6 ± 11.3° at the latest follow-up (p = 0.001). The mean difference of the HEA between the operated and contralateral sides was 16.9 ± 15.1° preoperatively, which decreased to 2.4 ± 12.4° at the latest follow-up (p = 0.008). The mean articulo-trochanteric distance of the operated side, which was 3.2 ± 5.5 mm longer than that of the contralateral side preoperatively, became 5.6 ± 9.1 mm shorter at the latest follow-up (p = 0.001). The ratio of femoral neck length of the operated side to that of the contralateral side decreased over the follow-up period. Acetabular shape as measured by the Sharp angle and acetabular roof angle and femoral head coverage as measured by lateral center-edge angle did not change significantly by the index operation. The ratio of medial joint space width of the operated side to that of the contralateral side did not change significantly. Screw medial hemi-epiphysiodesis can effectively correct caput valgum

  13. PERUBAHAN VIABILITAS DAN STRUKTUR SUBSELULER SPERMATOZOA DOMBA SETELAH PENGERINGBEKUAN

    Directory of Open Access Journals (Sweden)

    Takdir Saili

    2009-12-01

    Full Text Available Several methods i.e. cooling, freezing, and freeze-drying have been widely used to preserve spermatozoa with various degree of success. Freeze-drying appears to provide a method to preserve spermatozoa in a dry state without requiring liquid nitrogen for storing frozen spermatozoa. Freeze-drying procedures can have a detrimental effect on plasma membrane and acrosomal cap of the spermatozoa. In this experiment study, the viability and subcellular changes of freeze-dried ram spermatozoa were evaluated using staining method and scanning electron microscopy. The results revealed that all freeze-dried spermatozoa were dead following evaluation using eosin staining and Hoechst-propidium iodide staining methods. Morover, plasma membrane and acrosomal cap of freeze-dried ram spermatozoa was disrupted observed using scanning electron microscope.

  14. Genetic damage caused by methyl-parathion in mouse spermatozoa is related to oxidative stress

    International Nuclear Information System (INIS)

    Pina-Guzman, B.; Solis-Heredia, M.J.; Rojas-Garcia, A.E.; Uriostegui-Acosta, M.; Quintanilla-Vega, B.

    2006-01-01

    Organophosphorous (OP) pesticides are considered genotoxic mainly to somatic cells, but results are not conclusive. Few studies have reported OP alterations on sperm chromatin and DNA, and oxidative stress has been related to their toxicity. Sperm cells are very sensitive to oxidative damage which has been associated with reproductive dysfunctions. We evaluated the effects of methyl-parathion (Me-Pa; a widely used OP) on sperm DNA, exploring the sensitive stage(s) of spermatogenesis and the relationship with oxidative stress. Male mice (10-12-weeks old) were administered Me-Pa (3-20 mg/kg bw/i.p.) and euthanized at 7- or 28-days post-treatment. Mature spermatozoa were obtained and evaluated for chromatin structure through SCSA (Sperm Chromatin Structure Assay; DNA Fragmentation Index parameters: Mean DFI and DFI%) and chromomycin-A 3 (CMA 3 )-staining, for DNA damage through in situ-nick translation (NT-positive) and for oxidative stress through lipid peroxidation (LPO; malondialdehyde production). At 7-days post-treatment (mature spermatozoa when Me-Pa exposure), dose-dependent alterations in chromatin structure (Mean DFI and CMA 3 -staining) were observed, as well as increased DNA damage, from 2-5-fold in DFI% and NT-positive cells. Chromatin alterations and DNA damage were also observed at 28-days post-treatment (cells at meiosis at the time of exposure); suggesting that the damage induced in spermatocytes was not repaired. Positive correlations were observed between LPO and sperm DNA-related parameters. These data suggest that oxidative stress is related to Me-Pa alterations on sperm DNA integrity and cells at meiosis (28-days post-treatment) and epididymal maturation (7-days post-treatment) are Me-Pa targets. These findings suggest a potential risk of Me-Pa to the offspring after transmission

  15. Effect of Mucuna pruriens (Linn.) on mitochondrial dysfunction and DNA damage in epididymal sperm of streptozotocin induced diabetic rat.

    Science.gov (United States)

    Suresh, Sekar; Prithiviraj, Elumalai; Lakshmi, Nagella Venkata; Ganesh, Mohanraj Karthik; Ganesh, Lakshmanan; Prakash, Seppan

    2013-01-09

    Mucuna pruriens Linn. (M. pruriens) is a leguminous plant that has been recognized as an herbal medicine for improving fertility and related disorders in the Indian traditional system of medicine, however without proper scientific validations. To study the effect of ethanolic seed extract of M. pruriens on mitochondrial dysfunction and the DNA damage in hyperglycemic rat epididymal spermatozoa. Male Wistar albino rats were divided as control (Sham), diabetes induced [streptozotocin 60 mg/kg of body weight (b.w.) in 0.1M citrate buffer] (STZ), diabetic rats administered with 200mg/kg b.w. of extract (STZ+MP) and normal rats administered with 200mg/kg b.w. of extract (Sham+MP). M. pruriens was administered (gavage) once daily for a period of 60 days. On 60th day animals were sacrificed by cervical dislocation sperm were collected from epididymis and subjected various analysis like antioxidants, ROS, lipid peroxidation (LPO), DNA damage, chromosomal integrity and mitochondrial membrane potential (MMP). Significant reduction in the sperm count, motility, viability and significant increase in the number of abnormal sperm in STZ compared to sham was noticed. STZ rat sperm showed significant increase in LPO and DNA damage. Both the enzymic and non-enzymic were decreased; MMP and the mitochondrial functions were severely affected in STZ group. The diabetic rats supplemented with M. pruriens showed a remarkable recovery in antioxidant levels and reduced LPO with well preserved sperm DNA. MMP and mitochondrial function test were also preserved in STZ+MP rat sperm. The present study has clearly demonstrated the potency of M. pruriens to reduce the diabetic induced sperm damage induced by oxidative stress (OS). These observations are encouraging to perform similar studies in human. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  16. Detection of spermatozoa following consensual sexual intercourse

    DEFF Research Database (Denmark)

    Astrup, Birgitte Schmidt; Thomsen, Jørgen Lange; Lauritsen, Jens

    2012-01-01

    INTRODUCTION: In cases of sexual assault, the finding of semen can provide crucial evidence. The presence of spermatozoa serves as proof of a sexual act and may give the identity of the alleged perpetrator through DNA-profiling. In most western countries, there are guidelines for standardized...... examinations of sexual assault victims. For an objective evaluation of the findings, substantial knowledge of aspects regarding consensual sexual intercourse is crucial. The aim of this study was to examine detection frequencies and genital sampling sites of spermatozoa following consensual sexual intercourse....... METHODS: In a prospective setting, 60 women underwent forensic examination following consensual sexual intercourse. Specimens were obtained from the external genitalia, the posterior fornix and the cervical canal, and examined using the Papanicolau stain and standard light microscopy. RESULTS: We found...

  17. Study of ionizing radiation effect on human spermatozoa chromosomes

    International Nuclear Information System (INIS)

    Rousseaux, S.

    1990-02-01

    The purpose of this thesis is to study the radio-induced chromosomal aberrations in spermatozoa. After a brief recall on ionizing radiations, the author reviews the radio-induced chromosomal anomalies on somatic cells and on germinal line cells and spermatozoa. The author presents the technical aspects of human spermatozoa karyotype and finally studies the radio induced chromosomal anomalies of sperm to patients undergoing a radiotherapy. 13 tabs., 28 figs., 28 photos

  18. A Systematic Analysis of a Deep Mouse Epididymal Sperm Proteome

    Energy Technology Data Exchange (ETDEWEB)

    Chauvin, Theodore; Xie, Fang; Liu, Tao; Nicora, Carrie D.; Yang, Feng; Camp, David G.; Smith, Richard D.; Roberts, Kenneth P.

    2012-12-21

    Spermatozoa are highly specialized cells that, when mature, are capable of navigating the female reproductive tract and fertilizing an oocyte. The sperm cell is thought to be largely quiescent in terms of transcriptional and translational activity. As a result, once it has left the male reproductive tract, the sperm cell is essentially operating with a static population of proteins. It is therefore theoretically possible to understand the protein networks contained in a sperm cell and to deduce its cellular function capabilities. To this end we have performed a proteomic analysis of mouse sperm isolated from the cauda epididymis and have confidently identified 2,850 proteins, which is the most comprehensive sperm proteome for any species reported to date. These proteins comprise many complete cellular pathways, including those for energy production via glycolysis, β-oxidation and oxidative phosphorylation, protein folding and transport, and cell signaling systems. This proteome should prove a useful tool for assembly and testing of protein networks important for sperm function.

  19. Oxidative and antioxidative status in the testes of rats with acute epididymitis.

    Science.gov (United States)

    Kaya, Mete; Boleken, Mehmet Emin; Zeyrek, Fadile; Ozardali, Ilyas; Kanmaz, Turan; Erel, Ozcan; Yücesan, Selçuk

    2006-01-01

    Epididymitis is an inflammation or infection of the epididymis, a convoluted duct that lies on the posterior surface of the testicle. Oxidative stress due to excessive production of reactive oxygen species in epididymitis, impaired antioxidant defense mechanisms, or both, precipitates a range of pathologies that are currently believed to negatively affect the male reproductive function. How oxidative stress affects the testes is still unknown. We aimed to investigate the oxidative and antioxidative status of testes of rats with unilateral acute Escherichia coli epididymitis. The study included 36 male Wistar albino rats which were divided into three groups. In the epididymitis group (n = 12), an E. coli suspension was injected into the right ductus deferens of rats, and the same amount of saline was injected in the saline groups (n = 12). No surgery was performed in the control group (n = 12) for baseline values. Rats were sacrificed after 24 h and the epididymis and testes removed. The infection was confirmed by histopathologic evaluation and microbiological tests. The oxidative status of testes was evaluated by measuring myeloperoxidase (MPO) activity, and antioxidative status was evaluated by measuring total antioxidant response (TAR) and total antioxidant capacity levels (TAC). MPO activity in both the ipsilateral and contralateral testes of the epididymitis group was significantly higher than those of the saline and control groups (p antioxidants. 2006 S. Karger AG, Basel.

  20. Optimization of IVF pregnancy outcomes with donor spermatozoa.

    Science.gov (United States)

    Wang, Jeff G; Douglas, Nataki C; Prosser, Robert; Kort, Daniel; Choi, Janet M; Sauer, Mark V

    2009-03-01

    To identify risk factors for suboptimal IVF outcomes using insemination with donor spermatozoa and to define a lower threshold that may signal a conversion to fertilization by ICSI rather than insemination. Retrospective, age-matched, case-control study of women undergoing non-donor oocyte IVF cycles using either freshly ejaculated (N=138) or cryopreserved donor spermatozoa (N=69). Associations between method of fertilization, semen sample parameters, and pregnancy rates were analyzed. In vitro fertilization of oocytes with donor spermatozoa by insemination results in equivalent fertilization and pregnancy rates compared to those of freshly ejaculated spermatozoa from men with normal semen analyses when the post-processing motility is greater than or equal to 88%. IVF by insemination with donor spermatozoa when the post-processing motility is less than 88% is associated with a 5-fold reduction in pregnancy rates when compared to those of donor spermatozoa above this motility threshold. When the post-processing donor spermatozoa motility is low, fertilization by ICSI is associated with significantly higher pregnancy rates compared to those of insemination. While ICSI does not need to be categorically instituted when using donor spermatozoa in IVF, patients should be counseled that conversion from insemination to ICSI may be recommended based on low post-processing motility.

  1. CHARACTERIZATION OF THE OLFACTORY RECEPTORS EXPRESSED IN HUMAN SPERMATOZOA

    Directory of Open Access Journals (Sweden)

    Caroline eFlegel

    2016-01-01

    Full Text Available The detection of external cues is fundamental for human spermatozoa to locate the oocyte in the female reproductive tract. This task requires a specific chemoreceptor repertoire that is expressed on the surface of human spermatozoa, which is not fully identified to date. Olfactory receptors (ORs are candidate molecules and have been attributed to be involved in sperm chemotaxis and chemokinesis, indicating an important role in mammalian spermatozoa. An increasing importance has been suggested for spermatozoal RNA, which led us to investigate the expression of all 387 OR genes. This study provides the first comprehensive analysis of OR transcripts in human spermatozoa of several individuals by RNA-Seq. We detected 91 different transcripts in the spermatozoa samples that could be aligned to annotated OR genes. Using stranded mRNA-Seq, we detected a class of these putative OR transcripts in an antisense orientation, indicating a different function, rather than coding for a functional OR protein. Nevertheless, we were able to detect OR proteins in various compartments of human spermatozoa, indicating distinct functions in human sperm. A panel of various OR ligands induced Ca2+ signals in human spermatozoa, which could be inhibited by mibefradil. This study indicated that a variety of ORs are expressed at the mRNA and protein level in human spermatozoa and demonstrates that ORs are involved in the physiological processes.

  2. Orchitis and Epididymitis in Koalas (Phascolarctos cinereus) Infected With Chlamydia pecorum.

    Science.gov (United States)

    Johnston, S D; Deif, H H; McKinnon, A; Theilemann, P; Griffith, J E; Higgins, D P

    2015-11-01

    Although Chlamydia causes disease of the urethra and prostate of male koalas, its impact on the testis and epididymis has not been examined. This study describes chronic-active and granulomatous orchitis and epididymitis with interstitial fibrosis associated with infection by Chlamydia pecorum in 2 of 18 adult male koalas being euthanized at a koala hospital, 8 of which also had chlamydial prostatitis. By immunohistochemistry and transmission electron microscopy, chlamydial inclusions were demonstrated within Sertoli cells directly associated with mild inflammation surrounding intact seminiferous and epididymal tubules, marked pyogranulomatous inflammation around disrupted tubules, replacement of tubules by interstitial fibrosis, and aspermia. The presence of C. pecorum but not Chlamydia pneumoniae was detected by quantitative polymerase chain reaction of formalin-fixed tissues of the left and right testes and right epididymis in 1 animal. This is the first report of orchitis and epididymitis in a koala infected with C. pecorum. © The Author(s) 2015.

  3. Muellerian duct cyst as a cause of acute infantile-onset epididymitis

    Energy Technology Data Exchange (ETDEWEB)

    Donkol, Ragab H.; Moghazy, Khaled [Gulf Specialist Hospital, Department of Radiology, P.O. Box 1444, Qatif (Saudi Arabia); Monib, Sherif [Gulf Specialist Hospital, Department of General Surgery, Qatif (Saudi Arabia)

    2006-11-15

    Muellerian duct cyst is a congenital anomaly that originates from remnants of the muellerian duct that usually regresses in utero. It is most commonly detected in adult males during the third and fourth decades of life. Muellerian duct cyst is rarely reported in infants less than 1 year of age, in whom diagnosis is difficult because of immaturity of the genital tract. In this case report, we describe the sonographic and CT appearance of a large intrapelvic muellerian duct cyst presenting with acute epididymitis in a 9-month-old infant. The embryological basis, imaging findings and relation to epididymitis are discussed. (orig.)

  4. Muellerian duct cyst as a cause of acute infantile-onset epididymitis

    International Nuclear Information System (INIS)

    Donkol, Ragab H.; Moghazy, Khaled; Monib, Sherif

    2006-01-01

    Muellerian duct cyst is a congenital anomaly that originates from remnants of the muellerian duct that usually regresses in utero. It is most commonly detected in adult males during the third and fourth decades of life. Muellerian duct cyst is rarely reported in infants less than 1 year of age, in whom diagnosis is difficult because of immaturity of the genital tract. In this case report, we describe the sonographic and CT appearance of a large intrapelvic muellerian duct cyst presenting with acute epididymitis in a 9-month-old infant. The embryological basis, imaging findings and relation to epididymitis are discussed. (orig.)

  5. Molecular characterization and prevalence of two capulaviruses: Alfalfa leaf curl virus from France and Euphorbia caput-medusae latent virus from South Africa.

    Science.gov (United States)

    Bernardo, Pauline; Muhire, Brejnev; François, Sarah; Deshoux, Maëlle; Hartnady, Penelope; Farkas, Kata; Kraberger, Simona; Filloux, Denis; Fernandez, Emmanuel; Galzi, Serge; Ferdinand, Romain; Granier, Martine; Marais, Armelle; Monge Blasco, Pablo; Candresse, Thierry; Escriu, Fernando; Varsani, Arvind; Harkins, Gordon W; Martin, Darren P; Roumagnac, Philippe

    2016-06-01

    Little is known about the prevalence, diversity, evolutionary processes, genomic structures and population dynamics of viruses in the divergent geminivirus lineage known as the capulaviruses. We determined and analyzed full genome sequences of 13 Euphorbia caput-medusae latent virus (EcmLV) and 26 Alfalfa leaf curl virus (ALCV) isolates, and partial genome sequences of 23 EcmLV and 37 ALCV isolates. While EcmLV was asymptomatic in uncultivated southern African Euphorbia caput-medusae, severe alfalfa disease symptoms were associated with ALCV in southern France. The prevalence of both viruses exceeded 10% in their respective hosts. Besides using patterns of detectable negative selection to identify ORFs that are probably functionally expressed, we show that ALCV and EcmLV both display evidence of inter-species recombination and biologically functional genomic secondary structures. Finally, we show that whereas the EcmLV populations likely experience restricted geographical dispersion, ALCV is probably freely moving across the French Mediterranean region. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Predictive Factors for Natural Pregnancy after Microsurgical Reconstruction in Patients with Primary Epididymal Obstructive Azoospermia

    Directory of Open Access Journals (Sweden)

    Mihai Harza

    2014-01-01

    Full Text Available Primary epididymal obstructive azoospermia (OA is the most prevalent form of OA in nonvasectomized patients and has been less studied. We aim to assess the results with microsurgical vasoepididymostomy used in the treatment of men diagnosed with primary epididymal obstructive azoospermia and to identify the factors associated with natural pregnancy occurring after microsurgical reconstruction. This prospective study included consecutive patients with epididymal OA who underwent microsurgical reconstruction in our center. Clinical and biological data were obtained every three months during follow-up. Occurrence of natural pregnancy was the primary study outcome. In total, 36 patients underwent microsurgical reconstruction. The mean age was 34±4.5 years (range 24–46 years. Median follow-up time was 15 [IQR 12–21] months. The total patency rate was 77.7% (n=28. During follow-up, 8 (22.2% natural pregnancies occurred. The overall live birth rate was 100%. Low FSH levels (HR: 0.22; 95% CI: 0.052–0.88; P=0.032 and higher total motile sperm count (TMSC (HR: 1.001; 95% CI 1–1.001; P=0.012 were associated with a higher rate of natural pregnancy. Our data suggest that microsurgical vasoepididymostomy is an effective therapy of primary epididymal OA. Baseline lower FSH and higher TMSC were independent predictors for natural pregnancy occurrence.

  7. Predictive factors for natural pregnancy after microsurgical reconstruction in patients with primary epididymal obstructive azoospermia.

    Science.gov (United States)

    Harza, Mihai; Voinea, Sebastian; Ismail, Gener; Gagiu, Cristian; Baston, Catalin; Preda, Adrian; Manea, Ioan; Priporeanu, Tiberiu; Sinescu, Ioanel

    2014-01-01

    Primary epididymal obstructive azoospermia (OA) is the most prevalent form of OA in nonvasectomized patients and has been less studied. We aim to assess the results with microsurgical vasoepididymostomy used in the treatment of men diagnosed with primary epididymal obstructive azoospermia and to identify the factors associated with natural pregnancy occurring after microsurgical reconstruction. This prospective study included consecutive patients with epididymal OA who underwent microsurgical reconstruction in our center. Clinical and biological data were obtained every three months during follow-up. Occurrence of natural pregnancy was the primary study outcome. In total, 36 patients underwent microsurgical reconstruction. The mean age was 34 ± 4.5 years (range 24-46 years). Median follow-up time was 15 [IQR 12-21] months. The total patency rate was 77.7% (n = 28). During follow-up, 8 (22.2%) natural pregnancies occurred. The overall live birth rate was 100%. Low FSH levels (HR: 0.22; 95% CI: 0.052-0.88; P = 0.032) and higher total motile sperm count (TMSC) (HR: 1.001; 95% CI 1-1.001; P = 0.012) were associated with a higher rate of natural pregnancy. Our data suggest that microsurgical vasoepididymostomy is an effective therapy of primary epididymal OA. Baseline lower FSH and higher TMSC were independent predictors for natural pregnancy occurrence.

  8. Comparison of two different extenders for cryopreservation of epididymal dog sperm.

    Science.gov (United States)

    Martins, Mim; Justino, R C; Sant'anna, M C; Trautwein, L G C; Souza, F F

    2012-12-01

    The collection of epididymal sperm is an option for preservation of germplasm of genetically superior animals that need to be orchiectomized or have died. The extender type used to freeze sperm is important to avoid spermatozoal membrane damage and to preserve semen quality after cryopreservation. The objective of this study was to verify the effects of a commercial bovine extender (Bovimix(®); Nutricell, Campinas) and a traditional TRIS-citric acid-glucose-egg yolk-7% glycerol extender on cryopreservation of canine epididymal sperm. The testes of 13 adult dogs were kept at 5 °C for 24 h in saline solution, and epididymal sperm was recovered in Ringers solution without lactate and were evaluated for motility. Samples with ≥ 80% motility were pooled and then divided before dilution and packaging in 0.5 ml plastic straws, equilibration at 4 °C for 1 h, freezing in nitrogen vapour for 20 min and storing at -196 °C. The straws were thawed at 56 °C for 10 s and were evaluated for motility by computer assisted analysis (CASA). The semen parameters, sperm movement index, linearity, total motility and rapid progressive motility were statistically higher in Bovimix(®) than TRIS. In contrast, amplitude of lateral head displacement, slow sperm and static sperm were lower in Bovimix(®). Despite the high percentage of sperm defects in epididymal cells, regardless of the extender, we concluded that Bovimix(®) is a viable alternative for the freezing of canine epididymal sperm. © 2012 Blackwell Verlag GmbH.

  9. Morphological studies on the seasonal changes in the epididymal duct of the one-humped camel (camelus dromedarius

    Directory of Open Access Journals (Sweden)

    Ahmed El-Zuhry Zayed

    2012-02-01

    Full Text Available The present work was carried out on 20 testes and epididymis of sexually mature camels to elucidate the gross anatomical, morphometerical, light microscopical and scanning electron microscopical features of the epididymis in different seasons. Anatomically, the epididymal duct of a camel consists of three parts head, body and tail. Histomorphologically, the epididymal duct is subdivided into initial, middle and terminal segments, of which the middle segment is further subdivided into proximal, intermediate and distal parts. There is a gradual decrease in the epithelial height of the epididymal duct from the initial to the terminal segments. This mechanically facilities passage of the sperms toward the terminal segment. High epithelium in the initial segment may indicate a more absorptive power of the epithelium in this segment. The seasonal reproductivety of the epididymal duct in the camel expressed by variations in the weight and volume of the epididymis, total diameter of the epididymal duct, epithelial height, length of the stereocilia, thickness of the muscular coat and cellular distributions in different segments. The spring months offer ideal circumstances for maximal reproductive activity in this species. The cellular components of the epididymal duct epithelium of the camel displays important morphological changes from season to another showing signs of increasing activity during spring in comparison to decreasing activity in other seasons. PAS positive granules are demonstrated in different segments of the epididymal duct and intraepithelial glands in different seasons. These granules are relatively more numerous in spring. The lamina propria surrounding the epididymal duct contains a layer of the elastic fibers which is very thick in winter, thick in spring and thin in other seasons. This increase in thickness of the elastic fibers predisposes for the increase in the total diameter of the epididymal duct in spring. It was conclude

  10. MOLECULAR ANALYSIS OF HUMAN SPERMATOZOA: POTENTIAL FOR INFERTILITY RESEARCH

    Science.gov (United States)

    Gordon Research Conference: Mammalian Gametogenesis and Embryogenesis New London, CT, July 1-6, 2000Molecular Analysis of Human Spermatozoa: Potential for Infertility ResearchDavid Miller 1, David Dix2, Robert Reid 3, Stephen A Krawetz 3 1Reproductive ...

  11. Cryopreservation of spermatozoa of black marlin, Makaira indica (Teleostei: Istiophoridae).

    Science.gov (United States)

    van der Straten, K M; Leung, L K-P; Rossini, R; Johnston, S D

    2006-01-01

    As a first step towards the development of a method for the cryopreservation of black marlin spermatozoa, this study investigated the effect of dimethylsulfoxide (DMSO) concentration and pellet size on post-thaw spermatozoal motility. Spermatozoa were recovered from the spermatic duct of testes retrieved post-mortem from four adult black marlin caught in the Coral Sea spawning grounds of Australia. Undiluted spermatozoa were stored on ice for 4 to 10 hours during transport to shore, then evaluated for motility after activation in seawater (1:10 v:v). Spermatozoa were prepared for cryopreservation in pellets by extension (1:3 v:v) in a defined fish Ringer's solution to give two final DMSO concentrations of 2.5% or 5.0%. Diluted spermatozoa were frozen directly on a dry ice block in pellet sizes of either 0.25 ml or 0.50 ml. Frozen pellets were thawed in a water bath at 40 degrees C for 60 seconds and assessed for post-thaw motility following activation in seawater. Spermatozoa recovered within 50 minutes of death and chilled on ice for 4 to 10 hours showed a mean (+/- SEM) motility immediately following activation of 91.6 +/- 7.9%. 50% of the spermatozoa remained motile for approximately 4 to 5 minutes. Following cryopreservation, mean motility declined significantly across all cryoprotectant and pellet size combinations (P < 0.001) but spermatozoa frozen in 2.5% DMSO showed higher motility than those frozen in 5.0% DMSO (P = 0.014). Pellet size had no effect on post-thaw motility (P = 0.179).

  12. Influence of Macrophages on the Rooster Spermatozoa Quality.

    Science.gov (United States)

    Kuzelova, L; Vasicek, J; Chrenek, P

    2015-08-01

    The goal of this study was to evaluate the occurrence of macrophages in rooster semen and to investigate their impact on the spermatozoa quality. Ross 308 breeder males (n = 30) with no evidence of genital tract infections were used to determine the concentration of macrophages using fluorescently conjugated acetylated low-density lipoprotein (AcLDL). Subsequently, the roosters were divided into two groups on the basis of semen macrophage concentration, and semen quality was compared in two heterospermic samples. We applied computer-assisted semen analysis (CASA) system to determine motility parameters. Fluorescence microscopy and flow cytometry were used to evaluate occurrence of apoptotic and dead spermatozoa. Spermatozoa fertility potential was examined after intravaginal artificial insemination of hens. Eighteen roosters (control group) contained 0.2-3% of macrophages within spermatozoa population and ten roosters (macrophage group) had 10-15% of macrophages. Males from macrophage group had lower (p rooster semen may have a negative effect on some parameters of rooster spermatozoa evaluated in vitro. Furthermore, our study suggests that flow cytometry allows more precise examination of spermatozoa viability and apoptosis in a very short time compared with the fluorescent microscopy. © 2015 Blackwell Verlag GmbH.

  13. Effects of spermatozoa-oviductal cell coincubation time and oviductal cell age on spermatozoa-oviduct interactions.

    Science.gov (United States)

    Aldarmahi, Ahmed; Elliott, Sarah; Russell, Jean; Fazeli, Alireza

    2014-01-01

    The oviduct plays a crucial role in sperm storage, maintenance of sperm viability and sperm transport to the site of fertilisation. The aim of the present study was to investigate the effects of oviductal cell culture passage number, oviductal cell age and spermatozoa-oviduct coincubation times on gene expression in oviductal cells. Immortalised oviductal epithelial cells (OPEC) obtained from two different cell passages (36 and 57) were subcultured three times with and without spermatozoa for 24 h (control group). In a second study, OPEC were cocultured with spermatozoa for different time intervals (0, 4, 12 and 24 h). Expression of adrenomedullin (ADM), heat shock 70 kDa protein 8 (HSPA8) and prostaglandin E synthase (PGES) in OPEC was measured by quantitative polymerase chain reaction. The expression of ADM and HSPA8 was decreased significantly in OPEC cells from Passage 57, particularly in the later subculture group. These effects on HSPA8, but not ADM, expression in OPEC were further altered after coculture with spermatozoa for 24 h. We also demonstrated that spermatozoa-oviduct coculture for 12 and 24 h resulted in significantly higher expression of ADM, HSPA8 and PGES in OPEC. Overall, the data suggest that the OPEC lose some of their properties as a result of oviductal cell aging and that there are spermatozoa-oviduct interactions leading to increased oviductal cell gene expression.

  14. Ion transporters for fluid reabsorption in the rooster (Gallus domesticus) epididymal region.

    Science.gov (United States)

    Bahr, J M; Dalponte, M; Janssen, S; Bunick, D; Nakai, M

    2006-10-01

    Testicular fluid is highly condensed during its passage through the epididymal region in the avian species. In the present study, major ion transporters that are responsible for condensation mainly by water resorption in the reproductive tract as identified in the mammalian epididymis were localized within the rooster (Gallus domesticus) epididymis by immunohistochemistry. The results show that the efferent ductule epithelium expressed sodium-potassium ATPase (Na(+),K(+)-ATPase), carbonic anhydrase II (CAII) and sodium hydrogen exchanger isoform 3 (NHE3) and that the connecting ductule and epididymal duct epithelia expressed Na(+),K(+)-ATPase and CAII. These data suggest that a model proposed for reabsorption in mammalian efferent ductules can be applied to avian efferent ductules.

  15. Influence of recovery method and centrifugation on epididymal sperm from collared peccaries (Pecari tajacu Linnaeus, 1758).

    Science.gov (United States)

    Bezerra, José Artur Brilhante; da Silva, Andréia Maria; Peixoto, Gislayne Christianne Xavier; da Silva, Mariana de Araújo; Franco de Oliveira, Moacir; Silva, Alexandre Rodrigues

    2014-05-01

    In order to establish protocols for gamete recovery from accidentally killed wild animals, or to take advantage of those slaughtered by captive breeders, we assess the influence of two methods on the recovery of epididymal sperm from collared peccaries, and verify the effect of centrifugation on such gametes. Genitalia from nine animals were used. For each animal, one epididymis was processed by flotation and the other was processed by retrograde flushing, both using a buffered media based on Tris. Following recovery, sperm were evaluated for motility, vigor, viability, functional membrane integrity, and morphology. A 1-mL aliquot of each sample was centrifuged, the supernatant removed, and the pellet suspended and evaluated as fresh samples. The sperm characteristics did not differ between the samples collected by flotation or retrograde flushing (P methods was affected by centrifugation. In conclusion, epididymal sperm from collared peccaries can be efficiently collected through flotation or retrograde flushing, but not when either is followed by centrifugation.

  16. Morphology of the testes and epididymal ducts in the pampas cat Leopardus colocolo (Molina, 1782

    Directory of Open Access Journals (Sweden)

    Mahmoud Mehanna

    Full Text Available ABSTRACT: The pampas cat Leopardus colocolo (Molina, 1782 is a species of the Felidae family, widely distributed in South America, included on CITES Appendix II and classified as Near Threatened on the IUCN Red List, with population trend decreasing. Based on this information, the objective of this study is to describe morphologically the testes and epididymal ducts of pampas cat. The animal, coming from the Federal University of Mato Grosso Zoo, Brazil, had died after anesthesia procedure and the male reproductive system was dissected to collect the testicles. The samples taken were fragmented and histologically examined. From the microscopic analysis of the testes were identified: vaginal and tunica albuginea, formed by dense connective tissue modeled with large amount of collagen fibers. The tunica albuginea fibrous septa emits into the body. The seminiferous tubules are coiled and coated internally by spermatogenic epithelium consisting of Sertoli cells, surrounded by a basement membrane in the presence of myoid cells. The interstitial tissue between the seminiferous tubules, is composed of loose connective tissue, blood and lymph vessels, and Leydig cells in polyhedral shape. The epididymal ducts showed pseudostratified columnar epithelium with secretory cells of which stereocilia design, situated on a basement membrane filled by myoid cells. This epithelium has principal and basal cells, the main cell design stereocilia toward the lumen of the epididymal duct.

  17. Static magnetic field reduced exogenous oligonucleotide uptake by spermatozoa using magnetic nanoparticle gene delivery system

    Science.gov (United States)

    Katebi, Samira; Esmaeili, Abolghasem; Ghaedi, Kamran

    2016-03-01

    Spermatozoa could introduce exogenous oligonucleotides of interest to the oocyte. The most important reason of low efficiency of sperm mediated gene transfer (SMGT) is low uptake of exogenous DNA by spermatozoa. The aim of this study was to evaluate the effects of static magnetic field on exogenous oligonucleotide uptake of spermatozoa using magnetofection method. Magnetic nanoparticles (MNPs) associated with the labeled oligonucleotides were used to increase the efficiency of exogenous oligonucleotide uptake by rooster spermatozoa. We used high-field/high-gradient magnet (NdFeB) to enhance and accelerate exogenous DNA sedimentation at the spermatozoa surface. Flow cytometry analysis was performed to measure viability and percentage of exogenous oligonucleotide uptake by sperm. Flow cytometry analysis showed a significant increase in exogenous oligonucleotide uptake by rooster spermatozoa (Prooster spermatozoa; however unlike others studies, static magnetic field, was not only ineffective to enhance exogenous oligonucleotide uptake by rooster spermatozoa but also led to reduction in efficiency of magnetic nanoparticles in gene transfer.

  18. Influence of heavy nanocrystals on spermatozoa and fertility of mammals

    Energy Technology Data Exchange (ETDEWEB)

    Akhavan, Omid, E-mail: oakhavan@sharif.edu [Department of Physics, Sharif University of Technology, P.O. Box 11155-9161, Tehran (Iran, Islamic Republic of); Institute for Nanoscience and Nanotechnology, Sharif University of Technology, P.O. Box 14588-89694, Tehran (Iran, Islamic Republic of); Hashemi, Ehsan [National Research Center for Transgenic Mouse, National Institute of Genetic Engineering and Biotechnology, P.O. Box 14965-161, Tehran (Iran, Islamic Republic of); Zare, Hakimeh [Physics Department, Yazd University, Yazd, P.O. Box 89195-741 (Iran, Islamic Republic of); Shamsara, Mehdi [National Research Center for Transgenic Mouse, National Institute of Genetic Engineering and Biotechnology, P.O. Box 14965-161, Tehran (Iran, Islamic Republic of); Taghavinia, Nima [Department of Physics, Sharif University of Technology, P.O. Box 11155-9161, Tehran (Iran, Islamic Republic of); Institute for Nanoscience and Nanotechnology, Sharif University of Technology, P.O. Box 14588-89694, Tehran (Iran, Islamic Republic of); Heidari, Farid [National Research Center for Transgenic Mouse, National Institute of Genetic Engineering and Biotechnology, P.O. Box 14965-161, Tehran (Iran, Islamic Republic of)

    2016-12-01

    In recent years, quantum dots (QDs) have been widely used in upcoming nanotechnology-based solar cells, light-emitting diodes and even bioimaging, due to their tunable optical properties and excellent quantum yields. But, such nanostructures are currently constituted by heavy elements which can threat the human health and living environment. Hence, in this work, the in vivo effects of CdTe nanocrystals (NCs) (as one of the promising QDs) on spermatozoa of male mice and subsequently on fertility of female mice were investigated, for the first time. To do this, CdTe NCs were synthesized through an environment-friendly (aqueous-based solution) method. The sperm cells presented a high potential for uptake of the heavy QDs. Meantime, the NCs exhibited concentration-dependent adverse effects on morphology, viability, kinetic characteristics and DNA of the spermatozoa. At low concentration of 0.1 μg/mL, the NCs showed a moderate toxicity (~ 25% reduction in viability and motility of the spermatozoa), while remarkable toxicities were observed at higher concentrations of 1.0–100 μg/mL (~ 67% reduction in viability and motility for 100 μg/mL). Furthermore, significant in vitro DNA fragmentation of the spermatozoa was observed at CdTe concentrations ≥ 10 μg/mL. In vivo toxicity of the NCs was found lower than the in vitro toxicity. Nevertheless, the in vivo destructive effects of the NCs still caused ~ 34% reduction in viability as well as motility and ~ 5% damages in DNA of male mice spermatozoa. These resulted in ~ 26% decrease in fertility and gestation of female mice, along with an overall hormone secretion during the pregnancy, and ~ 39% reduction in viability of pups/pregnant females. - Highlights: • The cytotoxic effects of CdTe nanocrystals on spermatozoa of male mice • High uptake of CdTe by spermatozoa, resulting in inactivation of spermatozoa or pollution of the others • The adverse effects of polluted spermatozoa on fertility/gestation of female mice

  19. Effect of cholesterol supplementation on cryosurvival of goat spermatozoa

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    Sunita Behera

    2015-12-01

    Full Text Available Aim: Sperm membrane cholesterol influences cryodamage during cryopreservation. The present study was carried out to evaluate the effect of varying cholesterol levels in Tris based extenders on the freezability of sexually healthy Malabari buck semen. Materials and Methods: A total of 48 ejaculates from two adults healthy sexually healthy Malabari bucks were utilized for the study. The collected and pooled ejaculates were divided into four groups with Group I serving as Control - I, Group II and III were treated with 1 mg and 2 mg of cholesterol-loaded-cyclodextrin (CLC/120 × 106 spermatozoa, respectively, and Group IV treated with 1 mg methyl-β-cyclodextrin (MβCD served as Control - II. Manual freezing was carried out to cryopreserve the treated and control spermatozoa. Results: Treatment of semen samples with CLC resulted in improved maintenance of sperm motility at pre-freeze and post-thaw stages of cryopreservation without affecting hypo-osmotic swelling response. Treatment of semen with 1 mg of CLC/120 × 106 spermatozoa was observed to be better than treatment with 2 mg of CLC/120 × 106 spermatozoa. In general, MβCD treatment was found to result in significantly lower sperm characteristics than those of Control - I and CLC treatment at pre-feeze and post-thaw stages and when incubated up to 4 h. Conclusion: Cholesterol treatment of sexually healthy Malabari buck semen was found to hold promise for improving cryopreser-vability of spermatozoa.

  20. Cytogenetic effects of in vitro irradiation of human spermatozoa

    International Nuclear Information System (INIS)

    Miro, R.; Genesca, A.; Alvarez, R.; Tusell, L.; Ponsa, I.

    1997-01-01

    The effects of human mutagens, clastogens and aneugens have been studied almost exclusively in somatic tissues. However, currently there is a considerable discussion about the potential of ionizing radiation to induce heritable germ cell mutations. While the various viewpoints remain controversial. One of the aims of germ cell cytogenetic studies must be to improve the ability to identify and estimate the actual genetic risk in humans. One way to assess the risk of transmission of genetic anomalies by men occupationally or accidentally exposed to ionizing radiation is to determine whether there is a dose-related genetic damage in human spermatozoa. Cytogenetic analysis of human spermatozoa is possible after interspecific in vitro fertilization between zona pellucida-free hamster oocytes and human spermatozoa. Using this assay system we have analyzed the radiation induction of structural chromosome abnormalities in sperm derived complements at the first embryo cleavage, as well as the radiation induction of micronuclei and aneuploidy in two-cell hybrid embryos. (author)

  1. Non-genomic effects of vitamin D in human spermatozoa

    DEFF Research Database (Denmark)

    Blomberg Jensen, Martin; Dissing, Steen

    2012-01-01

    The spectrum for vitamin D (VD) mediated effects has expanded in recent years. Activated VD (1,25(OH)(2)D(3)) binds to the VD receptor (VDR) and mediates non-genomic effects through the alternative ligand binding-pocket (VDR-ap) or regulates gene transcription through the genomic binding......-pocket. VDR and VD-metabolizing enzymes are expressed in human testis, male reproductive tract and mature spermatozoa, and VD is considered important for male reproduction. Expression of the VD-inactivating enzyme CYP24A1 at the annulus of human spermatozoa distinguish normal and infertile men with high...... specificity, and CYP24A1 expression is positively correlated with all semen variables and suggested as a marker for both semen quality and VD responsiveness. Moreover, spermatozoa are transcriptionally silent and are therefore a unique model to study non-genomic effects. 1,25(OH)(2)D(3) induced a rapid...

  2. Techniques for augmentation of exogenous DNA uptake by ovine spermatozoa

    Science.gov (United States)

    Hoseini Pajooh, K.; Tajik, P.; Karimipoor, M.; Behdani, M.

    2016-01-01

    Sperm mediated gene transfer can be an inexpensive and simple method in animal transgenesis; however its efficiency is poor, mainly due to the spermatozoa’s lesser uptake of exogenous DNA. In the present study, the effects of lipofection and other augmentation techniques, such as sperm freezing and spermatozoa treatment with triton X100 and DMSO, on exogenous DNA uptake by sheep spermatozoa and motility of sperms with plasmid uptake were evaluated. In the first experiment, ram sperms were incubated with a complex of rhodamine labeled plasmid (p-EGFP) and Lipofectamine 2000TM. In the second, spermatozoa were treated with Triton X-100TM or DMSO or were frozen without cryoprotectant. The results indicated that there was no significant difference (Plipofected sperms with 300 and 600 ng of plasmid in comparison with control group, i.e. transfected without lipofectamine. Furthermore, lipofection could not improve sperm motility during true plasmid uptake. Almost all of triton X100 treated and frozen-thawed spermatozoa had absorbed foreign DNA, though all were immotile. In spermatozoa treated with 0.1% DMSO, plasmid absorption rate (69.40%) was significantly higher (P<0.05) than untreated spermatozoa (57.80%), but sperm motility was not significantly different from control group. In conclusion, lipofectamine® 2000 could neither improve transfection rate, nor support motility in transfected sperms. The methods inducing membrane disruption like, freeze-thaw and triton X100 treatment, can be used in ICSI-sperm mediated gene transfer without the need for sperm selection, provided that they cause no damage to sperm nucleus. PMID:27656225

  3. EFFECT OF DIFFERENT DILUENTS ON ROOSTER SPERMATOZOA APOPTOSIS

    Directory of Open Access Journals (Sweden)

    Lenka Kuželová

    2013-02-01

    Full Text Available The aim of this study was to observe the apoptosis of rooster spermatozoa and to compare the effect of commercial insemination diluent and saline solution on rooster spermatozoa apoptosis. Semen samples were collected once a week from roosters lines Lohmann Light (n=30. The one heterospermic sample was diluted with saline solution at a ratio of 1:100 and the second was diluted with commercial insemination diluent (Avian diluents; IMV Technologies, France in the same ratio 1:100 at room temperature. The percentage of apoptotic spermatozoa was estimated by fluorescent staining (Yo-Pro-1 0.5, 1 and 2 hours after semen collection and analyzed by fluorescent microscopy. There were no significant differences in sperms’ membranes quality after half an hour and one hour between diluted semen in both heterospermic samples. Significantly higher rate (P<0.05 of Yo-Pro-1 – positive cells was observed in sample diluted in commercial diluents (22.03% vs. 35.91% of apoptotic spermatozoa after two hours. We concluded that rooster semen could be diluted by saline solution which is cheaper and more accessible for practical The aim of this study was to observe the apoptosis of rooster spermatozoa and to compare the effect of commercial insemination diluent and saline solution on rooster spermatozoa apoptosis. Semen samples were collected once a week from roosters lines Lohmann Light (n=30. The one heterospermic sample was diluted with saline solution at a ratio of 1:100 and the second was diluted with commercial insemination diluent (Avian diluents; IMV Technologies, France in the same ratio 1:100 at room temperature. The percentage of apoptotic spermatozoa was estimated by fluorescent staining (Yo-Pro-1 0.5, 1 and 2 hours after semen collection and analyzed by fluorescent microscopy. There were no significant differences in sperms’ membranes quality after half an hour and one hour between diluted semen in both heterospermic samples. Significantly higher

  4. In vivo oxidative stress alters thiol redox status of peroxiredoxin 1 and 6 and impairs rat sperm quality

    Directory of Open Access Journals (Sweden)

    Yannan Liu

    2017-01-01

    Full Text Available Oxidative stress, the imbalance between the production of reactive oxygen species (ROS and antioxidant activity is a major culprit of male infertility. Peroxiredoxins (PRDXs are major antioxidant enzymes of mammalian spermatozoa and are thiol oxidized and inactivated by ROS in a dose-dependent manner. Their deficiency and/or inactivation have been associated with men infertility. The aim of this study was to elucidate the impact of oxidative stress, generated by the in vivo tert-butyl hydroperoxide (tert-BHP treatment on rat epididymal spermatozoa during their maturation process. Adult Sprague-Dawley males were treated with 300 μmoles tert-BHP/kg or saline (control per day intraperitoneal for 15 days. Lipid peroxidation (2-thibarbituric acid reactive substances assay, total amount and thiol oxidation of PRDXs along with the total amount of superoxide dismutase (SOD, motility and DNA oxidation (8-hydroxy-deoxyguanosine were determined in epididymal spermatozoa. Total amount of PRDXs and catalase and thiol oxidation of PRDXs were determined in caput and cauda epididymis. While animals were not affected by treatment, their epididymal spermatozoa have decreased motility, increased levels of DNA oxidation and lipid peroxidation along with increased PRDXs (and not SOD amounts. Moreover, sperm PRDXs were highly thiol oxidized. There was a differential regulation in the expression of PRDX1 and PRDX6 in the epididymis that suggests a segment-specific role for PRDXs. In conclusion, PRDXs are increased in epididymal spermatozoa in an attempt to fight against the oxidative stress generated by tert-BHP in the epididymis. These findings highlight the role of PRDXs in the protection of sperm function and DNA integrity during epididymal maturation.

  5. Effects of dietary polysaccharides from the submerged fermentation concentrate of Hericium caput-medusae (Bull.:Fr.) Pers. on fat deposition in broilers.

    Science.gov (United States)

    Shang, Hong Mei; Song, Hui; Shen, Si Jie; Yao, Xu; Wu, Bo; Wang, Li Na; Jiang, Yun Yao; Ding, Guo Dong

    2015-01-01

    The present study was conducted to investigate the lipid-lowering effect of polysaccharides from the submerged fermentation concentrate of Hericium caput-medusae (Bull.:Fr.) Pers. (HFCP) in broilers. A total of 480 female Arbor Acres broilers were randomly divided into four dietary treatments, each consisting of six pens as replicates, and fed diets containing 0 (control), 1, 3 or 5 g kg(-1) HFCP. The results revealed that the average daily gain of broilers increased (linear (L), P < 0.01; quadratic (Q), P < 0.01) when the HFCP levels increased. The serum cholesterol, triglyceride and low-density lipoprotein cholesterol levels decreased (Q, P < 0.05) while the high-density lipoprotein cholesterol level increased (Q, P < 0.05) when the HFCP levels increased. The caecum Escherichia coli count and pH decreased (Q, P < 0.01) while the lactobacilli count and bifidobacteria count increased (L, P < 0.05; Q, P < 0.05) when the HFCP levels increased. The propionic acid and butyric acid concentrations increased (L, P < 0.001; Q, P < 0.001) while the abdominal fat rate and liver fat content decreased (L, P < 0.01; Q, P < 0.05) when the HFCP levels increased. Dietary supplementation with HFCP may lead to the development of low abdominal fat of broilers as demanded by health-conscious consumers. © 2014 Society of Chemical Industry.

  6. Static magnetic field reduced exogenous oligonucleotide uptake by spermatozoa using magnetic nanoparticle gene delivery system

    International Nuclear Information System (INIS)

    Katebi, Samira; Esmaeili, Abolghasem; Ghaedi, Kamran

    2016-01-01

    Spermatozoa could introduce exogenous oligonucleotides of interest to the oocyte. The most important reason of low efficiency of sperm mediated gene transfer (SMGT) is low uptake of exogenous DNA by spermatozoa. The aim of this study was to evaluate the effects of static magnetic field on exogenous oligonucleotide uptake of spermatozoa using magnetofection method. Magnetic nanoparticles (MNPs) associated with the labeled oligonucleotides were used to increase the efficiency of exogenous oligonucleotide uptake by rooster spermatozoa. We used high-field/high-gradient magnet (NdFeB) to enhance and accelerate exogenous DNA sedimentation at the spermatozoa surface. Flow cytometry analysis was performed to measure viability and percentage of exogenous oligonucleotide uptake by sperm. Flow cytometry analysis showed a significant increase in exogenous oligonucleotide uptake by rooster spermatozoa (P<0.001) when spermatozoa were incubated in exogenous oligonucleotide solution and MNPs. However, by applying static magnetic field during magnetofection method, a significant decrease in exogenous oligonucleotide uptake was observed (P<0.05). Findings of this study showed that MNPs were effective to increase exogenous oligonucleotide uptake by rooster spermatozoa; however unlike others studies, static magnetic field, was not only ineffective to enhance exogenous oligonucleotide uptake by rooster spermatozoa but also led to reduction in efficiency of magnetic nanoparticles in gene transfer. - Highlights: • Core/shell type Iron oxide nanoparticles were used as a novel and efficient method. • This method increases exogenous DNA uptake by rooster spermatozoa. • Static magnetic field decreased DNA uptake by rooster spermatozoa.

  7. Static magnetic field reduced exogenous oligonucleotide uptake by spermatozoa using magnetic nanoparticle gene delivery system

    Energy Technology Data Exchange (ETDEWEB)

    Katebi, Samira; Esmaeili, Abolghasem, E-mail: aesmaeili@sci.ui.ac.ir; Ghaedi, Kamran

    2016-03-15

    Spermatozoa could introduce exogenous oligonucleotides of interest to the oocyte. The most important reason of low efficiency of sperm mediated gene transfer (SMGT) is low uptake of exogenous DNA by spermatozoa. The aim of this study was to evaluate the effects of static magnetic field on exogenous oligonucleotide uptake of spermatozoa using magnetofection method. Magnetic nanoparticles (MNPs) associated with the labeled oligonucleotides were used to increase the efficiency of exogenous oligonucleotide uptake by rooster spermatozoa. We used high-field/high-gradient magnet (NdFeB) to enhance and accelerate exogenous DNA sedimentation at the spermatozoa surface. Flow cytometry analysis was performed to measure viability and percentage of exogenous oligonucleotide uptake by sperm. Flow cytometry analysis showed a significant increase in exogenous oligonucleotide uptake by rooster spermatozoa (P<0.001) when spermatozoa were incubated in exogenous oligonucleotide solution and MNPs. However, by applying static magnetic field during magnetofection method, a significant decrease in exogenous oligonucleotide uptake was observed (P<0.05). Findings of this study showed that MNPs were effective to increase exogenous oligonucleotide uptake by rooster spermatozoa; however unlike others studies, static magnetic field, was not only ineffective to enhance exogenous oligonucleotide uptake by rooster spermatozoa but also led to reduction in efficiency of magnetic nanoparticles in gene transfer. - Highlights: • Core/shell type Iron oxide nanoparticles were used as a novel and efficient method. • This method increases exogenous DNA uptake by rooster spermatozoa. • Static magnetic field decreased DNA uptake by rooster spermatozoa.

  8. Birefringence characteristics in sperm heads allow for the selection of reacted spermatozoa for intracytoplasmic sperm injection.

    Science.gov (United States)

    Gianaroli, Luca; Magli, M Cristina; Ferraretti, Anna P; Crippa, Andor; Lappi, Michela; Capitani, Serena; Baccetti, Baccio

    2010-02-01

    To verify clinical outcome after injection of spermatozoa that have undergone the acrosome reaction (reacted spermatozoa) vs. those still having an intact acrosome (nonreacted spermatozoa). Prospective, randomized study. Reproductive Medicine Unit, Italian Society for the Study of Reproductive Medicine, Bologna, Italy. According to a prospective randomization including 71 couples with severe male factor infertility, intracytoplasmic sperm injection (ICSI) was performed under polarized light that permitted analysis of the pattern of birefringence in the sperm head. Twenty-three patients had their oocytes injected with reacted spermatozoa, 26 patient's oocytes were injected with nonreacted spermatozoa, and in 22 patients both reacted and nonreacted spermatozoa were injected. Intracytoplasmic sperm injection was performed under polarized light to selectively inject acrosome-reacted and acrosome-nonreacted spermatozoa. Rates of fertilization, cleavage, pregnancy, implantation, and ongoing implantation. There was no effect on the fertilizing capacity and embryo development of either type of sperm, whereas the implantation rate was higher in oocytes injected with reacted spermatozoa (39.0%) vs. those injected with nonreacted spermatozoa (8.6%). The implantation rate was 24.4% in the group injected with both reacted and nonreacted spermatozoa. The delivery rate per cycle followed the same trend. Spermatozoa that have undergone the acrosome reaction seem to be more prone to supporting the development of viable ICSI embryos. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  9. Progression of Epididymal Maldevelopment Into Hamartoma-like Neoplasia in VHL Disease

    Directory of Open Access Journals (Sweden)

    Gautam U. Mehta

    2008-10-01

    Full Text Available Inactivation of the von Hippel-Lindau (VHL gene and activation of the hypoxia-inducible factor (HIF in susceptible cells precedes formation of tumorlets and frank tumor in the epididymis of male VHL patients. We performed detailed histologic and molecular pathologic analysis of tumor-free epididymal tissues from VHL patients to obtain further insight into early epididymal tumorigenesis. Four epididymides from two VHL patients were serially sectioned to allow for three-dimensional visualization of morphologic changes. Areas of interest were genetically analyzed by tissue microdissection, immunohistochemistry for HIF and markers for mesonephric differentiation, and in situ hybridization for HIF downstream target vascular endothelial growth factor. Structural analysis of the epididymides revealed marked deviations from the regular anatomic structure resulting from impaired organogenesis. Selected efferent ductules were represented by disorganized mesonephric cells, and the maldeveloped mesonephric material was VHL-deficient by allelic deletion analysis. Furthermore, we observed maldeveloped mesonephric material near cystic structures, which were also VHL-deficient and were apparent derivatives of maldeveloped material. Finally, a subset of VHL-deficient cells was structurally integrated in regular efferent ductules; proliferation of intraductular VHL-deficient cells manifests itself as papillary growth into the ductular lumen. Furthermore, we clarify that that there is a pathogenetic continuum between microscopic tumorlets and formation of tumor. In multiple locations, three-dimensional reconstruction revealed papillary growth to extend deeply into ductular lumina, indicative of progression into early hamartoma-like neoplasia. We conclude epididymal tumorigenesis in VHL disease to occur in two distinct sequential steps: maldevelopment of VHL-deficient mesonephric cells, followed by neoplastic papillary proliferation.

  10. Predictors of patency after two-stitch invagination vaso-epididymal anastomosis for idiopathic obstructive azoospermia

    Directory of Open Access Journals (Sweden)

    G Gautam

    2005-01-01

    Full Text Available Objectives: Anastomotic patency with return of sperm in the ejaculate following microsurgical vasoepididymostomy (VEA is not universal and may be delayed. The ability to predict the result of VEA based on preoperative or intra-operative parameters would enable the surgeon to offer the best treatment to the infertile couple. We used the two-stitch invagination technique of VEA in patients of idiopathic obstructive azoospermia and prospectively analyzed factors that could predict a patent anastomosis. While such studies have previously been done for patients undergoing VEA for secondary infertility following a vasectomy, to the best of our knowledge this is the first study analyzing these parameters for patients with primary infertility and idiopathic obstruction. Methods and materials: Over a 2-year period, 29 men underwent the 2-suture invagination VEA for idiopathic obstructive azoospermia. Twenty-four patients provided at least one postoperative semen sample. Preoperative and intra-operative parameters were compared between patients with a patent anastomosis with sperm in ejaculate (n = 12 and those with no sperm in the ejaculate (n = 12 using the t-test, Fisher′s exact test or chi-square test, as appropriate and a multivariate statistical analysis to determine any significant difference. Results: The mean follow up of the 24 patients was 7.6 months (2-30 months. A significantly greater number of patients with patent anastomosis had motile epididymal sperms (P = 0.034 and higher surgeon′s technical satisfaction with the procedure (P = 0.034. However, this difference was seen only on a univariate analysis and did not persist when a multivariate analysis was used. Conclusions: The presence of motile sperms in the epididymal fluid and a high level of technical satisfaction with the anastomosis may indicate a higher likelihood of success following a vaso-epididymal anastomosis for idiopathic obstruction. However, these parameters are not

  11. Single-layer centrifugation through colloid selects improved quality of epididymal cat sperm.

    Science.gov (United States)

    Chatdarong, K; Thuwanut, P; Morrell, J M

    2010-06-01

    The objectives were to determine the: 1) extent of epithelial and red blood cell contamination in epididymal cat sperm samples recovered by the cutting method; 2) efficacy of simple washing, single-layer centrifugation (SLC), and swim-up for selecting epididymal cat sperm; and 3) effects of freezing and thawing on cat sperm selected by various techniques. Ten unit samples were studied; each contained sperm from the cauda epididymides of four cats (total, approximately 200 x 10(6) sperm) and was equally allocated into four treatments: 1) simple washing, 2) single-layer centrifugation through colloid prior to cryopreservation (SLC-PC), 3) single-layer centrifugation through colloid after cryopreservation (SLC-AC), and 4) swim-up. Centrifugation (300 x g for 20 min) was done for all methods. The SLC-PC had a better recovery rate than the SLC-AC and swim-up methods (mean+/-SD of 16.4+/-8.7, 10.7+/-8.9, and 2.3+/-1.7%, respectively; Pblood cell contamination than simple washed samples (0.02+/-0.01, 0.02+/-0.04, 0.03+/-0.04, and 0.44+/-0.22 x 10(6) cells/mL, respectively; P0.05), SLC-PC yielded the highest percentage of sperm with normal midpieces and tails (P0.05). In conclusion, both SLC-PC and swim-up improved the quality of epididymal cat sperm, including better morphology, membrane and DNA integrity, and removal of cellular contamination. However, SLC had a better sperm recovery rate than swim-up. 2010 Elsevier Inc. All rights reserved.

  12. CHARACTERISTICS AND FERTILITY OF SUMATRAN TIGER SPERMATOZOA CRYOPRESERVED WITH DIFFERENT SUGARS.

    Science.gov (United States)

    Wayan Kurniani Karja, Ni; Fahrudin, Mokhamad; Setiadi, Mohamad Agus; Tumbelaka, Ligaya Ita; Sudarwati, Retno; Hastuti, Yohana Tri; Mulia, Bongot Huas; Widianti, Ardyta; Sultan, Keni; Terazono, Tsukasa; Namula, Zhao; Taniguchi, Masayasu; Tanihara, Fuminori; Takemoto, Tatsuya; Kikuchi, Kazuhiro; Sato, Yoko; Otoi, Takeshige

    Cryopreservation of semen is one of the most important methods for the preservation of endangered tigers. This study evaluated the effects of sugar supplementation on the cryosurvival of spermatozoa from Sumatran tigers (Panthera tigris sumatrae). The post-thaw characteristics and fertility of spermatozoa cryopreserved with different sugars (glucose, lactose, and trehalose) were evaluated using heterologous in-vitro fertilisation with cat oocytes. All parameters of post-thaw spermatozoa significantly decreased as compared with those of fresh spermatozoa. The index of sperm motility for semen cryopreserved with lactose was significantly higher than that for semen cryopreserved with trehalose. The percentage of total fertilisation for tiger spermatozoa cryopreserved with trehalose was significantly lower than that for control cat spermatozoa. Our findings indicated that supplementation with lactose or glycerol as the main sugar in the egg yolk extender resulted in a better motility and fertility potential for post-thawed spermatozoa.

  13. Impact of 5'-amp-activated Protein Kinase on Male Gonad and Spermatozoa Functions.

    Science.gov (United States)

    Nguyen, Thi Mong Diep

    2017-01-01

    As we already know, the male reproductive system requires less energetic investment than the female one. Nevertheless, energy balance is an important feature for spermatozoa production in the testis and for spermatozoa properties after ejaculation. The 5'-AMP-activated protein kinase, AMPK, is a sensor of cell energy, that regulates many metabolic pathways and that has been recently shown to control spermatozoa quality and functions. It is indeed involved in the regulation of spermatozoa quality through its action on the proliferation of testicular somatic cells (Sertoli and Leydig), on spermatozoa motility and acrosome reaction. It also favors spermatozoa quality through the management of lipid peroxidation and antioxidant enzymes. I review here the most recent data available on the roles of AMPK in vertebrate spermatozoa functions.

  14. Uniflagellate spermatozoa in Nemertoderma (Turbellaria) and their phylogenetic significance.

    Science.gov (United States)

    Tyler, S; Rieger, R M

    1975-05-16

    An ultrastructural study of Nemertoderma (Turbellaria, Nemertodermatida) has revealed that its spermatozoa have only a single falgellum. This is the first uniflagellate spermatozoon known in the Turbellaria; it is indicative of the primitiveness of Nemertoderma and is evidence in support of the view that the Turbellaria as a whole are among the most primitive living Bilateria.

  15. The effect of sperm morphology and testicular spermatozoa ...

    African Journals Online (AJOL)

    Objective. To determine the correlation between sperm morphology groups (strict criteria) and testicular spermatozoa, and day 2 and 3 embryo quality in intracytoplasmic sperm injection (ICSI) and in vitro fertilisation (IVF) cases. Methods. A retrospective study was done of 2 402 IVF and ICSI-fertilised embryos classified as ...

  16. Ekstrak Biji Klabet Menurunkan Jumlah Sel Spermatozoa pada Kelinci

    Directory of Open Access Journals (Sweden)

    I Gusti Nyoman Sri Wiryawan

    2009-06-01

    Full Text Available Fenugreek seed (Trigonella foenum-graecum contains saponin diosgenin, wich has an antifertility effect on spermatozoa so it can be used as an oral contraceptive drug. This study was aimedto investigate the effect of fenugreek seed extract to spermatogenic process of rabbit, especially onviability spermatozoa. “Completely randomized control group post-test only design” was used inthis study. The animals were divided into four groups; one control group and three treatmentgroups with six replicates (P0 = control group; P1 = group were given 10 % fenugreek seed extract,1 cc/day; P2 = group were given 20 % fenugreek seed extract, 1 cc/day; P3 = group were given 30 %fenugreek seed extract, 1 cc/day. The extract was given orally once a day in 50 days. After treatment,testicles were sectioned and stained with Hematoxylin Eosin; for qualitative and quantitativemicroscopic analysis. The result of this study showed that the number of spermatozoa were decreasedsignificantly (p<0,05 after receiving 10% fenugreek seed extract 1 cc per day. In conclusion,fenugreek seed extract could reduce the number of spermatozoa.

  17. The Role of Oviductal Cells in Activating Stallion Spermatozoa

    NARCIS (Netherlands)

    Leemans, Bart; Gadella, Bart M.; Stout, Tom A.E.; De Schauwer, Catharina; Nelis, Hilde; Hoogewijs, Maarten; Van Soom, Ann

    2016-01-01

    Conventional in vitro fertilization is poorly successful with equine gametes. Although stallion spermatozoa bind to the zona pellucida in vitro, they fail to acrosome react and cannot therefore penetrate into the perivitelline space. Failed sperm penetration most likely relates to the absence in in

  18. Cryosurvival of goat spermatozoa in tris-egg yolk extender ...

    African Journals Online (AJOL)

    The effect of melatonin supplementation in semen extenders on cryosurvival of spermatozoa obtained from West African Dwarf (WAD) goat bucks was studied. Tris-egg yolk extenders supplemented with different levels of melatonin (0, 2, 4, 6 and 8 mM) were diluted with semen samples. The diluted semen samples were ...

  19. Coitus Interruptus: Are there spermatozoa in the pre-ejaculate ...

    African Journals Online (AJOL)

    Methods: A literature search was performed using PubMed, and Google Scholar search engines. Literature reviewed included reviews, and original articles that evaluated the presence of spermatozoa in the pre-ejaculatory fluid. Articles reporting about coitus interruptus as a method of contraception were also reviewed.

  20. The cryoprotective effect of trehalose supplementation on boar spermatozoa quality.

    Science.gov (United States)

    Hu, Jian-Hong; Li, Qing-Wang; Li, Gang; Jiang, Zhong-Liang; Bu, Shu-hai; Yang, Hai; Wang, Li-Qiang

    2009-05-01

    In order to improve boar sperm quality during frozen-thawed process, the influence of the presence of trehalose on success of cryopreservation of boar sperm were investigated. We evaluated freeze-thawing tolerance of boar spermatozoa in a base cooling extender with the addition of different trehalose concentrations (0, 25, 50, 100 and 200mmol/l), and tried to determine the optimum concentration of trehalose. We chose sperm motility, acrosome integrity, membrane integrity and cryocapacitation as parameters to evaluate cryopreservation capacity of boar spermatozoa. We obtained the best results for 100mmol/l trehalose-supplemented extenders, with values of 49.89% for motility, 66.52% for acrosome integrity and 44.61% for membrane integrity, while freeze-thawing tolerance was diminished significantly for 200mmol/l of trehalose. Before and after capacitation, the CTC score for semen diluted by extender containing 100mmol/l trehalose was 3.68% and 43.82%, respectively. In conclusion, trehalose could confer a greater cryoprotective capacity to boar spermatozoa. Trehalose-supplementation with 100mmol/l concentration in basic extender could significantly improve sperm motility, membrane integrity and acrosome integrity parameters, and reduce boar spermatozoa cryocapacitation during the cryopreservation process.

  1. Cholangiocarcinoma presenting as a solitary epididymal metastasis: a case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Bailey David M

    2007-08-01

    Full Text Available Abstract Background Solid tumor metastasis to the epididymis is a rare occurrence and is mostly discovered incidentally at autopsy or after therapeutic orchidectomy for prostate cancer. Other primary carcinomas that have been demonstrated to metastasize to the paratesticular region include those originating in the stomach, kidney, ileum, and colon. Case presentation A 72-year-old gentleman presented with a firm and tender mass involving the right epididymis. On examination, he was jaundiced. Computed tomography of the abdomen demonstrated an obstructive stricture of the extra-hepatic bile ducts, in keeping with a cholangiocarcinoma, through which a metal stent was endoscopically inserted for symptomatic relief. Subsequent right radical orchidectomy yielded a diffusely infiltrative adenocarcinoma obliterating the epididymis, extending into the rete testis, vas deferens and spermatic cord and showing widespread vascular and perineural invasion. Residual epididymal, rete, and testicular tubules showed no in situ neoplasia. Morphologically and immunohistochemically the features were in keeping with a metastasis from a primary cholangiocarcinoma. Conclusion Only two cases of bile duct carcinoma metastasising to the male genital tract have previously been reported in the literature, the testis being the main site of metastasis in both cases. To our knowledge, this is the first described case of cholangiocarcinoma metastasising primarily to the epididymis, and presenting as a solitary epididymal metastasis in the absence of disseminated disease. It serves to highlight the importance of performing a thorough examination of the male external genitalia both clinically, in the follow up of cancer patients, and at autopsy.

  2. Effect of Three Days Storage of Coated Spermatozoa at Cooling and Adding Seminal Plasma on Ram Fertility

    Directory of Open Access Journals (Sweden)

    Alireza Vaferi

    2016-08-01

    Full Text Available Introduction Artificial insemination (AI has only been used as a supplement to natural mating. AI, when used in conjunction with accurate progeny testing schemes, can substantially increase the rate of genetic progress compared with that of natural service. Moreover, the use of AI causes the limitation of the transmitted diseases. Cervical insemination with frozen-thawed ram semen has not been widely adopted, probably because of the relative poor fertility obtained. Thus using fresh and diluted semen is only approach for performing AI. AI is currently limited by the poor fertility achieved after cervical insemination with the storage of liquid semen at sub-ambient temperature. The success of this procedure in sheep is restricted by the short length of time that ram sperm can be stored in a liquid state. Moreover, the effect of cooling on sperm differs depending on species. It is also well known that ram spermatozoa are more sensitive to cold-shock stress than those of other species. Seminal plasma, as physiological secretion, is a complex mixture of secretions originating from testis, epididymis and accessory sex glands which is mixed with epididymal sperm at ejaculation; it serves as the carrier of sperm to the female genital tract. This mixture contains numerous factors such as organic and nonorganic material which play an important role in the final maturation of the spermatozoa through hormonal, enzymatic and surface-modifying events. During natural mating, a mechanism may be activated to separate spermatozoa from seminal plasma. After being ejaculated into the vagina, sperm swim through cervical mucus and enter the uterus within minutes (>30 min; cervical mucus acts as a barrier for seminal plasma. In the artificial insemination industry, seminal plasma with all the useful and harmful components is not removed from semen and is in contact with sperm throughout cooling, freezing and storage. On the other hand, it was demonstrated that the

  3. Effects of dietary fermentation concentrate of Hericium caput-medusae (Bull.:Fr.) Pers. on growth performance, digestibility, and intestinal microbiology and morphology in broiler chickens.

    Science.gov (United States)

    Shang, Hong Mei; Song, Hui; Xing, Ya Li; Niu, Shu Li; Ding, Guo Dong; Jiang, Yun Yao; Liang, Feng

    2016-01-15

    The present study was undertaken to investigate the effects of fermentation concentrate of Hericium caput-medusae (Bull.:Fr.) Pers. (HFC) on growth performance, digestibility, intestinal microbiology, and intestinal morphology in broiler chickens. A total of 600 male Arbor Acres broilers were randomly divided into five dietary treatments (20 broilers per pen with six pens per treatment): CON (basal diet), ANT (basal diet supplemented with 5 mg kg(-1) flavomycin) and HFC (basal diet supplemented with 6, 12, and 18 g kg(-1) HFC). The experimental lasted for 42 days. The results revealed that the average daily gain [linear (L), P < 0.01; quadratic (Q), P < 0.01] of broilers increased when the HFC levels increased during the starter (days 1-21), finisher (days 22-42), and the overall experiment period (days 1 to 42). In the small intestinal digesta and the caecum digesta, the Escherichia coli count (L, P < 0.05; Q, P < 0.001) decreased while the Lactobacilli count (L, P < 0.01; Q, P < 0.001) and Bifidobacteria count (L, P < 0.001; Q, P < 0.001) increased when the HFC levels increased. The crude protein digestibility of broilers (L, P < 0.01; Q, P < 0.001) increased when the HFC levels increased. In the duodenum, jejunum, and ileum of broilers, the villus height and villus height to crypt depth ratio (L, P < 0.001; Q, P < 0.001) increased when the HFC levels increased. Dietary supplementation with HFC increased gut Lactobacilli and Bifidobacteria counts and inhibited E. coli growth, improved nutrient utilisation and intestine villus structure, and thus improved the growth of broilers. © 2015 Society of Chemical Industry.

  4. Recovery and cryopreservation of epididymal sperm from agouti (Dasiprocta aguti) using powdered coconut water (ACP-109c) and Tris extenders.

    Science.gov (United States)

    Silva, M A; Peixoto, G C X; Santos, E A A; Castelo, T S; Oliveira, M F; Silva, A R

    2011-10-01

    The objective was to compare the use of powdered coconut water (ACP-109c; ACP Biotecnologia, Fortaleza, CE, Brazil) and Tris extenders for recovery and cryopreservation of epididymal sperm from agouti. The caudae epididymus and proximal ductus deferens from 10 sexually mature agoutis were subjected to retrograde washing using ACP-109c (ACP Biotecnologia) or Tris. Epididymal sperm were evaluated for motility, vigor, sperm viability, membrane integrity, and morphology. Samples were centrifuged, and extended in the same diluents plus egg yolk (20%) and glycerol (6%), frozen in liquid nitrogen, and subsequently thawed at 37°C for 1 min, followed by re-evaluation of sperm characteristics. The two extenders were similarly efficient for epididymal recovery, with regard to the number and quality of sperm recovered. However, for both extenders, sperm quality decreased (P Biotecnologia) group, which was significantly better than 9.7 ± 2.6% motile sperm with 1.2 ± 0.3 vigor in Tris. In conclusion, agouti epididymal sperm were successfully recovered using either ACP-109c (ACP Biotecnologia) or Tris extenders; however, ACP-109c (ACP Biotecnologia) was a significantly better extender for processing and cryopreserving these sperm. Copyright © 2011 Elsevier Inc. All rights reserved.

  5. Human spermatozoa: revelations on the road to conception

    OpenAIRE

    Aitken, R. John

    2013-01-01

    Human spermatozoa are highly complex specialized cells designed to survive a long and perilous journey from the site of insemination to the upper reaches of the female reproductive tract where fertilization occurs. During this journey, these cells have to run the gauntlet laid down by the female immune system and time their physiological maturation so that as soon as an egg appears in the Fallopian tube, they are equipped to recognize this cell and participate in a remarkable cascade of cellu...

  6. Human spermatozoa: revelations on the road to conception.

    Science.gov (United States)

    Aitken, R John

    2013-10-01

    Human spermatozoa are highly complex specialized cells designed to survive a long and perilous journey from the site of insemination to the upper reaches of the female reproductive tract where fertilization occurs. During this journey, these cells have to run the gauntlet laid down by the female immune system and time their physiological maturation so that as soon as an egg appears in the Fallopian tube, they are equipped to recognize this cell and participate in a remarkable cascade of cellular interactions culminating in fertilization. Despite their high level of specialization, human spermatozoa are notoriously inadequate and appear to be major contributors to the poor fertility that characterizes our species. Defective spermatozoa are also known to have a major impact on the progress of pregnancy and the health trajectory of the offspring, resulting in paternally mediated increases in miscarriage rate and a range of diseases in the progeny, including dominant genetic diseases and cancer. The causes of defective sperm function are complex and involve both genetic and environmental impacts, as well as paternal age. Where genetic factors are involved, there is a concern that the widespread use of assisted conception technologies will serve to enhance the retention of poor fertility genes in the population such that the more we use assisted reproductive technologies in one generation the more we shall need them in the next. These observations may have important implications for the health and well-being of children and for the provision of reproductive healthcare services for future generations.

  7. Current perspectives of CASA applications in diverse mammalian spermatozoa.

    Science.gov (United States)

    van der Horst, Gerhard; Maree, Liana; du Plessis, Stefan S

    2018-03-26

    Since the advent of computer-aided sperm analysis (CASA) some four decades ago, advances in computer technology and software algorithms have helped establish it as a research and diagnostic instrument for the analysis of spermatozoa. Despite mammalian spermatozoa being the most diverse cell type known, CASA is a great tool that has the capacity to provide rapid, reliable and objective quantitative assessment of sperm quality. This paper provides contemporary research findings illustrating the scientific and commercial applications of CASA and its ability to evaluate diverse mammalian spermatozoa (human, primates, rodents, domestic mammals, wildlife species) at both structural and functional levels. The potential of CASA to quantitatively measure essential aspects related to sperm subpopulations, hyperactivation, morphology and morphometry is also demonstrated. Furthermore, applications of CASA are provided for improved mammalian sperm quality assessment, evaluation of sperm functionality and the effect of different chemical substances or pathologies on sperm fertilising ability. It is clear that CASA has evolved significantly and is currently superior to many manual techniques in the research and clinical setting.

  8. Characterization of the testicular, epididymal and endocrine phenotypes in the Leuven Vdr-deficient mouse model

    DEFF Research Database (Denmark)

    Blomberg Jensen, Martin; Lieben, Liesbet; Nielsen, John E

    2013-01-01

    Vitamin D is a key factor for calcium and bone homeostasis, but signalling through the vitamin D receptor (VDR) seems also to be important for testicular function. To test the functional role of vitamin D signalling we examined the male reproductive system of the Leuven Vdr-ablated (Vdr(-/-)) mice......, previously established as a model for hereditary vitamin D-resistant rickets. We investigated reproductive hormones, changes in gene expression and histological phenotype of eleven Vdr(-/-), eight Vdr(+/-) and nine Vdr(+/+) mice. Testicular and epididymal histology were grossly normal in Vdr(-/-) mice...... expressions of ERα and the estrogen-target gene Aqp9 were higher. In conclusion, vitamin D seems dispensable for murine spermatogenesis and sex hormone production, but aberrant estrogen-signalling may elicit some of the VDR-mediated effects on male reproduction....

  9. In-vitro effect of estrogen-antagonist on motility and penetration ability of human spermatozoa.

    Science.gov (United States)

    Allag, I S; Rangari, K

    1997-08-01

    Antiestrogens affect spermatozoa through their action on Leydig and Sertoli cells. Direct effect of antiestrogens namely tamoxifen and centchroman in concentration of 1, 2.5, 5, 10 and 20 micrograms/ml in incubation medium was determined on motility and penetration ability of human spermatozoa. Motility (%) was invariably reduced after 15, 30 and 60 min. of incubation. Addition of 17 beta-estradiol to medium with antagonist caused inhibition of motility in dose related manner. The distance travelled by spermatozoa treated with tamoxifen or centchroman in media was reduced by 30% and addition of estradiol along with antiestrogen reduced it to 50% compared to that of untreated spermatozoa.

  10. Freeze-dried spermatozoa: An alternative biobanking option for endangered species.

    Science.gov (United States)

    Anzalone, Debora Agata; Palazzese, Luca; Iuso, Domenico; Martino, Giuseppe; Loi, Pasqualino

    2018-03-01

    In addition to the iconic wild species, such as the pandas and Siberian tigers, an ever-increasing number of domestic species are also threatened with extinction. Biobanking of spermatozoa could preserve genetic heritages of extinct species, and maintain biodiversity of existing species. Because lyophilized spermatozoa retain fertilizing capacity, the aim was to assess whether freeze-dried spermatozoa are an alternative option to save endangered sheep breeds. To achieve this objective, semen was collected from an Italian endangered sheep breed (Pagliarola), and a biobank of cryopreserved and freeze-dried spermatozoa was established, and evaluated using IVF (for frozen spermatozoa) and ICSI procedures (for frozen and freeze-dried spermatozoa). As expected, the fertilizing capacity of cryopreserved Pagliarola's spermatozoa was comparable to commercial semen stocks. To evaluate the activating capability of freeze-dried spermatozoa, 108 MII sheep oocytes were subjected to ICSI, and allocated to two groups: 56 oocytes were activated by incubation with ionomycin (ICSI-FDSa) and 52 were not activated (ICSI-FDSna). Pronuclear formation (2PN) was investigated at 14-16 h after ICSI in fixed presumptive zygotes. Only artificially activated oocytes developed into blastocysts after ICSI. In the present study, freeze-dried ram spermatozoa induced blastocyst development following ICSI at a relatively high proportion, providing evidence that sperm lyophilization is an alternative, low cost storage option for biodiversity preservation of domestic species. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. Conjugated linoleic acid supplementation caused reduction of perilipin1 and aberrant lipolysis in epididymal adipose tissue

    International Nuclear Information System (INIS)

    Cai, Demin; Li, Hongji; Zhou, Bo; Han, Liqiang; Zhang, Xiaomei; Yang, Guoyu; Yang, Guoqing

    2012-01-01

    Highlights: ► Conjugated linoleic acid supplementation suppresses perilipin1 in epididymal fat. ► Conjugated linoleic acid inhibits promoter activity of perilipin1 in 3T3-L1 cells. ► Conjugated linoleic acids elevate basal but blunt hormone-stimulated lipolysis. -- Abstract: Perilipin1, a coat protein of lipid droplet, plays a key role in adipocyte lipolysis and fat formation of adipose tissues. However, it is not clear how the expression of perilipin1 is affected in the decreased white adipose tissues (WAT) of mice treated with dietary supplement of conjugated linoleic acids (CLA). Here we obtained lipodystrophic mice by dietary administration of CLA which exhibited reduced epididymal (EPI) WAT, aberrant adipocytes and decreased expression of leptin in this tissue. We found both transcription and translation of perilipin1 was suppressed significantly in EPI WAT of CLA-treated mice compared to that of control mice. The gene expression of negative regulator tumor necrosis factor α (TNFα) and the positive regulator Peroxisome Proliferator-Activated Receptor-γ (PPARγ) of perilipin1 was up-regulated and down-regulated, respectively. In cultured 3T3-L1 cells the promoter activity of perilipin1 was dramatically inhibited in the presence of CLA. Using ex vivo experiment we found that the basal lipolysis was elevated but the hormone-stimulated lipolysis blunted in adipose explants of CLA-treated mice compared to that of control mice, suggesting that the reduction of perilipin1 in white adipose tissues may at least in part contribute to CLA-mediated alternation of lipolysis of WAT.

  12. The effect of citric acid and citrate on protoplasmic droplet of bovine epididymal sperms

    Directory of Open Access Journals (Sweden)

    Keivan Abdy

    2011-11-01

    Full Text Available AbstractFor evaluation of citric acid and citrate effects on bovine epididymal protoplasmic droplets, fifty bovine testes were collected in the October 2007 till June 2008 from Urmia slaughterhouse and transported to the laboratory in a cool container filled with 5 °C ice pack. Caudal epididymis was incised and sperm cells were put into Petri dishes containing hams f10 media with 10% fetal calf serum (FCS, which were kept in 37 °C, CO2 incubator. Then sperm cells were counted and 50-milion per mL concentration was prepared. After this stage, three dilutions of citric acid (0.1, 0.2, 0.3 N and one dilution of citrate (1N, based on normal osmolarity and normal pH were added to a micro tube containing 25 milion per mL sperm. Then one-step eosin-nigrosin staining in 30-60-120-240-360 minutes was performed and slides were evaluated with 1000-x phase contrast microscope and 200 sperm cells per slide were counted. The results revealed significant difference between blank and citric acid 0.3 N. The proportion of protoplasmic droplet in group consisting of 0.3 N acid citric in 120-240-360 minutes, was significantly lower than that of blank (P < 0.05. There was no significant difference between citrate – blank and citric acid 0.1N-blank groups, but after 240 minutes significant difference was observed between blank & citric acid 0.2 N (P < 0.05. In conclusions citric acid based on dilution and time duration can reduce the proportion of bovine epididymal sperm cytoplasmic droplets.

  13. Conjugated linoleic acid supplementation caused reduction of perilipin1 and aberrant lipolysis in epididymal adipose tissue

    Energy Technology Data Exchange (ETDEWEB)

    Cai, Demin [College of Animal Sciences and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Li, Hongji [Key Laboratory of Animal Biochemistry and Nutrition, Ministry of Agriculture, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Zhou, Bo [College of Animal Sciences and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Han, Liqiang [Key Laboratory of Animal Biochemistry and Nutrition, Ministry of Agriculture, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Zhang, Xiaomei [College of Animal Sciences and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Yang, Guoyu, E-mail: haubiochem@163.com [Key Laboratory of Animal Biochemistry and Nutrition, Ministry of Agriculture, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China); Yang, Guoqing, E-mail: gqyang@yeah.net [College of Animal Sciences and Veterinary Medicine, Henan Agricultural University, Zhengzhou 450002, Henan Province, People' s Republic of China (China)

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer Conjugated linoleic acid supplementation suppresses perilipin1 in epididymal fat. Black-Right-Pointing-Pointer Conjugated linoleic acid inhibits promoter activity of perilipin1 in 3T3-L1 cells. Black-Right-Pointing-Pointer Conjugated linoleic acids elevate basal but blunt hormone-stimulated lipolysis. -- Abstract: Perilipin1, a coat protein of lipid droplet, plays a key role in adipocyte lipolysis and fat formation of adipose tissues. However, it is not clear how the expression of perilipin1 is affected in the decreased white adipose tissues (WAT) of mice treated with dietary supplement of conjugated linoleic acids (CLA). Here we obtained lipodystrophic mice by dietary administration of CLA which exhibited reduced epididymal (EPI) WAT, aberrant adipocytes and decreased expression of leptin in this tissue. We found both transcription and translation of perilipin1 was suppressed significantly in EPI WAT of CLA-treated mice compared to that of control mice. The gene expression of negative regulator tumor necrosis factor {alpha} (TNF{alpha}) and the positive regulator Peroxisome Proliferator-Activated Receptor-{gamma} (PPAR{gamma}) of perilipin1 was up-regulated and down-regulated, respectively. In cultured 3T3-L1 cells the promoter activity of perilipin1 was dramatically inhibited in the presence of CLA. Using ex vivo experiment we found that the basal lipolysis was elevated but the hormone-stimulated lipolysis blunted in adipose explants of CLA-treated mice compared to that of control mice, suggesting that the reduction of perilipin1 in white adipose tissues may at least in part contribute to CLA-mediated alternation of lipolysis of WAT.

  14. Variabilidade sazonal no ducto epididimário de codorna doméstica: observações morfológicas Seasonal variability in epididymal duct of the domestic quail: morphologic features

    Directory of Open Access Journals (Sweden)

    Antônio M. Orsi

    2007-12-01

    Full Text Available O ducto epididimário (DE de codorna doméstica mostrou, ao longo do ano, variabilidade pequena, porém muito expressiva no outono, o qual corresponde à fase quiescente do ciclo testicular anual. A morfologia do DE na primavera foi, em termos, similar à verificada no verão e inverno. Nestas fases notaram-se aumento significante do calibre tubular do DE; estocagem intraluminal de espermatozóides e ocorrência de mitocôndrias, lamelas do RE, vesículas variáveis quanto à forma, dimensões e conteúdos e presença de alguns lisossomos localizados, principalmente, no citoplasma apical das células principais (P, no epitélio epididimário. Estas características ultra-estruturais das células P parecem ser indicativas da ocorrência de processos ativos de endocitose e de secreção micromerócrina. A quiescência outonal foi caracterizada pelo aspecto anfractuoso do DE; ausência de espermatozóides e pouco material intraluminal, observados à microscopia de luz. Características ultra-estruturais degenerativas foram verificadas ao nível do citoplasma supranuclear das células P epididimárias no outono.Small but expressive variability was noted on the epididymidis duct (ED of domestic quail along the year, with more evidence in autumn of the quiescent phase of the annual testis cycle in this species. Spring features of ED had a general similar pattern in summer and winter. They were characterized by enlargement of epididymis tubule, storage of spermatozoa into the luminal compartment and presence of mitochondria, ER lamellae, several variable vesicles, and lysosomes localized mainly on the apical cytoplasm of principal cells (P of the epididymal epithelium. These P cells features indicated a process of endocytosis and perhaps protein secretion. Autumn quiescence was marked by a convolute pattern of the epididymis tubule, lacking of spermatozoa and small amount of exfoliate heterogeneous material inside the luminal compartment at light

  15. The effect of sperm concentration in the ejaculate on morphological traits of bull spermatozoa.

    Science.gov (United States)

    Kondracki, Stanisław; Banaszewska, Dorota; Wysokńjska, Anna; Iwanina, Maria

    2012-01-01

    Experiments were performed on 75 ejaculates obtained from 19 bulls representing different cattle breeds used at the Masovian Centre for Animal Breeding and Reproduction in Łowicz. Fresh ejaculates were measured in respect to their volume and sperm count in the ejaculates was determined. The ejaculates were classified based on the criterion of sperm concentration and divided into five groups. Sperm morphometric measurements were taken from each bull and assessment of semen morphology was done on the basis of examination under a microscope using preparations made from fresh ejaculates. For each slide, morphometric measurements were taken of 15 randomly selected spermatozoa characterised by normal morphology and well visible under the microscope. Additionally, in each preparation morphometry of 500 spermatozoa was evaluated, numbers of spermatozoa with normal morphology and morphological abnormalities were recorded and these were categorized into spermatozoa with major and minor defects. An insignificant correlation was observed between the sperm concentration in the ejaculate and morphological traits, dimensions and shapes of bull spermatozoa. The less concentrated ejaculates contained spermatozoa with a slightly larger head circumference and a more elongated head shape in comparison with the spermatozoa in the more concentrated ejaculates. The highest frequency of morphologically malformed spermatozoa, both in the case of primary and secondary alterations, was observed in ejaculates with sperm concentration of no more than 1000 x 10(3)/mm3.

  16. Selection of High-Quality Spermatozoa May Be Promoted by Activated Vitamin D in the Woman

    DEFF Research Database (Denmark)

    Bøllehuus Hansen, Lasse; Rehfeld, Anders; de Neergaard, Rosanna

    2017-01-01

    Context: The vitamin D receptor (VDR) and enzymes involved in activation (CYP2R1, CYP27B1) and inactivation (CYP24A1) of vitamin D are expressed in ovary, testes, and spermatozoa. Objective: Determine responsiveness to 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] in spermatozoa from normal and infertil...

  17. Simultaneous vitality and DNA-fragmentation measurement in spermatozoa of smokers and non-smokers.

    Science.gov (United States)

    De Bantel, A; Fleury-Feith, J; Poirot, C; Berthaut, I; Garcin, C; Landais, P; Ravel, C

    2015-03-01

    Because cigarette smoke is a powerful ROS producer, we hypothesized that the spermatozoa of smokers would be more at risk of having increased DNA fragmentation than spermatozoa of non-smoking men. A cross-sectional study was performed on consenting smokers and non-smokers, consulting in an infertility clinic for routine sperm analysis. The application of a novel TUNEL assay coupled to a vitality marker, LIVE/DEAD®, allowed both DNA fragmentation and viability measurement within spermatozoa of participants to be analyzed by flow cytometry. The coupled vitality-DNA fragmentation analysis revealed that non-smokers and smokers, respectively presented medians of 3.6% [0.6-36.8] and 3.3% [0.9-9.6] DNA fragmented spermatozoa among the living spermatozoa population (P > 0.05). No deleterious effect of smoking on spermatozoa was found in our study. More studies concerning potential mutagenic capacities of cigarette smoke on spermatozoa are necessary. In addition, the coupled vitality-DNA fragmentation analysis may orient Assisted Reproductive Technology teams when confronted with patients having a high percentage of DNA-fragmented living spermatozoa. © 2014 International Clinical Cytometry Society.

  18. Chromosomal integrity of freeze-dried mouse spermatozoa after 137Cs γ-ray irradiation

    International Nuclear Information System (INIS)

    Kusakabe, Hirokazu; Kamiguchi, Yujiroh

    2004-01-01

    This study demonstrated that freeze-dried mouse spermatozoa possess strong resistance to 137 Cs γ-ray irradiation at doses of up to 8 Gy. Freeze-dried mouse spermatozoa were rehydrated and injected into mouse oocytes with an intracytoplasmic sperm injection (ICSI) technique. Most oocytes can be activated after ICSI by using spermatozoa irradiated with γ-rays before and after freeze-drying. Sperm chromosome complements were analyzed at the first cleavage metaphase. Chromosome aberrations increased in a dose-dependent manner in the spermatozoa irradiated before freeze-drying. However, no increase in oocytes with chromosome aberrations was observed when fertilized by spermatozoa that had been irradiated after freeze-drying, as compared with freeze-dried spermatozoa that had not been irradiated. These results suggest that both the chromosomal integrity of freeze-dried spermatozoa, as well as their ability to activate oocytes, were protected from γ-ray irradiation at doses at which chromosomal damage is found to be strongly induced in spermatozoa suspended in solution

  19. Application of quantum dot nanoparticles for potential non-invasive bio-imaging of mammalian spermatozoa

    Science.gov (United States)

    Various obstacles are encountered by mammalian spermatozoa during their journey through the female genital tract, and only few or none will reach the site of fertilization. Currently, there are limited technical approaches for non-invasive investigation of spermatozoa migration after insemination. A...

  20. SPERM HY-LITER™ for the identification of spermatozoa from sexual assault evidence

    DEFF Research Database (Denmark)

    Westring, Christian Gustav; Wiuf, Morten; Nielsen, S Jock

    2014-01-01

    Accurate microscopic identification of human spermatozoa is important in sexual assault cases. We have compared the results of examinations with (1) a fluorescent microscopy method, SPERM HY-LITER™, and (2) Baecchi's method for identification of human spermatozoa. In 35 artificial, forensic type...

  1. In vitro phagocytosis of boar spermatozoa by neutrophils from peripheral blood of sows

    NARCIS (Netherlands)

    Matthijs, A.; Harkema, W.; Engel, B.; Woelders, H.

    2000-01-01

    A considerable number of spermatozoa are used in each sow in routine artificial insemination. However, within a few hours after insemination, many spermatozoa are phagocytosed by polymorphonuclear leucocytes. Some aspects of sperm transport in the female genital tract in the sow have been thoroughly

  2. Vitamin D is positively associated with sperm motility and increases intracellular calcium in human spermatozoa

    DEFF Research Database (Denmark)

    Blomberg Jensen, Martin; Bjerrum, Poul J; Jessen, Torben E

    2011-01-01

    BACKGROUND The vitamin D receptor (VDR) is expressed in human spermatozoa, and VDR-knockout mice and vitamin D (VD) deficiency in rodents results in impaired fertility, low sperm counts and a low number of motile spermatozoa. We investigated the role of activated VD (1,25(OH)(2)D(3)) in human...... spermatozoa and whether VD serum levels are associated with semen quality. METHODS Cross-sectional association study of semen quality and VD serum level in 300 men from the general population, and in vitro studies on spermatozoa from 40 men to investigate the effects of VD on intracellular calcium, sperm......M). 1,25(OH)(2)D(3) increased intracellular calcium concentration in human spermatozoa through VDR-mediated calcium release from an intracellular calcium storage, increased sperm motility and induced the acrosome reaction in vitro. CONCLUSIONS 1,25(OH)(2)D(3) increased intracellular calcium...

  3. EXTENDING THE VIABILITY OF SPERMATOZOA AND EGGS OF THE SEA URCHIN LYTECHINUS VARIEGATUS.

    Science.gov (United States)

    Malgarin, Jéssica; Resgalla, Charrid

    2015-01-01

    The storage of spermatozoa and eggs of the sea urchin Lytecninus variegatus can meet the demand of different human activities. To develop a protocol easy to reproduce for spermatozoa cryopreservation and cooling of the eggs of the sea urchin. Different formulations of artificial sea water were tested for their effectiveness in the freezing of sea urchin spermatozoa and storage of the eggs. Protocol for freezing of spermatozoa in liquid nitrogen presented the positive results when the cryoprotectant solution was diluted in artificial seawater free of calcium and magnesium. For the conservation of the eggs by cooling, the calcium-free artificial sea water, the calcium- and magnesium-free sea water, and the low-sodium water proved more efficient in preserving the integrity of the eggs. The results showed success in the freezing protocol of spermatozoa and cooling of the eggs mainly in artificial calcium- and magnesium-free sea water.

  4. Persistent scrotal pain and suspected orchido-epididymitis of a young boy during pinworm (Enterobius vermicularis) infection in the bowel.

    Science.gov (United States)

    Sinikumpu, J-J; Serlo, W

    2011-08-01

    Pinworm (Enterobius vermicularis) is the most common parasite among mankind. Ectopic pinworm infections in number of organs are broadly reported. The aim of this report is to review the pinworm infection in the scrotum on the basis of our case. Our patient is a young boy with persistent pain in the scrotums together with abdominal symptoms. He underwent several urgent operations. Pinworm was finally found in appendix vermiformis. We hypothesize that concurrent unresponsive orchido-epididymitis was caused by pinworm as well. Eradication happened finally with pyvrinembonate. Pinworm is a parasite that lives usually in the bowel. Many ectopic locations, like scrotum, are known. Orchido-epididymitis by pinworm has to be kept in mind when treating boys with persistent scrotal pain. © 2011 The Author(s)/Acta Paediatrica © 2011 Foundation Acta Paediatrica.

  5. Evaluation of the damage in fish spermatozoa cryopreservation

    Science.gov (United States)

    Li, Jun; Liu, Qinghua; Zhang, Shicui

    2006-12-01

    Cryodamages occur during sperm cryopreservation. Cryopreservation of fish sperm usually results in marked decrease in sperm quality, such as swelling or disruption of the plasma membrane, mitochondrial dysfunction, diminished sperm motility, impaired velocity, shorter motility period, denaturation, and release of some enzymes from spermatozoa. In this paper, damages in morphology, physiology, biochemistry and metabolism, and genetic integrity of fish semen after cryopreservation are discussed. New approaches in assessment of fish thawed sperm quality such as computer assisted sperm analysis, flow cytometic analysis combined with fluorescent probes and single cell gel electrophoresis are also briefly reviewed.

  6. Outlining adequate protocols for Lidia bull epididymal storage and sperm cryopreservation: use of glycerol, dimethylformamide and N-acetylcysteine

    Directory of Open Access Journals (Sweden)

    Elvira Matilla

    2017-12-01

    Full Text Available The Lidia bovine breed is an important hallmark of the Spanish cattle industry. Bulls are selected based upon aggressiveness and epididymal sperm cryopreservation is the way to obtain and store their genetics. There are not specifically designed protocols yet to perform Lidia bull sperm cryopreservation. The present study aimed to determine if a tris-fructose-citrate-egg yolk (20% v/v; TFY extender supplemented with 7% glycerol (TFY1 or 3.5% glycerol plus 3.5% dimethylformamide (DMF; TFY2 are suitable media for cryopreservation of epididymal Lidia bull sperm. Moreover, the effect of N-acetylcysteine (NAC, a potent antioxidant, was evaluated. The epididymis were stored at 4°C for 24, 48, 72 or 96 h, and both freezing media were tested as such or supplemented with 1 or 2.5 mM of NAC. Our data demonstrated that post-thaw viability was well maintained (TFY1: 50.8% ± 1.9 at 24 h and 52.4% ± 0.8 at 96 h and TFY2: 52.6% ± 1.6 at 24 h and 56.1% ± 1.8 at 96 h; mean % ± SEM; p>0.05 as also were total and progressive sperm motility, high mitochondrial membrane potential, ROS production, DNA status and acrosomal intactness of Lidia bull sperm up to 96 h of epididymal storage, all extender variations being similar (p>0.05. In conclusion, the use of TFY medium supplemented either with 7% glycerol alone or the combination of 3.5% glycerol and 3.5% DMF were equally safe choices for epididymal Lidia bull sperm cryopreservation, and NAC addition did not significantly improve sperm post-thaw quality.

  7. [IFN-gamma enzyme-linked immunospot assay versus PPD tuberculin skin test in the diagnosis of tuberculous epididymitis].

    Science.gov (United States)

    Huang, Hao; Yang, Xi-Fei; Deng, Qun-Yi; Li, Bing; Liu, Guo-Hui; Zhang, Jie-Yun; Yang, Da-Fei

    2012-06-01

    To explore the potential application of IFN-gamma enzyme-linked immunospot (ELISPOT) assay in the diagnosis of tuberculous epididymitis (TE) by comparing ELISPOT assay with the traditional purified protein derivative (PPD) tuberculin skin test. We examined 13 TE patients using an in-house ELISPOT kit, another 11 TE patients by PPD skin testing, and 57 healthy male volunteers by parallel test with both the methods. Twelve (92.3%) of the 13 TE cases were positive on ELISPOT assay, and 10 (90.9%) of the 11 TE cases positive on PPD skin test, with no statistically significant differences between the two groups (P > 0.05). Among the 57 healthy male volunteers, 8 (14.0%) were positive on ELISPOT, and 28 (49.1%) positive on PPD test, the latter significantly higher than the former (P PPD test in the examination of tuberculous epididymitis. As for specificity, ELISPOT assay seems better than PPD test in differentiating tuberculous epididymitis patients from healthy males.

  8. Evaluation of Lasting Effects of Heat Stress on Sperm Profile and Oxidative Status of Ram Semen and Epididymal Sperm

    Directory of Open Access Journals (Sweden)

    Thais Rose dos Santos Hamilton

    2016-01-01

    Full Text Available Higher temperatures lead to an increase of testicular metabolism that results in spermatic damage. Oxidative stress is the main factor responsible for testicular damage caused by heat stress. The aim of this study was to evaluate lasting effects of heat stress on ejaculated sperm and immediate or long-term effects of heat stress on epididymal sperm. We observed decrease in motility and mass motility of ejaculated sperm, as well as an increase in the percentages of sperm showing major and minor defects, damaged plasma and acrosome membranes, and a decrease in the percentage of sperm with high mitochondrial membrane potential in the treated group until one spermatic cycle. An increased enzymatic activity of glutathione peroxidase and an increase of stressed cells were observed in ejaculated sperm of the treated group. A decrease in the percentage of epididymal sperm with high mitochondrial membrane potential was observed in the treated group. However, when comparing immediate and long-term effects, we observed an increase in the percentage of sperm with low mitochondrial membrane potential. In conclusion, testicular heat stress induced oxidative stress that led to rescuable alterations after one spermatic cycle in ejaculated sperm and also after 30 days in epididymal sperm.

  9. The effect of different concentrations of citric acid on motility patterns of bovine epididymal sperms in Hams F10 milieu

    Directory of Open Access Journals (Sweden)

    K Abdy

    2008-11-01

    Full Text Available The aim of this study was to investigate the effect of three concentration of citric acid on motility patterns of bovine epididymal sperms. For this purpose, 50 pairs of bovine testicles were collected immediately after slaughter form urmia abattoir and transferred to the laboratory alongside 5oc ice pack. Epididymal tail sperms were collected with a few incisions in vascular areas and transferred to hams f10 milieu with 10% fetal calf serum and counted after 15 minutes of incubation at 37oc in Co2 incubator. Concentrations of 50 million sperms per ml were proposal and in the normal sperm pH rang of 6.7-7.4, 0.1, 0.2 and 0.3 normal concentration of citric acid were added to sperm continuity micro tubes (normal concentration of acid equals 7 mg/ml of bovine serum and at 15, 30, 45, 60, 90, 120, 180, 240 and 360 minutes the motility patterns of epididymal sperms were evaluated using the computer assisted sperms analyzing (CASA method. Data were analyzed with one-way ANOVA using the SPSS 15 software. The results indicated significant differences in various indices of sperm motility patterns (Curvilinear   Velocity, Straight-line Velocity, Average Path Velocity, Mean Angel Degree, Amplitude of Lateral Head Displacement, Beat-Cross Frequency, Linearity, Wobble particularly at 0.3 normal concentration of citric acid compared with the control.

  10. The binding patterns of antisera to sex steroids and human gonadotropins on human and rhesus monkey spermatozoa.

    Science.gov (United States)

    Allag, I S; Das, R P; Roy, S

    1983-01-01

    The presence of different hormones on the surface of ejaculated spermatozoa was determined by immunofluorescence studies of the binding patterns of specific antisera to these hormones. There were striking similarities in the binding pattern of antisera to steroid hormones found on human and monkey spermatozoa. Assuming the intensity of fluorescence is proportional to the concentration of the hormone, concentrations of testosterone on the acrosomal and the postacrosomal regions were higher than levels of progesterone and estrogens. Spermatozoa with a "tapering head" had more hCG bound on the acrosomal and postacrosomal regions than spermatozoa with "normal head" (oval shaped). Correlating these findings to the functions of spermatozoa will require further studies.

  11. Identification of phosphoproteins coupled to initiation of motility in live epididymal mouse sperm

    Science.gov (United States)

    Tash, J. S.; Bracho, G. E.

    1998-01-01

    A method for collecting live immotile cauda epididymal mouse sperm that initiate motility by dilution into an activation buffer is described. Sperm in collection buffer showed low percent motility (MOT) and population progression (PRG) that increased 10-fold and 9-fold, respectively, during the first 2 min after dilution into activation buffer. Western phosphoserine (pS), phosphothreonine (pT), and phosphotyrosine (pY) analysis revealed a 120 kDa protein that markedly increased in pT content during initiation of motility and may be related to FP130, the motility-coupled axonemal protein of sea urchin sperm. A prominent 82 kDa protein that was pS and pT-phosphorylated in immotile and motile sperm is likely the fibrous sheath component AKAP82 that is phosphorylated during spermatogenesis. Analysis of live human sperm also identified a prominent 120 kDa pT protein. Thus it appears that phosphorylation of FP130 and related 120 kDa proteins in mouse, and perhaps human sperm, represent common targets during motility initiation in sperm. Copyright 1998 Academic Press.

  12. [Expression of DKKL1 in spermatozoa of men with asthenospermia].

    Science.gov (United States)

    Yan, Qiu-Xia; Ma, Yi; Chen, Run-Qiang; Zhou, Xiu-Qin; Qiao, Jing; Xian, Ying-Jie; Feng, Ling; Chen, Cai-Rong

    2018-03-20

    To compare the expression of DKKL1 in ejaculated spermatozoa of normal fertile men and men with asthenospermia and investigate the role of DKKL1 in the pathogenesis of asthenospermia. The characteristics of semen samples collected from normal fertile men and men with asthenospermia were analyzed using computer-assisted sperm analysis according to WHO criteria. The ejaculated sperms were isolated by Percoll discontinuous density gradients to detect the expression of DKKL1 mRNA and protein using real-time PCR and Western blotting. The expression of DKKL1 mRNA was significantly down-regulated by 11.1 times in asthenospermic men as compared with that in normal fertile men (P<0.01). Western blotting showed that the expression of DKKL1 protein was down-regulated by 2.4 times in asthenospermic men compared to normal fertile men. The expression of DKKL1, which may play an important role in sperm motility,is significantly decreased in ejaculated spermatozoa of men with asthenospermia.

  13. Modification of spermatozoa quality in mature small ruminants.

    Science.gov (United States)

    Martin, G B; de St Jorre, T Jorre; Al Mohsen, F A; Malecki, I A

    2011-01-01

    This review is based largely, but not entirely, on the assumption that gamete quality is directly linked to sperm output and thus testicular mass, an approach made necessary by the absence of a large body of data on factors that affect gamete quality in ruminants. On the other hand, there is a change in the efficiency of sperm production per gram of testicular tissue when the testis is growing or shrinking, a clear indicator of changes in the rates of cell loss during the process of spermatogenesis, probably through apoptosis. We therefore postulate that the spermatozoa that do survive when the testis is shrinking are of a lower quality than those that are produced when the testis is growing and the rate of sperm survival is increasing. In adult small ruminants in particular, testicular mass and sperm production are highly labile and can be manipulated by management of photoperiod (melatonin), nutrition, genetics and behaviour ('mating pressure'). Importantly, these factors do not act independently of each other - rather, the outcomes in terms of sperm production are dictated by interactions. It therefore seems likely that spermatozoa quality will be affected by these same factors, but definitive answers await detailed studies.

  14. Ultrastructure of spermatozoa in cobia, Rachycentron canadum (Linnaeus, 1766).

    Science.gov (United States)

    Dhanasekar, Krishnamoorthy; Selvakumar, Narasimman; Munuswamy, Natesan

    2018-02-01

    Ultrastructure and development of spermatozoa in cobia, Rachycentron canadum are described. Sections through the testis show different developmental stages viz, Spermatocytes, spermatids and sperm. Spermatozoa of R. canadum exhibit the configuration of uniflagellated, anacrosomal Type I aquasperm, typical for externally fertilizing fish. Mature spermatozoon is seen with a prominent head and long cylindrical flagellum. Ultrastructure of sperm shows invaginated 'U' shaped nucleus and other organelles. The mitochondrial matrix is electron-dense with irregular arrangement of the cristae. The nucleus reveals a deep invagination (nuclear fossa) in which the centriolar complex is located. The centriolar complex lies inside the nuclear fossa and is composed of a proximal and a distal centriole. The two centrioles are placed perpendicular to each other. The flagellum has a typical eukaryotic organization (microtubule doublets 9 + 2 pattern) and measures around 36.21 ± 0.42 μm in length. This study for the first time provides a comprehensive detail on the ultrastructure and developmental process of sperm in cobia, R. canadum. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Successful use of the Cryolock device for cryopreservation of scarce human ejaculate and testicular spermatozoa.

    Science.gov (United States)

    Stein, A; Shufaro, Y; Hadar, S; Fisch, B; Pinkas, H

    2015-03-01

    The existing methods for cryopreservation of very low count sperm samples are complex and sub-optimal for individual spermatozoa. Our purpose is to establish an effective simple method for cryoprotecting individual spermatozoa. Samples from patients with OTA were mixed with TYB or HEPES-buffered salt solution with glycerol + glucose and placed on a Cryolock that was plunged directly into liquid nitrogen or exposed to its vapors. Thawing was performed by direct immersion into a drop of warmed medium. The favorable method was tested on diluted samples (10-50 cells) and leftover TESE specimens from patients with azoospermia. Cryopreservation was considered successful if >30 spermatozoa, (>3 motile), or >5 spermatozoa (>1 motile) in diluted and TESE samples, were detected post-thawing. A significantly higher survival rate of seminal spermatozoa was obtained when using the Cryolock with TYB and freezing with liquid nitrogen vapor, compared to HEPES glycerol-glucose (95 vs. 35% respectively). Plunging the Cryolock into liquid nitrogen was detrimental. Cryolock combined with TYB cryoprotection and liquid nitrogen vapor freezing was highly effective for cryopreservation of individual spermatozoa in diluted and TESE samples. The Cryolock may serve for freezing very low-count sperm samples and individual spermatozoa. This method offers simplicity, efficacy, use of available materials, without requiring micromanipulation equipment or skills. © 2015 American Society of Andrology and European Academy of Andrology.

  16. In-depth proteomic analysis of carp (Cyprinus carpio L) spermatozoa.

    Science.gov (United States)

    Dietrich, Mariola A; Arnold, Georg J; Fröhlich, Thomas; Ciereszko, Andrzej

    2014-12-01

    Using a combination of protein fractionation by one-dimensional gel electrophoresis and high performance liquid chromatography-electrospray ionization tandem mass spectrometry, we identified 348 proteins in carp spermatozoa, most of which were for the first time identified in fish. Dynein, tubulin, HSP90, HSP70, HSP60, adenosylhomocysteinase, NKEF-B, brain type creatine kinase, mitochondrial ATP synthase, and valosin containing enzyme represent high abundance proteins in carp spermatozoa. These proteins are functionally related to sperm motility and energy production as well as the protection of sperm against oxidative injury and stress. Moreover, carp spermatozoa are equipped with functionally diverse proteins involved in signal transduction, transcription, translation, protein turnover and transport. About 15% of proteins from carp spermatozoa identified here were also detected in seminal plasma which may be a result of leakage from spermatozoa into seminal plasma, adsorption of seminal plasma proteins on spermatozoa surface, and expression in both spermatozoa and cells secreting seminal plasma proteins. The availability of a catalog of carp sperm proteins provides substantial advances for an understanding of sperm function and for future development of molecular diagnostic tests of carp sperm quality, the evaluation of which is currently limited to certain parameters such as sperm count, morphology and motility or viability. The mass spectrometry data are available at ProteomeXchange with the dataset identifier PXD000877 (DOI: http://dx.doi.org/10.6019/PXD000877). Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Quality of human spermatozoa: relationship between high-magnification sperm morphology and DNA integrity.

    Science.gov (United States)

    Maettner, R; Sterzik, K; Isachenko, V; Strehler, E; Rahimi, G; Alabart, J L; Sánchez, R; Mallmann, P; Isachenko, E

    2014-06-01

    The aim of this work is to establish the relationship between the morphology of Intracytoplasmic Morphologically Selected Sperm Injection (IMSI)-selected spermatozoa and their DNA integrity. The 45 ejaculates were randomly distributed into three treatment groups: normozoospermic, oligoasthenozoospermic and oligoasthenotheratozoospermic samples. The evaluation of DNA integrity was performed using the sperm chromatin dispersion test. It was established that DNA integrity of spermatozoa is strongly dependent on ejaculate quality (P count of spermatozoa with nonfragmented DNA in normozoospermic samples was high and independent from IMSI-morphological classes (Class 1 versus Class 3, respectively) (P > 0.1). With decreased ejaculate quality, the percentage of spermatozoa with nonfragmented DNA decreased significantly (P < 0.05) independent from morphological class. Nevertheless, the rate of IMSI-selected spermatozoa with fragmented DNA within of Class 1 in normozoospermic (Group 1), in oligoasthenozoospermic (Group 2) and in oligoasthenotheratozoospermic (Group 3) samples was 21.1%, 31.8% and 54.1%, respectively. In conclusion, there is a direct relationship between morphological parameters of spermatozoa and their DNA integrity. However, the IMSI technique alone is not enough for the selection of spermatozoa with intact nuclei. © 2013 Blackwell Verlag GmbH.

  18. A method for tracing exogenous DNA uptake in live spermatozoa and embryos.

    Science.gov (United States)

    Mu, Y; Jiao, M; Zhao, Y; Lv, J; Wang, J; Hao, J; Zhang, X; Kong, Q; Liu, Z

    2018-03-01

    Sperm-mediated gene transfer(SMGT) is a simple method for producing transgenic animals. Due to the lack of repeatability in spermatozoa binding and internalization of exogenous DNA, the efficiency of SMGT is still low. Considering this point, the present work aims to develop a method for evaluating the spermatozoa capacity of binding exogenous DNA after co-incubation with DNA. The main approach is using a Cy5-labelled DNA to trace the exogenous DNA and assess the ability of spermatozoa to take up exogenous DNA. Using this technique, we found that the percentage of spermatozoa that are binding and uptaking DNA is higher at concentration of 10 μg/mL and 100 μg/mL than 5 μg/mL, 1 μg/mL and 0 μg/mL after incubation with Cy5-DNA for 30min at 37oC. After fertilization, the DNA fluorescence signal was also detected in zygotes in groups where spermatozoa were incubated with 10 μg/mL and 100 μg/mL of Cy5-DNA. These results showed a simple and convenient method to trace the exogenous DNA in spermatozoa and zygote when compared to conventional methods of labeling DNA during fertilization, resulting in a real-time observation of the exogenous DNA in spermatozoa and zygote. Copyright© by the Polish Academy of Sciences.

  19. Oxidation flux change on spermatozoa membrane in important pathologic conditions leading to male infertility.

    Science.gov (United States)

    Wiwanitkit, V

    2008-06-01

    Free radicals or reactive oxygen species mediate their action through proinflammatory cytokines and this mechanism has been proposed as a common underlying factor for male infertility. There is extensive literature on oxidative stress and its role in male infertility and sperm DNA damage and its effects on assisted reproductive techniques. However, there has never been a report on the oxidation flux change in spermatozoa. Here, the author determined the oxidation flux change in such hypoxic cases, using the simulation test based on nanomedicine technique is used. Of interest, change of flux can be detected. The main pathogenesis should be the direct injury of membrane structure of spermatozoa by free radicals which can lead to sperm defect. Therefore, this work can support the finding that the oxidation flux change corresponding to oxygen pressure change in spermatozoa does not exist. However, the flux change can be seen if the membrane thickness of spermatozoa is varied. Thin membrane spermatozoa are more prone to oxidative stress than thick membrane ones. The defect in the enzymatic system within the spermatozoa should be a better explanation for vulnerability of spermatozoa to oxidative stress. The use of enzymatic modification technique by antioxidants can be useful alternative in management of male infertility.

  20. Triglyceride synthesis in epididymal adipose tissue: contribution of glucose and non-glucose carbon sources.

    Science.gov (United States)

    Bederman, Ilya R; Foy, Steven; Chandramouli, Visvanathan; Alexander, James C; Previs, Stephen F

    2009-03-06

    The obesity epidemic has generated interest in determining the contribution of various pathways to triglyceride synthesis, including an elucidation of the origin of triglyceride fatty acids and triglyceride glycerol. We hypothesized that a dietary intervention would demonstrate the importance of using glucose versus non-glucose carbon sources to synthesize triglycerides in white adipose tissue. C57BL/6J mice were fed either a low fat, high carbohydrate (HC) diet or a high fat, carbohydrate-free (CF) diet and maintained on 2H2O (to determine total triglyceride dynamics) or infused with [6,6-(2)H]glucose (to quantify the contribution of glucose to triglyceride glycerol). The 2H2O labeling data demonstrate that although de novo lipogenesis contributed approximately 80% versus approximately 5% to the pool of triglyceride palmitate in HC- versus CF-fed mice, the epididymal adipose tissue synthesized approximately 1.5-fold more triglyceride in CF- versus HC-fed mice, i.e. 37+/-5 versus 25+/-3 micromolxday(-1). The [6,6-(2)H]glucose labeling data demonstrate that approximately 69 and approximately 28% of triglyceride glycerol is synthesized from glucose in HC- versus CF-fed mice, respectively. Although these data are consistent with the notion that non-glucose carbon sources (e.g. glyceroneogenesis) can make substantial contributions to the synthesis of triglyceride glycerol (i.e. the absolute synthesis of triglyceride glycerol from non-glucose substrates increased from approximately 8 to approximately 26 micromolxday(-1) in HC- versus CF-fed mice), these observations suggest (i) the importance of nutritional status in affecting flux rates and (ii) the operation of a glycerol-glucose cycle.

  1. Fructose increases corticosterone production in association with NADPH metabolism alterations in rat epididymal white adipose tissue.

    Science.gov (United States)

    Prince, Paula D; Santander, Yanina A; Gerez, Estefania M; Höcht, Christian; Polizio, Ariel H; Mayer, Marcos A; Taira, Carlos A; Fraga, Cesar G; Galleano, Monica; Carranza, Andrea

    2017-08-01

    Metabolic syndrome is an array of closely metabolic disorders that includes glucose intolerance/insulin resistance, central obesity, dyslipidemia, and hypertension. Fructose, a highly lipogenic sugar, has profound metabolic effects in adipose tissue, and has been associated with the etiopathology of many components of the metabolic syndrome. In adipocytes, the enzyme 11 β-HSD1 amplifies local glucocorticoid production, being a key player in the pathogenesis of central obesity and metabolic syndrome. 11 β-HSD1 reductase activity is dependent on NADPH, a cofactor generated by H6PD inside the endoplasmic reticulum. Our focus was to explore the effect of fructose overload on epididymal white adipose tissue (EWAT) machinery involved in glucocorticoid production and NADPH and oxidants metabolism. Male Sprague-Dawley rats fed with a fructose solution (10% (w/v) in tap water) during 9 weeks developed some characteristic features of metabolic syndrome, such as hypertriglyceridemia, and hypertension. In addition, high levels of plasma and EWAT corticosterone were detected. Activities and expressions of H6PD and 11 β-HSD1, NAPDH content, superoxide anion production, expression of NADPH oxidase 2 subunits, and indicators of oxidative metabolism were measured. Fructose overloaded rats showed an increased potential in oxidant production respect to control rats. In parallel, in EWAT from fructose overloaded rats we found higher expression/activity of H6PD and 11 β-HSD1, and NADPH/NADP + ratio. Our in vivo results support that fructose overload installs in EWAT conditions favoring glucocorticoid production through higher H6PD expression/activity supplying NADPH for enhanced 11 β-HSD1 expression/activity, becoming this tissue a potential extra-adrenal source of corticosterone under these experimental conditions. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Recovery of sperm after epididymal refrigeration from domestic cats using ACP-117c and Tris extenders

    Directory of Open Access Journals (Sweden)

    D.B.C. Lima

    2016-08-01

    Full Text Available ABSTRACT We aimed to compare fresh sperm and sperm cooled to 4ºC that had been recovered from the epididymides of cats using powdered coconut water (ACP-117c and Tris extenders. Sixty epididymides were divided into 6 groups: 10 fresh epididymides were recovered using Tris (T0h; 10 were kept at 4°C/2h and recovered using Tris (T2h; 10 were kept at 4°C/4h and recovered using Tris (T4h; 10 fresh were recovered using ACP-117c (A0h; 10 were kept at 4°C/2h and recovered using ACP-117c (A2h, and 10 were kept at 4°C/4h and recovered using ACP-117c (A4h. The testis-epididymis complexes (TEC control were not cooled. The others were cooled at 4°C for 2 or 4h. The epididymis was separated and the sperm was recovered by the modified flotation method. Sperm kinetic parameters were evaluated by a computer-system analysis, and vigor, viability, concentration, membrane function and morphology of the sperm were assessed under a light microscope. The progressive motility with ACP-117c declined after 2h of cooling, but did not differ between fresh and 4h. The vigor and membrane function were higher in A4h than A0h. The vigor at T2h and T4h were decreased compared to T0h. T0h was higher than A0h for vigor and sperm membrane function. However, after 4h of cooling, ACP-117c maintained a higher percentage of living cells. Feline epididymal sperm quality can be maintained to the degree necessary for artificial breeding programs following cooling and ACP-117c may be successfully used to recover cat sperm that have been cooled for up to 4h.

  3. VIABILITAS SPERMATOZOA KAMBING BOER PASCA PENDINGINAN DAN PEMBEKUAN MENGGUNAKAN PENGENCER DASAR TRIS DENGAN LEVEL TREHALOSA YANG BERBEDA

    Directory of Open Access Journals (Sweden)

    Nurul Isnaini

    2012-04-01

    Full Text Available ABSTRAK Penyimpanan semen bisa dilakukan dengan pendinginan atau pembekuan. Setelah pendinginan atau pembekuan viabilitas spermatozoa akan menurun. Untuk dapat mempertahankan viabilitas spermatozoa selama pendinginan atau pembekuan diperlukan krioprotektan dengan level tepat. Trehalosa merupakan krioprotektan ekstraseluler yang mampu menstabilkan membrane dan sebagai anti oksidan bagi spermatozoa selama pendinginan dan pembekuan sehingga diharapkan dapat mempertahankan viabilitas spermatozoa. Masalah dalam penelitian ini adalah bagaimanakah peranan trehalosa pada level yang berbeda dalam pengencer dasar tris mampu menekan penurunan viabilitas spermatozoa kambing Boer selama pendinginan dan pembekuan? Tujuan  dari penelitian ini adalah mengkaji pengaruh berbagai level trehalosa dalam pengencer dasar tris terhadap viabilitas spermatozoa kambing Boer setelah  pendinginan dan pembekuan. Metode penelitian yang digunakan adalah eksperimental laboratorium. Penelitian terdiri atas 2 tahap. Tahap I. Pengaruh level trehalosa dalam pengencer tris terhadap viabilitas spermatozoa kambing Boer setelah pendinginan. Tahap 2. Pengaruh level trehalosa dalam pengencer tris terhadap viabilitas spermatozoa kambing Boer setelah pembekuan. Level trehalosa yang dicobakan pada masing-masing tahap adalah: 1,5%; 2,5% dan 3,5%, dan ulangan: 10 kali. Variabel yang diamati adalah viabilitas spermatozoa. Hasil penelitian menunjukkan bahwa level trehalosa berpengaruh terhadap viabilitas spermatozoa, baik setelah pendinginan maupun pembekuan. Dari hasil penelitian dapat disimpulkan bahwa dalam pendinginan dan pembekuan semen kambing Boer, masing-masing penambahan level 1,5% dan 2,5% dalam pengencer dasar tris menghasilkan viabilitas spermatozoa yang optimal. Dari hasil penelitian disarankan bahwa dalam pendinginan dan pembekuan semen kambing Boer sebaiknya ditambahkan trehalosa masing-masing 1,5% dan 2,5% dalam pengencer dasar tris agar mendapatkan viabilitas spermatozoa yang

  4. Influence of selected factors on bovine spermatozoa cold shock resistance

    Directory of Open Access Journals (Sweden)

    Luděk Stádník

    2015-01-01

    Full Text Available The objectives of this study were to determine the effects of sire, extender, and addition of Low Density Lipoprotein (LDL to extenders used on the percentage rate of spermatozoa survival after cold shock. Two groups of extenders were compared: without LDL addition (control variants and LDL enriched (experimental variants. Three extenders were used: AndroMed®, Bioxcell®, and Triladyl®. Experimental variants included 4–8% LDL addition into the AndroMed® and Bioxcell® extenders, and 6–10% LDL addition into the Triladyl® extender. In total, 12 samples of fresh semen were collected from 4 bulls during a period of 8 weeks. Bovine spermatozoa cold shock resistance (1 ± 1 °C, 10 min was evaluated by the percentage rate of live sperm using eosin-nigrosine staining immediately and after heat incubation (37 ± 1 °C, 120 min. The results showed the effect of sire as important and individual differences between selected sires in their sperm resistance against cold shock were confirmed. AndroMed® and Bioxcell® were found to be providing better protection of bull semen to cold shock compared to Triladyl® due to lower decline of live sperm proportion. Our results detected a positive effect of LDL addition on sperm resistance against cold shock, especially on lower decrease of live sperm percentage rate after 120 min of the heat test (P < 0.05. Further studies are needed to assess the optimal concentration of LDL in various kinds of extenders as well to state ideal time and temperature conditions for ensuring LDL reaction with sperm.

  5. Induction of oxidative stress in rat epididymal sperm after exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin

    Energy Technology Data Exchange (ETDEWEB)

    Latchoumycandane, C.; Chitra, K.C.; Mathur, P.P. [School of Life Sciences, Pondicherry University (India)

    2002-03-01

    The ability of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to induce oxidative stress in various tissues of animals has been reported. The nature and mechanism of action of TCDD on the antioxidant system of sperm has not been studied. In the present study we have sought to investigate whether TCDD induces oxidative stress in the epididymal sperm of rats. Subchronic doses of TCDD (1, 10, and 100 ng/kg body weight per day) were administered orally to male Wistar strain rats for 45 days. After 24 h of the last treatment the rats were killed using diethyl ether. The epididymides were removed and cleared from the adhering tissues. Epididymal sperm were collected by cutting the epididymides into small pieces in Ham's F12 medium, and counted using a hemocytometer. The epididymal sperm counts in the TCDD-treated groups decreased in a dose-dependent manner from the control value of 8.2{+-}0.14 x 10{sup 8} to 5.31{+-}0.15 x 10{sup 8}. Since a positive correlation (r=0.95; n=24) was observed between sperm count and DNA content of the epididymal sperm, DNA content was routinely used as an indicator of sperm count, and the results were expressed in terms of both protein and DNA. There was a significant decline in the activities of superoxide dismutase (40{+-}2.17 to 27.1{+-}0.76/mg protein and 32.41 to 18.07{+-}0.76/mg DNA), catalase (2.49{+-}0.13 to 2.03{+-}0.05/mg protein and 2.01{+-}0.05 to 1.35{+-}0.05/mg DNA), glutathione reductase (71.2{+-}3.87 to 48{+-}1.79/mg protein and 57.58{+-}1.52 to 31.94/mg DNA) and glutathione peroxidase (22.4{+-}1.43 to 16.9{+-}1.57/mg protein and 18.08{+-}0.61 to 11.38{+-}1.22/mg DNA) while there were increases in the levels of hydrogen peroxide (20.8{+-}1.96 to 55.3{+-}0.88/ mg protein and 16.18{+-}1.88 to 36.87{+-}0.88/ mg DNA) and lipid peroxidation (2.17{+-}0.2 to 6.08/mg protein and 1.75{+-}0.12 to 4.05{+-}0.12/mg DNA) in the epididymal sperm. The results suggest that graded doses of TCDD elicit depletion of antioxidant defense system

  6. Chronic glucocorticoid exposure-induced epididymal adiposity is associated with mitochondrial dysfunction in white adipose tissue of male C57BL/6J mice.

    Directory of Open Access Journals (Sweden)

    Jie Yu

    Full Text Available Prolonged and excessive glucocorticoids (GC exposure resulted from Cushing's syndrome or GC therapy develops central obesity. Moreover, mitochondria are crucial in adipose energy homeostasis. Thus, we tested the hypothesis that mitochondrial dysfunction may contribute to chronic GC exposure-induced epididymal adiposity in the present study. A total of thirty-six 5-week-old male C57BL/6J mice (∼20 g were administrated with 100 µg/ml corticosterone (CORT or vehicle through drinking water for 4 weeks. Chronic CORT exposure mildly decreased body weight without altering food and water intake in mice. The epididymal fat accumulation was increased, but adipocyte size was decreased by CORT. CORT also increased plasma CORT, insulin, leptin, and fibroblast growth factor 21 concentrations as measured by RIA or ELISA. Interestingly, CORT increased plasma levels of triacylglycerols and nonesterified fatty acids, and up-regulated the expression of both lipolytic and lipogenic genes as determined by real-time RT-PCR. Furthermore, CORT impaired mitochondrial biogenesis and oxidative function in epididymal WAT. The reactive oxygen species production was increased and the activities of anti-oxidative enzymes were reduced by CORT treatment as well. Taken together, these findings reveal that chronic CORT administration-induced epididymal adiposity is, at least in part, associated with mitochondrial dysfunction in mouse epididymal white adipose tissue.

  7. Effect of D-valine and cytosine arabinoside on [3H]thymidine incorporation in rat and rabbit epididymal epithelial cell cultures

    International Nuclear Information System (INIS)

    Orgebin-Crist, M.C.; Jonas-Davies, J.; Storey, P.; Olson, G.E.

    1984-01-01

    Epithelial cell enriched primary cultures were established from the rat and the rabbit epididymis. Epithelial cell aggregates, obtained after pronase digestion of minced epididymis, attached to the culture dish and after 72 h in vitro spread out to form discrete patches of cells. These cells have an epithelioid morphology and form a monolayer of closely apposed polygonal cells where DNA synthesis, as judged by [ 3 H]thymidine uptake, is very low. In L-valine medium the nonepithelial cell contamination was no more than 10% in rat and rabbit epididymal primary cultures. The labeling index of rat epididymal cells cultured in D-valine medium was significantly lower than that of cells cultured in L-valine medium. In contrast, the labeling index of rabbit epididymal cells cultured in D-valine medium was significantly higher than that of cells cultured in L-valine medium. Cytosine arabinoside decreased the number of labeled cells in both L-valine and D-valine cultures. From these results, it appears that D-valine is a selective agent for rat epididymal epithelial cells, but not for rabbit epithelial cells, and that cytosine arabinoside is a simple and effective means to control the proliferation of fibroblast-like cells in both rat and rabbit epididymal cell cultures

  8. Gold in semen: Level in seminal plasma and spermatozoa of normal ...

    African Journals Online (AJOL)

    K.P. Skandhan

    2016-07-01

    Jul 1, 2016 ... Gold level in sediment (spermatozoa) of normal was almost same as observed in its seminal plasma ... showed, gold was not detected in semen by Direct Couple ... Diffraction Analysis, revealed presence of gold throughout.

  9. Interactions among motility, fertilizing ability, and testosterone binding on spermatozoa of bonnet monkey (Macaca radiata).

    Science.gov (United States)

    Warikoo, P K; Majumdar, S S; Allag, I S; Das, R P; Roy, S

    1986-01-01

    Fresh ejaculates of bonnet monkeys were separated into fractions rich with highly motile and sluggishly motile spermatozoa. The motility, ability to fertilize zona-free hamster eggs, and distribution of testosterone-binding sites on spermatozoa were assessed to determine the relation between these sperm functions. Two parameters of objective assessment of motility--velocity and degree of flagellar bending--were significantly correlated with the ability to form pronuclei in zona-free hamster eggs. Only spermatozoa with good motility could form pronuclei, which might be important for assessment of the fertilizing ability. The motility was directly related to the distribution of testosterone-binding sites; the fraction having mostly motile spermatozoa was distributed over the sperm surface. The technique is simple and may be used to evaluate semen of nonhuman primates.

  10. Antimicrobial susceptibility of Haemophilus influenzae strains isolated from the urethra of men with acute urethritis and/or epididymitis.

    Science.gov (United States)

    Deguchi, Takashi; Ito, Shin; Hatazaki, Kyoko; Horie, Kengo; Yasuda, Mitsuru; Nakane, Keita; Mizutani, Kosuke; Tsuchiya, Tomohiro; Yokoi, Shigeaki; Hanaoka, Nozomu; Shimuta, Ken; Ohnishi, Makoto; Muratani, Tetsuro; Nakano, Masahiro

    2017-11-01

    We determined minimum inhibitory concentrations (MICs) of 41 antimicrobial agents for 73 clinical strains of Haemophilus influenzae isolated from the urethra of men with acute urethritis and/or epididymitis and examined the strains for the production of β-lactamase. We also compared their antimicrobial susceptibilities with those of H. influenzae strains from respiratory tract or otorhinolaryngological infections that were reported in Japan. The proportion of β-lactamase-nonproducing ampicillin-resistant strains from acute urethritis and/or epididymitis appeared to be lower, but that of β-lactamase-producing ampicillin-resistant strains appeared to be higher, compared with those from respiratory tract or otorhinolaryngological infections. However, their antimicrobial susceptibilities to a variety of other antimicrobial agents would be similar to those from respiratory tract or otorhinolaryngological infections. Almost all of the strains of H. influenzae from acute urethritis and/or epididymitis were susceptible to the agents, including ceftriaxone, quinolones, macrolides, and tetracyclines, commonly prescribed for treatment of acute urethritis based on the MIC breakpoints recommended by the Clinical and Laboratory Standards Institute. Ceftriaxone and quinolones could be effective on H. influenzae-induced urethritis. However, azithromycin treatment failures were reported in acute urethritis caused by H. influenzae strains considered susceptible to azithromycin. Further studies will be needed to determine MIC breakpoints of antimicrobial agents, which are recommended for treatment of urogenital infections, for H. influenzae strains causing these infections. Nevertheless, this study provides useful data regarding antimicrobial susceptibilities of H. influenzae strains isolated from the urogenital tract, which have rarely been studied. Copyright © 2017 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier

  11. Hatchery-scale trials using cryopreserved spermatozoa of black-lip pearl oyster, Pinctada margaritifera

    OpenAIRE

    Hui, Belinda; Vonau, Vincent; Moriceau, Jacques; Tetumu, Roger; Vanaa, Vincent; Demoy-schneider, Marina; Suquet, Marc; Le Moullac, Gilles

    2011-01-01

    Cryopreservation is a valuable tool for genetic improvement programs. Several bivalve mollusc species have already been the subject of such programs and the Tahitian black pearl oyster industry is now planning the development of selective breeding for desirable traits in Pinctada margaritifera. The ability to cryopreserve spermatozoa would, therefore, offer significant benefits to the cultured black pearl industry. Spermatozoa were cryopreserved with CPA 0.7 M trehalose in 0.8 M Me2SO and...

  12. Fragmentasi DNA Spermatozoa: Penyebab, Deteksi, dan Implikasinya pada Infertilitas Laki-Laki

    Directory of Open Access Journals (Sweden)

    Silvia W. Lestari

    2015-12-01

    Full Text Available Prediksi fertilitas laki-laki dapat dilakukan dengan analisis semen. Analisis semen konvensionalmerupakan pemeriksaan sederhana dan tidak mahal, tetapi memiliki variabilitas yang tinggi.Integritas DNA spermatozoa penting untuk transmisi informasi genetik. Fragmentasi DNAspermatozoa sebagai akibat gangguan spermatogenesis, maturasi spermatozoa, stres oksidatifdan infeksi, dapat menyebabkan infertilitas laki-laki, gangguan perkembangan embrio dan abortusberulang. Hubungan fragmentasi DNA spermatozoa dengan luaran teknologi reproduksi berbantu(TRB mengarahkan fragmentasi DNA spermatozoa sebagai pemeriksaan infertilitas laki-laki. Dariberbagai metode fragmentasi DNA spermatozoa yang umum dilakukan, sperm chromatin dispersion(SCD merupakan metode pemeriksaan fragmentasi DNA spermatozoa yang sederhana, akuratdan tidak mahal, sehingga dapat dilaksanakan di laboratorium andrologi. Selain menghasilkandiagnosis yang lebih baik, pemeriksaan fragmentasi DNA spermatozoa juga menggambarkanprognosis infertilitas termasuk luaran program TRB. Kata kunci: infertilitas laki-laki, fragmentasi DNA spermatozoa, SCD   Sperm DNA Fragmentation: Etiology, Detection and Implicationto Male Infertility Abstract The prediction of male fertility is determined by semen analysis. The conventional semenanalysis is simple and inexpensive but prone to variability. The integrity of sperm DNA is essentialfor the transmission of genetic information. Fragmentation of sperm DNA as result of disruptionin spermatogenesis and sperm maturation, oxidative stress, and infection may lead to maleinfertility, abnormal embryonic development and recurrent abortion. The association betweensperm DNA fragmentation and diminished reproductive outcomes has led to the introduction ofsperm DNA fragmentation testing on the clinical assessment of male infertility. Of all the spermDNA fragmentation tests, sperm chromatin dispersion (SCD test is quite simple, accurate, andinexpensive to be conducted on

  13. Posttranslational Modifications in Spermatozoa and Effects on Male Fertility and Sperm Viability.

    Science.gov (United States)

    Brohi, Rahim Dad; Huo, Li-Jun

    2017-05-01

    Spermatogenesis is a complex and highly regulated process. The ability of spermatozoa to perform its function depends on multiple physiological and genetic factors that are not fully understood. Notably, due to lack of transcriptional and translational activity in spermatozoa, posttranslational modifications (PTMs) play key roles in determining their viability. PTMs not only confer structural changes in the proteome of the spermatozoa cells, but also increase the diversity of the proteome and introduce specific modifications that could be translated into functional changes in the affected spermatozoa. Multiple PTMs of active proteins have been identified in the developing spermatogonia. This review summarizes a diverse range of PTMs taking place in the developing spermatozoa, and analyzes their effects on male fertility and sperm viability. In particular, we discuss how SUMOylation, ubiquitination, phosphorylation, acetylation, glycosylation, and disulphide bond formation in proteins play a role in spermatogenesis, sperm maturation, movement of maturing spermatozoa to epididymis, capacitation, hyperactivation, spermatozoa motility, subversion of immune detection by spermatozoa, sperm to egg recognition and fusion, and the fertilization process. When possible, the specific proteins involved in these processes are highlighted. We point to existing knowledge gaps in the field of proteomics, and provide suggestions for future research on sperm viability and male fertility. We discuss briefly, as an example, the observations in water buffalo, Bubalus bubalis, which provides both meat and milk, and therefore is a reliable source for energy and protein needs of human populations. In conclusions, understanding the ways in which PTMs impact mammalian fertility and reproduction is important to make significant strides for diagnostic and therapeutic strategies in the near future.

  14. pH controls spermatozoa motility in the Pacific oyster (Crassostrea gigas

    Directory of Open Access Journals (Sweden)

    Myrina Boulais

    2018-03-01

    Full Text Available Investigating the roles of chemical factors stimulating and inhibiting sperm motility is required to understand the mechanisms of spermatozoa movement. In this study, we described the composition of the seminal fluid (osmotic pressure, pH, and ions and investigated the roles of these factors and salinity in initiating spermatozoa movement in the Pacific oyster, Crassostrea gigas. The acidic pH of the gonad (5.82±0.22 maintained sperm in the quiescent stage and initiation of flagellar movement was triggered by a sudden increase of spermatozoa external pH (pHe when released in seawater (SW. At pH 6.4, percentage of motile spermatozoa was three times higher when they were activated in SW containing 30 mM NH4Cl, which alkalinizes internal pH (pHi of spermatozoa, compared to NH4Cl-free SW, revealing the role of pHi in triggering sperm movement. Percentage of motile spermatozoa activated in Na+-free artificial seawater (ASW was highly reduced compared to ASW, suggesting that change of pHi triggering sperm motility was mediated by a Na+/H+ exchanger. Motility and swimming speed were highest in salinities between 33.8 and 42.7‰ (within a range of 0 to 50 ‰, and pH values above 7.5 (within a range of 4.5 to 9.5.

  15. Birth of healthy twins after intracytoplasmic sperm injection using ejaculated immotile spermatozoa from a patient with Kartagener's syndrome.

    Science.gov (United States)

    Geber, S; Lemgruber, M; Taitson, P F; Valle, M; Sampaio, M

    2012-05-01

    This case report demonstrates a successful pregnancy after ICSI combined with hypo-osmotic swelling test in a couple with Kartagener's syndrome with complete immotile ejaculated spermatozoa. Our result suggests that even for complete immotile spermatozoa, the use of hypo-osmotic swelling test is a good alternative to identify viable spermatozoa. When associated with ICSI, it can be a valuable tool to get fertilisation and pregnancy. © 2011 Blackwell Verlag GmbH.

  16. AKTIVITAS SPERMATOPROTECTIVE EKSTRAK DAUN JAMBU BIJI (Psidium guajava PADA JUMLAH SPERMATOZOA TIKUS PUTIH TERINDUKSI KADMIUM

    Directory of Open Access Journals (Sweden)

    W Christijanti

    2014-06-01

    Full Text Available Kadmium dapat bersifat meningkatkan aktivitas oksigen reaktif yang memicu munculnya radikal bebas. Zat aktif dalam daun jambu biji bersifat antioksidan yaitu suatu senyawa yang dapat berikatan atau menghambat terbentuknya radikal bebas. Penelitian ini bertujuan untuk mengkaji adanya hubungan  antara ekstrak daun jambu biji dengan jumlah spermatozoa tikus terinduksi kadmium. Penelitian eksperimental ini menggunakan 20 ekor tikus yang terinduksi kadmium melalui air minum dengan dosis 100ppm/ekor/hari. Kelompok kontrol mendapatkan ekstrak daun jambu biji 0 mg/ekor sedangkan kelompok I, II dan III berturut turut adalah 50 mg/ekor, 100 mg/ekor dan 150 mg/ekor. Pemberian ekstrak daun jambu biji selama 30 hari. Data jumlah spermatozoa dan kadar kadmium dianalisis dengan uji korelasi pada taraf uji 5%. Pada hari ke 31, testis diambil untuk diukur kadar kadmium dan jumlah spermatozoanya. Hasil penelitian menunjukkan adanya korelasi antara jumlah spermatozoa dan kadar kadmium dengan nilai sig 0,009 <0,05 dan berbanding terbalik sebesar  -0,567. Hal ini berarti semakin besar kadar kadium dalam testis maka semakin sedikit jumlah spermatozoa tikus dan semakin kecil kadar kadium dalam testis maka semakin banyak jumlah spermatozoa tikus. Simpulan dari penelitian ini adalah ekstrak daun jambu biji dapat berperan sebagai spermatoprotective tikus yang terpapar Cadmium. Cadmium can be used to increase the activity of reactive oxygen that triggers the emergence of free radicals. The active substances in guava leaves is an antioxidant compound that can bind or inhibit the formation of free radicals . This study aims to examine the relationship between guava leaf extract with the amount of rat spermatozoa induced by cadmium. This experimental study used 20 mice induced by cadmium through drinking water at  dose of 100ppm/mice/day. The control group received guava leaf extract of 0 mg/mice , while group I , II and III received 50 mg , 100 mg and 150 mg in 30 days

  17. Pemisahan Spermatozoa Berkromosom X dan Y Kambing Boer dan Aplikasinya Melalui Inseminasi Buatan Untuk Mendapatkan Jenis Kelamin Anak Sesuai Harapan

    Directory of Open Access Journals (Sweden)

    Dasrul Dasrul

    2013-04-01

    Full Text Available Separation of spermatozoa with x and y chromosome at boer goat and its application by artificial insemination for kid sex purpose ABSTRACT. The purposes of this experiment are to investigate the separation of X and Y spermatozoa by measuring the spermatozoa quality, sex ratio between X and Y, capacity of fertility indicated by conception rate and sex ratio of goat boar kids. Samples, used in this experiment, are fresh semen from Boer goat with high quality consists of 4 group treatments with 6 replications 1 group of spermatozoa without separation (control, 2 group of spermatozoa separated by percoll gradient density centrifugation 3 levels (P1, 5 levels (P2 and swine up (P3. The observed parameters are spermatozoa quality, X and Y spermatozoa ratio, fertility’s capacity and sex ratio on the birth. Quality examination of spermatozoa and identify X and Y spermatozoa is based on the standard method of WHO. The conception rate was based on the ratio of pregnant goat after the first insemination. Data of spermatozoa quality and spermatozoa ratio were analyzed by using analisis of variance (ANOVA and further analysis by LSD if there were differences between treatments. The results of this experiment showed that spermatozoa quality Boer goat significantly reduced (p<0,05 after separation with percoll gradient density centrifugation  and swim up. Percentage spermatozoa X after percoll gradient density centrifugation was significantly higher (P<0,05 compared to control and swim up. Meanwhile, the Y spermatozoa population was significantly higher (P<0,05 after swim up treatment compared to percoll gradient density centrifugation and control. The percentage of sex ratio (male: female after insemination from percoll gradient density centrifugation produced more female than male. On the other hand, insemination from swim up produced more male than female. Sex ratio produced from separation of percoll gradient density centrifugation, swim up was

  18. Genotoxicity of diuron and glyphosate in oyster spermatozoa and embryos.

    Science.gov (United States)

    Akcha, F; Spagnol, C; Rouxel, J

    2012-01-15

    We investigated the effects of genotoxicant exposure in gametes and embryos to find a possible link between genotoxicity and reproduction/developmental impairment, and explore the impact of chemical genotoxicity on population dynamics. Our study focused on the genotoxic effects of two herbicides on oyster gametes and embryos: glyphosate (both as an active substance and in the Roundup formulation) and diuron. France is Europe's leading consumer of agrochemical substances and as such, contamination of France's coastal waters by pesticides is a major concern. Glyphosate and diuron are among the most frequently detected herbicides in oyster production areas; as oyster is a specie with external reproduction, its gametes and embryos are in direct contact with the surrounding waters and are hence particularly exposed to these potentially dangerous substances. In the course of this study, differences in genotoxic and embryotoxic responses were observed in the various experiments, possibly due to differences in pollutant sensitivity between the tested genitor lots. Glyphosate and Roundup had no effect on oyster development at the concentrations tested, whereas diuron significantly affected embryo-larval development from the lowest tested concentration of 0.05 μg L⁻¹, i.e. an environmentally realistic concentration. Diuron may therefore have a significant impact on oyster recruitment rates in the natural environment. Our spermiotoxicity study revealed none of the tested herbicides to be cytotoxic for oyster spermatozoa. However, the alkaline comet assay showed diuron to have a significant genotoxic effect on oyster spermatozoa at concentrations of 0.05 μg L⁻¹ upwards. Conversely, no effects due to diuron exposure were observed on sperm mitochondrial function or acrosomal membrane integrity. Although our initial results showed no negative effect on sperm function, the possible impact on fertilization rate and the consequences of the transmission of damaged DNA for

  19. Quantitative analysis of gene-specific DNA damage in human spermatozoa

    International Nuclear Information System (INIS)

    Sawyer, Dennis E.; Mercer, Belinda G.; Wiklendt, Agnieszka M.; Aitken, R. John

    2003-01-01

    Recent studies have suggested that human spermatozoa are highly susceptible to DNA damage induced by oxidative stress. However, a detailed analysis of the precise nature of this damage and the extent to which it affects the mitochondrial and nuclear genomes has not been reported. To induce DNA damage, human spermatozoa were treated in vitro with hydrogen peroxide (H 2 O 2 ; 0-5 mM) or iron (as Fe(II)SO 4 , 0-500 μM). Quantitative PCR (QPCR) was used to measure DNA damage in individual nuclear genes (hprt, β-pol and β-globin) and mitochondrial DNA. Single strand breaks were also assessed by alkaline gel electrophoresis. H 2 O 2 was found to be genotoxic toward spermatozoa at concentrations as high as 1.25 mM, but DNA damage was not detected in these cells with lower concentrations of H 2 O 2 . The mitochondrial genome of human spermatozoa was significantly (P 2 O 2 -induced DNA damage than the nuclear genome. However, both nDNA and mtDNA in human spermatozoa were significantly (P<0.001) more resistant to damage than DNA from a variety of cell lines of germ cell and myoblastoid origin. Interestingly, significant DNA damage was also not detected in human spermatozoa treated with iron. These studies report, for the first time, quantitative measurements of DNA damage in specific genes of male germ cells, and challenge the commonly held belief that human spermatozoa are particularly vulnerable to DNA damage

  20. Resistance of human spermatozoa to cryoinjury in repeated cycles of thaw-refreezing

    Directory of Open Access Journals (Sweden)

    Sidney Verza Jr.

    2009-10-01

    Full Text Available Objective: To study the resistance of human spermatozoa to cryoinjury in repeated cycles of thaw-refreezing by using the fast liquid nitrogen vapor method. Material and Methods: Semen specimens were obtained from sixteen normal and oligozoospermic individuals who required disposal at the sperm bank. Five of them had testicular cancer. Specimens were thawed and an aliquot was removed for analysis. The remaining specimens were refrozen without removing the cryomedia. Repeated freeze-thaw cycles were performed until no motile sperm were observed. Sperm motility, number of motile spermatozoa and viability were determined after thawing. Resistance to cryoinjury was compared between groups and also after each refreezing cycle within groups. Results: Motile spermatozoa were recovered after five and two refreeze-thawing cycles in normozoospermic and oligozoospermic specimens, respectively. There were no significant differences in the recovery of motile spermatozoa between thaws within each group of normal and oligozoospermic specimens, but percentage motility and total number of motile spermatozoa were significantly lower in the oligozoospermic one. Specimens from men with cancer were exposed to six refreeze-thawing cycles. Although recovery of motile spermatozoa was significantly impaired after each thawing, there were no significant differences in the recovery of motile sperm between thaws in cancer and non-cancer groups. Conclusions: Human spermatozoa resist repeated cryopreservation using the fast liquid nitrogen vapor method. Normozoospermic specimens withstand refreezing for an average two cycles longer than oligozoospermic ones. Specimens from cancer patients seem to resist repeated cryoinjury similarly to non-cancer counterparts. Resistance to repeated cryoinjury was related to the initial semen quality.

  1. Migration of fresh and cryopreserved human spermatozoa in polyacrylamide gel.

    Science.gov (United States)

    Goldstein, M C; Wix, L S; Foote, R H; Feldschuh, R; Feldschuh, J

    1982-05-01

    The ability of freshly collected and frozen human spermatozoa to migrate in round capillary tubes containing specially formulated polyacrylamide gel was investigated, using 33 ejaculates from 27 donors. Each semen sample was divided; one portion was left undiluted, and the other portion was diluted to 50 x 10(6) sperm/ml. Glycerol was used as the cryoprotectant. The percentage of motile sperm cells was determined before and after freezing. Fresh semen contained a higher percentage of motile cells, which migrated farther than those of cryopreserved-thawed semen. Various correlations between the percentage of motile sperm and migration distance ranged from 0.57 to 0.62. There was a low positive correlation of migration distance with sperm cell concentration per milliliter, r = 0.25 to 0.34; and thus adjusting semen samples to a standard sperm concentration improved the accuracy of the test only slightly. The regression coefficient of migration distance on the percentage of motile sperm in fresh semen was 0.65, indicating that for each 10% increase in sperm motility, migration distance is predicted to increase 6.5 mm. Five batches of polyacrylamide gel gave uniform results, and the application of this stable gel to fertility investigations is discussed.

  2. Quantitative determination of creatine kinase release from herring (Clupea harengus) spermatozoa induced by tributyltin.

    Science.gov (United States)

    Grzyb, Katarzyna; Rychłowski, Michał; Biegniewska, Anna; Skorkowski, Edward F

    2003-02-01

    Creatine kinase (CK, ATP creatine phosphotransferase, EC 2.7.3.2) is an enzyme participating in ATP regeneration, which is the primary source of energy in living organisms. We demonstrated that CK from herring spermatozoa has high activity ( approximately 452 micromol/min/g of fresh semen) and has a different electrophoretic mobility from isoenzymes present in skeletal muscle. In our study, we investigated toxic effect of tributyltin (TBT) on herring spermatozoa using a specific sperm viability kit to observe live and dead sperm cells with a confocal microscope. Treatment of herring spermatozoa with TBT caused a time-dependent decrease of viability: 35% nonviable cells with 5 microM TBT and more than 90% nonviable cells with 10 microM TBT after 6 h exposure. We also monitored CK release from damaged spermatozoa into surrounding medium containing different concentrations of TBT. The higher concentration of TBT was used the more CK release from spermatozoa was observed. We suggest that CK could be a good biomarker of sperm cell membranes degradation in the case when lactate dehydrogenase release from permeabilized cells is not possible for rapid determination of the effect of TBT.

  3. Sneaker Male Squid Produce Long-lived Spermatozoa by Modulating Their Energy Metabolism.

    Science.gov (United States)

    Hirohashi, Noritaka; Tamura-Nakano, Miwa; Nakaya, Fumio; Iida, Tomohiro; Iwata, Yoko

    2016-09-09

    Spermatozoa released by males should remain viable until fertilization. Hence, sperm longevity is governed by intrinsic and environmental factors in accordance with the male mating strategy. However, whether intraspecific variation of insemination modes can impact sperm longevity remains to be elucidated. In the squid Heterololigo bleekeri, male dimorphism (consort and sneaker) is linked to two discontinuous insemination modes that differ in place and time. Notably, only sneaker male spermatozoa inseminated long before egg spawning can be stored in the seminal receptacle. We found that sneaker spermatozoa exhibited greater persistence in fertilization competence and flagellar motility than consort ones because of a larger amount of flagellar glycogen. Sneaker spermatozoa also showed higher capacities in glucose uptake and lactate efflux. Lactic acidosis was considered to stabilize CO2-triggered self-clustering of sneaker spermatozoa, thus establishing hypoxia-induced metabolic changes and sperm survival. These results, together with comparative omics analyses, suggest that postcopulatory reproductive contexts define sperm longevity by modulating the inherent energy levels and metabolic pathways. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  4. Sneaker Male Squid Produce Long-lived Spermatozoa by Modulating Their Energy Metabolism *

    Science.gov (United States)

    Hirohashi, Noritaka; Tamura-Nakano, Miwa; Nakaya, Fumio; Iida, Tomohiro; Iwata, Yoko

    2016-01-01

    Spermatozoa released by males should remain viable until fertilization. Hence, sperm longevity is governed by intrinsic and environmental factors in accordance with the male mating strategy. However, whether intraspecific variation of insemination modes can impact sperm longevity remains to be elucidated. In the squid Heterololigo bleekeri, male dimorphism (consort and sneaker) is linked to two discontinuous insemination modes that differ in place and time. Notably, only sneaker male spermatozoa inseminated long before egg spawning can be stored in the seminal receptacle. We found that sneaker spermatozoa exhibited greater persistence in fertilization competence and flagellar motility than consort ones because of a larger amount of flagellar glycogen. Sneaker spermatozoa also showed higher capacities in glucose uptake and lactate efflux. Lactic acidosis was considered to stabilize CO2-triggered self-clustering of sneaker spermatozoa, thus establishing hypoxia-induced metabolic changes and sperm survival. These results, together with comparative omics analyses, suggest that postcopulatory reproductive contexts define sperm longevity by modulating the inherent energy levels and metabolic pathways. PMID:27385589

  5. Laser irradiation effects and its possible mechanisms of action on spermatozoa functions in domestic animals

    Directory of Open Access Journals (Sweden)

    S A Lone

    2017-01-01

    Full Text Available This article presents a review pertains the laser irradiation effects and its possible mechanisms of action on spermatozoa functions in domestic animals. To improve artificial insemination, laser is sensitive and cost effective technique, when compared to other conventional methods. Laser may have both positive and negative effects on spermatozoa functions. Since the effects of light are mediated by reactive oxygen species, and the levels of these reactive oxygen species following irradiating spermatozoa with laser may be responsible for determining the effects of laser on sperm. Dose of laser may be regarded as of great significance and this dosage of laser may be responsible for determining its effects on spermatozoa. Optimum dosage of laser for improving seminal attributes may vary among various species and this need to be standardized in each of them. The beneficial effects include improving sperm livability, acrosomal integrity, hypo-osmotic swelling response, mitochondrial function and computer-aided sperm analysis parameters. The increase in cytochrome c oxidase activity, ATP levels and mitochondrial membrane potential, in laser irradiated cells may be responsible for enhanced sperm quality parameters. Improving fertility with laser irradiated spermatozoa has been reported in few species like boar and need to be elaborated in other species. In conclusion laser may be regarded as an easy, cheap and time saving technology for improving artificial insemination; in addition, laser may have various potential applications in the field of reproductive biotechnology as well as in livestock farms and veterinary polyclinics.

  6. Effects of alginate on frozen-thawed boar spermatozoa quality, lipid peroxidation and antioxidant enzymes activities.

    Science.gov (United States)

    Hu, Jinghua; Geng, Guoxia; Li, Qingwang; Sun, Xiuzhu; Cao, Hualin; Liu, Yawei

    2014-06-30

    Although alginate was reported to play an important role as free radical scavengers in vitro and could be used as sources of natural antioxidants, there was no study about the cryoprotective effects of alginate on boar spermatozoa freezing. The objective of this research was to evaluate the effects of different concentrations of alginate added to the freezing extenders on boar spermatozoa motility, plasma membrane integrity, acrosomal integrity, mitochondrial activities, lipid peroxidation and antioxidative enzymes activities (SOD and GSH-Px) after thawing. Alginate was added to the TCG extender to yield six different final concentrations: 0, 0.2, 0.4, 0.6, 0.8, and 1.0mg/mL. The semen extender supplemented with various doses of alginate increased (Pboar spermatozoa acrosomal integrity at concentrations of 0.6, 0.8, 1.0mg/mL, compared with that of the control (Pextenders with the presence of alginate led to higher SOD and GSH-Px activities and lower MDA levels, in comparison to the control (Pboar spermatozoa motility, functional integrity and antioxidative capacity at appropriate concentrations. Therefore alginate could be employed as an effective cryoprotectant in boar spermatozoa cryopreservation. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. MeCP2 Expression and Promoter Methylation of Cyclin D1 Gene Are Associated with Cyclin D1 Expression in Developing Rat Epididymal Duct

    International Nuclear Information System (INIS)

    Darwanto, Agus; Kitazawa, Riko; Mori, Kiyoshi; Kondo, Takeshi; Kitazawa, Sohei

    2008-01-01

    Hypermethylation-dependent silencing of the gene is achieved by recruiting methyl-CpG binding proteins (MeCPs). Among the MeCPs, MeCP2 is the most abundantly and ubiquitously expressed in various types of cells. We first screened the distribution and expression pattern of MeCP2 in adult and developing rat tissues and found strong MeCP2 expression, albeit rather ubiquitously among normal tissues, in ganglion cells and intestinal epithelium in the small intestine, in Purkinje cells and neurons in the brain, in spermatogonia and in epithelial cells in the epididymal duct of the testis. We then assessed the expression and the methylation pattern of the promoter region of cyclin D1 by immunohistochemistry and sodium bisulfite mapping, and found that cyclin D1 expression in the epididymal duct decreased rapidly during rat development: strong in newborn rats and very weak or almost negative in 7-day-old rats. Mirroring the decrease of cyclin D1 expression, methylated cytosine at both CpG and non-CpG loci in the cyclin D1 promoter was frequently observed in the epididymal duct of 7-day-old rats but not in that of newborn rats. Interestingly, MeCP2 expression also increased concomitant with the increase of methylation. Cyclin D1 expression in the epididymal duct may be efficiently regulated by the epigenetic mechanism of the cooperative increase of MeCP2 expression and promoter methylation

  8. False-positive “halo” sign on testicular scintigraphy in a 5-year-old boy with epididymitis and hydrocele

    International Nuclear Information System (INIS)

    Santhosh, Sampath; Guha, Poonam; Bhattacharya, Anish; Bawa, Monika; Mittal, Bhagwant Rai

    2011-01-01

    Scintigraphic differentiation between acute torsion, hydrocele and testicular or scrotal abscess can be difficult. Doppler sonography may provide useful complimentary information toward diagnosis. The authors describe a 5-year-old child where epididymitis with hydrocele was misdiagnosed as testicular torsion on scrotal scintigraphy

  9. Hericium caput-medusae (Bull.:Fr.) Pers. polysaccharide enhance innate immune response, immune-related genes expression and disease resistance against Aeromonas hydrophila in grass carp (Ctenopharyngodon idella).

    Science.gov (United States)

    Gou, Changlong; Wang, Jiazhen; Wang, Yuqiong; Dong, Wenlong; Shan, Xiaofeng; Lou, Yujie; Gao, Yunhang

    2018-01-01

    The objective was to add 0, 400, 800 or 1200 mg/kg of Hericium caput-medusae polysaccharide (HCMP) to the basal diet of grass carp (Ctenopharyngodon idella) and determine effects on humoral innate immunity, expression of immune-related genes and disease resistance. Adding HCMP enhanced (P < 0.05) bactericidal activity at 1, 2 and 3 weeks and also lysozyme activity, complement C3, and SOD activity at 2 and 3 weeks. Supplementing 800 or 1200 mg/kg of HCMP for 2 or 3 weeks increased (P < 0.05) serum concentrations of total protein, albumin and globulin. Two immune-related genes (IL-1β and TNF-α) were up-regulated (P < 0.05) in HCMP supplemented groups given 800 or 1200 mg/kg HCMP after 2 and 3 weeks of feeding. Expression of anti-inflammatory cytokine IL-10 was down-regulated (P < 0.05) after receiving 800 or 1200 mg/kg HCMP for 2 or 3 weeks. Fish fed 800 mg/kg HCMP had maximal disease resistance against Aeromonas hydrophila (65.4%). In conclusion, HCMP enhanced immune response and expression of immune-related genes and increased disease resistance against Aeromonas hydrophila in grass carp, with greatest effects in fish given 800 mg/kg HCMP for 3 weeks. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Effects of storage in different semen extenders on the pre-freezing and post-thawing quality of boar spermatozoa.

    Science.gov (United States)

    Dziekońska, A; Zasiadczyk, Ł; Lecewicz, M; Strzeżek, R; Koziorowska-Gilun, M; Fraser, L; Mogielnicka-Brzozowska, M; Kordan, W

    2015-01-01

    The aim of this study was to investigate the effects of storage of semen in different commercial extenders on the pre-freezing and post-thawing quality of boar spermatozoa. Semen was diluted in BTS, Androhep (AH) and Gedil (GD), stored for 24 h at 17°C, and then frozen in accordance with the cryopreservation protocol. Analyses of the quality of spermatozoa included: motility, normal apical ridge (NAR) acrosome, plasma membrane integrity (PMI), mitochondrial membrane potential (MMP), measurements of ATP content and activity of superoxidase dismutase (SOD) and glutathione peroxidase (GPx). Prior to the freezing process, no significant effect of the extender on the sperm quality parameters was noted. After thawing the spermatozoa it was demonstrated that the type of extender used influenced PMI, MMP, ATP content and activity of GPx. In the AH extender the percentage of spermatozoa with PMI and ATP content in spermatozoa was significantly higher (Pextenders. In addition, semen stored in the AH was characterised by a statistically higher (Pboar spermatozoa stored for 24 hours in liquid state can be used. However, the type of extender used prior to freezing may have a significant effect on the post-thawing quality of the spermatozoa. The AH extender better secured the quality of thawed boar spermatozoa as compared with the BTS or GD.

  11. Comparison of spermatozoa parameters, fine structures, and energy-related factors among tetraploid, hyper-tetraploid, and hyper-triploid loaches (Misgurnus anguillicaudatus).

    Science.gov (United States)

    Zhao, Yan; Saito, Taiju; Pšenička, Martin; Fujimoto, Takafumi; Arai, Katsutoshi

    2014-04-01

    To evaluate the influence of ploidy elevation and aneuploidy on spermatozoa in the loach Misgurnus anguillicaudatus, we investigated some parameters (motility, concentration, and viability), fine structures (gross morphology, head size, and flagellum length), and energy-related biochemical factors (volume of mitochondrial mass per cell and ATP content) in diploid, hyper-diploid, and hexaploid-range spermatozoa produced in natural tetraploid, hyper-tetraploid, and hyper-triploid male loaches, respectively. Diploid spermatozoa exhibited vigorous movement and sufficient duration of motility similar to those in haploid spermatozoa. They had longer flagella, higher numbers and larger volume of mitochondria, and higher ATP content than haploid spermatozoa of wild-type diploids. No differences were observed in parameters and morphological characteristics between diploid and hyper-diploid spermatozoa. In contrast, the hexaploid-range spermatozoa of hyper-triploid males exhibited poor progressive motility in spite of a higher ATP content of spermatozoa. Spermatozoa with no flagella (36.0%) or multiple flagella (18.6%) were also observed in hyper-triploids. Ratios of head to flagellum length in hexaploid-range spermatozoa were significantly different from those of haploid spermatozoa. In addition to the normal 9+2 microtubule structure of the flagellum, an abnormal 9+1 microtubule structure was also observed in the spermatozoa of hyper-triploids. © 2014 Wiley Periodicals, Inc.

  12. Effect of Air Space in Storage Vials on Motility of Spermatozoa in Chilled Buck Semen

    Directory of Open Access Journals (Sweden)

    Magnus Paul K and Lali F Anand 1

    Full Text Available This study was conducted in order to find out the effect of air space on the top of glass vial in which semen is stored, on the motility of spermatozoa. 45 samples collected from two bucks over a span of 6 months were used for experiment. Goat milk extender was the diluent used. Two ml each of diluted semen after noting their initial motility was stored in 2 ml and 5 ml vials. Samples were stored at 5°C and motility of spermatozoa noted at 24 and 48 hours. Semen without air space was found to preserve the motility better than semen with air space on 24 and 48 hours of incubation. This could be better attributed to reactive oxygen species production by the spermatozoa, but further investigation is needed in this aspect to confirm it. [Veterinary World 2010; 3(9.000: 421-423

  13. Evaluation of cheetah and leopard spermatozoa developmental capability after interspecific ICSI with domestic cat oocytes.

    Science.gov (United States)

    Moro, L N; Sestelo, A J; Salamone, D F

    2014-08-01

    The ICSI procedure is potentially of great value for felids, and it has not been extensively studied in these species. The objectives of this work were to determine the best conditions for ICSI in the domestic cat (DC) to generate interspecific embryos by injecting cheetah (Ch) and leopard (Leo) spermatozoa. Firstly, DC oocytes were matured with insulin-transferrin-selenium (ITS) or without it (MM) and cultured using atmospheric (21%) or low (5%) oxygen tension after ICSI. The group ITS-5%O2 showed the highest blastocyst rate (p cheetah and leopard spermatozoa were able to generate blastocysts without artificial activation, which suggests that developmental capacity of wild felid spermatozoa can be evaluated by interspecific ICSI. This technique should be used to assist wild felid reproduction. © 2014 Blackwell Verlag GmbH.

  14. MiR-200a is involved in rat epididymal development by targeting β-catenin mRNA

    Institute of Scientific and Technical Information of China (English)

    Xiaojiang Wu; Botao Zhao; Wei Li; Yue Chen; Ruqiang Liang; Lin Li; Youxin Jin; Kangcheng Ruan

    2012-01-01

    The expression of 350 microRNAs (miRNAs) in epididymis of rat from postnatal development to adult (from postnatal days 7-70) was profiled with home-made miRNA microarray.Among them,48 miRNAs changed significantly, in which the expression of miR-200a increased obviously with time,in a good agreement with that obtained from northern blot analysis.The real-time quantitative-polymerase chain reaction result indicated that temporal expression of rat β-catenin was exactly inversed to that of miR-200a during rat epididymal development,implying that miR-200a might also target β-catenin mRNA in rat epididymis as reported by Saydam et al.in humans.The bioinformatic analysis indicated that 3' untranslated region of rat β-catenin mRNA did contain a putative binding site for miR-200a.Meanwhile,it was found that the sequence of this binding site was different from that of human β-catenin mRNA with a deletion of two adjacent nucleotides (U and C).But the results of luciferase targeting assay in HEK 293T cells and the overexpression of miR-200a in rat NRK cells demonstrated that miR-200a did target rat β-catenin mRNA and cause the suppression of its expression.All these results show that miR-200a should be involved in rat epididymal development by targeting β-catenin mRNA of rat and suppressing its expression.

  15. Characterization of NAADP-mediated calcium signaling in human spermatozoa

    Energy Technology Data Exchange (ETDEWEB)

    Sánchez-Tusie, A.A. [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico); Vasudevan, S.R.; Churchill, G.C. [Department of Pharmacology, University of Oxford, Oxford OX1 3QT, England (United Kingdom); Nishigaki, T. [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico); Treviño, C.L., E-mail: ctrevino@ibt.unam.mx [Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos (Mexico)

    2014-01-10

    Highlights: •Human sperm cells synthesize NAADP. •NAADP-AM mediates [Ca{sup 2+}]{sub i} increases in human sperm in the absence of [Ca{sup 2+}]{sub o}. •Human sperm have two acidic compartments located in the head and midpiece. -- Abstract: Ca{sup 2+} signaling in spermatozoa plays a crucial role during processes such as capacitation and release of the acrosome, but the underlying molecular mechanisms still remain unclear. Nicotinic acid adenine dinucleotide phosphate (NAADP) is a potent Ca{sup 2+}-releasing second messenger in a variety of cellular processes. The presence of a NAADP synthesizing enzyme in sea urchin sperm has been previously reported, suggesting a possible role of NAADP in sperm Ca{sup 2+} signaling. In this work we used in vitro enzyme assays to show the presence of a novel NAADP synthesizing enzyme in human sperm, and to characterize its sensitivity to Ca{sup 2+} and pH. Ca{sup 2+} fluorescence imaging studies demonstrated that the permeable form of NAADP (NAADP-AM) induces intracellular [Ca{sup 2+}] increases in human sperm even in the absence of extracellular Ca{sup 2+}. Using LysoTracker®, a fluorescent probe that selectively accumulates in acidic compartments, we identified two such stores in human sperm cells. Their acidic nature was further confirmed by the reduction in staining intensity observed upon inhibition of the endo-lysosomal proton pump with Bafilomycin, or after lysosomal bursting with glycyl-L-phenylalanine-2-naphthylamide. The selective fluorescent NAADP analog, Ned-19, stained the same subcellular regions as LysoTracker®, suggesting that these stores are the targets of NAADP action.

  16. Effects of semen preservation on boar spermatozoa head membranes.

    Science.gov (United States)

    Buhr, M M; Canvin, A T; Bailey, J L

    1989-08-01

    Head plasma membranes were isolated from the sperm-rich fraction of boar semen and from sperm-rich semen that had been subjected to three commercial preservation processes: Extended for fresh insemination (extended), prepared for freezing but not frozen (cooled), and stored frozen for 3-5 weeks (frozen-thawed). Fluorescence polarization was used to determine fluidity of the membranes of all samples for 160 min at 25 degrees C and also for membranes from the sperm-rich and extended semen during cooling and reheating (25 to 5 to 40 degrees C, 0.4 degrees C/min). Head plasma membranes from extended semen were initially more fluid than from other sources (P less than 0.05). Fluidity of head membranes from all sources decreased at 25 degrees C, but the rate of decrease was significantly lower for membranes from cooled and lower again for membranes from frozen-thawed semen. Cooling to 5 degrees C reduced the rate of fluidity change for plasma membranes from the sperm-rich fraction, while heating over 30 degrees C caused a significantly greater decrease. The presence of Ca++ (10 mM) lowered the fluidity of the head plasma membranes from sperm-rich and extended semen over time at 25 degrees C but did not affect the membranes from the cooled or frozen-thawed semen. The change in head plasma membrane fluidity at 25 degrees C may reflect the dynamic nature of spermatozoa membranes prior to fertilization. Extenders, preservation processes and temperature changes have a strong influence on head plasma membrane fluidity and therefore the molecular organization of this membrane.

  17. Slow cryopreservation is not superior to vitrification in human spermatozoa; an experimental controlled study

    Directory of Open Access Journals (Sweden)

    Mohamed Shehata Ali Mohamed

    2015-10-01

    Full Text Available Background: Spermatozoa cryopreservation is used for the management of infertility and some other medical conditions. The routinely applied cryopreservation technique depends on permeating cryoprotectants, whose toxic effects have raised the attention towards permeating cryoprotectants-free vitrification technique. Objective: To compare between the application of slow cryopreservation and vitrification on human spermatozoa. Materials and Methods: This was an experimental controlled study involving 33 human semen samples, where each sample was divided into three equal parts; fresh control, conventional slow freezing, and permeating cryoprotectants-free vitrification. Viability and mitochondrial membrane potential (MMP of control and post-thawing spermatozoa were assessed with the sperm viability kit and the JC-1 kit, respectively, using fluorescence-activated cell sorting analysis. Results: Significant reduction of the progressive motility, viability and MMP was observed by the procedure of freezing and thawing, while there was not any significant difference between both cryopreservation techniques. Cryopreservation resulted in 48% reduction of the percentage of viable spermatozoa and 54.5% rise in the percentage of dead spermatozoa. In addition, high MMP was reduced by 24% and low MMP was increased by 34.75% in response to freezing and thawing. Progressive motility of spermatozoa was correlated significantly positive with high MMP and significantly negative with low MMP in control as well as post-thawing specimens (r=0.8881/ -0.8412, 0.7461/ -0.7510 and 0.7603/ -0.7839 for control, slow and vitrification respectively, p=0.0001. Conclusion: Although both cryopreservation techniques have similar results, vitrification is faster, easier and associated with less toxicity and costs. Thus, vitrification is recommended for the clinical application.

  18. Methyl-parathion decreases sperm function and fertilization capacity after targeting spermatocytes and maturing spermatozoa

    International Nuclear Information System (INIS)

    Pina-Guzman, B.; Sanchez-Gutierrez, M.; Marchetti, F.; Hernandez-Ochoa, I.; Solis-Heredia, M.J.; Quintanilla-Vega, B.

    2009-01-01

    Paternal germline exposure to organophosphorous pesticides (OP) has been associated with reproductive failures and adverse effects in the offspring. Methyl-parathion (Me-Pa), a worldwide-used OP, has reproductive adverse effects and is genotoxic to sperm, possibly via oxidative damage. This study investigated the stages of spermatogenesis susceptible to be targeted by Me-Pa exposure that impact on spermatozoa function and their ability to fertilize. Male mice were exposed to Me-Pa (20 mg/kg bw, i.p.) and spermatozoa from epididymis-vas deferens were collected at 7 or 28 days post-treatment (dpt) to assess the effects on maturing spermatozoa and spermatocytes, respectively. Spermatozoa were examined for DNA damage by nick translation (NT-positive cells) and SCSA (%DFI), lipoperoxidation (LPO) by malondialdehyde production, sperm function by spontaneous- and induced-acrosome reactions (AR), mitochondrial membrane potential (MMP) by using the JC-1 fluorochrome, and fertilization ability by an in vitro assay and in vivo mating. Alterations on DNA integrity (%DFI and NT-positive cells) in spermatozoa collected at 7 and 28 dpt, and decreases in sperm quality and induced-AR were observed; reduced MMP and LPO were observed at 7 dpt only. Negative correlations between LPO and sperm alterations were found. Altered sperm functional parameters evaluated either in vitro or in vivo were associated with reduced fertilization rates at both times. These results show that Me-Pa exposure of maturing spermatozoa and spermatocytes affects many sperm functional parameters that result in a decreased fertilizing capacity. Oxidative stress seems to be a likely mechanism of the detrimental effects of Me-Pa exposure in male germ cells.

  19. The effect of the freezing curve type on bull spermatozoa motility after thawing

    Directory of Open Access Journals (Sweden)

    Martina Doležalová

    2015-01-01

    Full Text Available The objective of this work was to determine the effect of selected freezing curves on spermatozoa survivability after thawing, defined by its motility. The ejaculates of nine selected sires of the same age, breed, and frequency of collecting, bred under the same breeding conditions including handling, stabling, feeding system and feeding ratio composition, were repeatedly collected and evaluated. Sperm samples of each sire were diluted using only one extender and divided into four parts. Selected four freezing curves – the standard, commercially recommended three-phase curve; a two-phase curve; a slow three-phase curve; and a fast three-phase curve, differing in the course of temperature vs time, were applied. The percentage rate of progressive motile spermatozoa above head was determined immediately after thawing, and after 30, 60, 90, and 120 min of the thermodynamic test (TDT. Moreover, average spermatozoa motility (AMOT and spermatozoa motility decrease (MODE throughout the entire TDT were evaluated. Insemination doses frozen using the simpler two-phase curve demonstrated the highest motility values (+2.97% to +10.37%; P < 0.05–0.01 immediately after thawing and during the entire TDT. Concurrently, the highest AMOT (+4.37% to +8.82%; P < 0.01 was determined. The highest spermatozoa motility values were detected after thawing doses frozen by the two-phase freezing curve in eight out of nine sires. Simultaneously, a significant effect of sire individuality was clearly confirmed. Inter-sire differences of spermatozoa motility during TDT as well as AMOT and MODE were significant (P < 0.01. The findings describing both factors of interaction indicate the necessity of individual cryopreservation of the ejaculate to increase its fertilization capability after thawing.

  20. Identification of the inorganic pyrophosphate metabolizing, ATP substituting pathway in mammalian spermatozoa.

    Science.gov (United States)

    Yi, Young-Joo; Sutovsky, Miriam; Kennedy, Chelsey; Sutovsky, Peter

    2012-01-01

    Inorganic pyrophosphate (PPi) is generated by ATP hydrolysis in the cells and also present in extracellular matrix, cartilage and bodily fluids. Fueling an alternative pathway for energy production in cells, PPi is hydrolyzed by inorganic pyrophosphatase (PPA1) in a highly exergonic reaction that can under certain conditions substitute for ATP-derived energy. Recombinant PPA1 is used for energy-regeneration in the cell-free systems used to study the zymology of ATP-dependent ubiquitin-proteasome system, including the role of sperm-borne proteasomes in mammalian fertilization. Inspired by an observation of reduced in vitro fertilization (IVF) rates in the presence of external, recombinant PPA1, this study reveals, for the first time, the presence of PPi, PPA1 and PPi transporter, progressive ankylosis protein ANKH in mammalian spermatozoa. Addition of PPi during porcine IVF increased fertilization rates significantly and in a dose-dependent manner. Fluorometric assay detected high levels of PPi in porcine seminal plasma, oviductal fluid and spermatozoa. Immunofluorescence detected PPA1 in the postacrosomal sheath (PAS) and connecting piece of boar spermatozoa; ANKH was present in the sperm head PAS and equatorial segment. Both ANKH and PPA1 were also detected in human and mouse spermatozoa, and in porcine spermatids. Higher proteasomal-proteolytic activity, indispensable for fertilization, was measured in spermatozoa preserved with PPi. The identification of an alternative, PPi dependent pathway for ATP production in spermatozoa elevates our understanding of sperm physiology and sets the stage for the improvement of semen extenders, storage media and IVF media for animal biotechnology and human assisted reproductive therapies.

  1. Identification of the inorganic pyrophosphate metabolizing, ATP substituting pathway in mammalian spermatozoa.

    Directory of Open Access Journals (Sweden)

    Young-Joo Yi

    Full Text Available Inorganic pyrophosphate (PPi is generated by ATP hydrolysis in the cells and also present in extracellular matrix, cartilage and bodily fluids. Fueling an alternative pathway for energy production in cells, PPi is hydrolyzed by inorganic pyrophosphatase (PPA1 in a highly exergonic reaction that can under certain conditions substitute for ATP-derived energy. Recombinant PPA1 is used for energy-regeneration in the cell-free systems used to study the zymology of ATP-dependent ubiquitin-proteasome system, including the role of sperm-borne proteasomes in mammalian fertilization. Inspired by an observation of reduced in vitro fertilization (IVF rates in the presence of external, recombinant PPA1, this study reveals, for the first time, the presence of PPi, PPA1 and PPi transporter, progressive ankylosis protein ANKH in mammalian spermatozoa. Addition of PPi during porcine IVF increased fertilization rates significantly and in a dose-dependent manner. Fluorometric assay detected high levels of PPi in porcine seminal plasma, oviductal fluid and spermatozoa. Immunofluorescence detected PPA1 in the postacrosomal sheath (PAS and connecting piece of boar spermatozoa; ANKH was present in the sperm head PAS and equatorial segment. Both ANKH and PPA1 were also detected in human and mouse spermatozoa, and in porcine spermatids. Higher proteasomal-proteolytic activity, indispensable for fertilization, was measured in spermatozoa preserved with PPi. The identification of an alternative, PPi dependent pathway for ATP production in spermatozoa elevates our understanding of sperm physiology and sets the stage for the improvement of semen extenders, storage media and IVF media for animal biotechnology and human assisted reproductive therapies.

  2. Reproductive Parameters of the West African Dwarf Bucks Raised in ...

    African Journals Online (AJOL)

    Reproductive samples (scrotal circumference, testicular density, scrotal weight, left and right testicular weights, left and right tunica weights, left and right testicular length, left and right caput epididymal weights, left and right corpus epididymal weights, left and right caudal epididymal weights) from the bucks were collected ...

  3. Validation of simple and cost-effective stains to assess acrosomal status, DNA damage and mitochondrial activity in rooster spermatozoa.

    Science.gov (United States)

    Rui, Bruno R; Angrimani, Daniel S R; Losano, João Diego A; Bicudo, Luana de Cássia; Nichi, Marcílio; Pereira, Ricardo J G

    2017-12-01

    Several methods have been developed to evaluate spermatozoa function in birds but many of these are sometimes complicated, costly and not applicable to field studies (i.e., performed within poultry breeding facilities). The objective was, therefore, to validate efficient, practical and inexpensive procedures to determine DNA fragmentation, acrosomal integrity, and mitochondrial activity in poultry spermatozoa. Initially, ejaculates were individually diluted and divided into control (4°C, 4h) and UV-irradiated aliquots (room temperature, 4h), and then samples containing different percentages of DNA-damaged spermatozoa (0%, 25%, 50%, 75% and 100%) were subjected to Toluidine Blue (TB) and Sperm Chromatin Dispersion assessments (SCD). Fast Green-Rose Bengal (FG-RB) and FITC-PSA staining protocols were subsequently used to assess acrosome status in aliquots comprising assorted amounts of acrosome-reacted spermatozoa. Furthermore, to validate 3,3'-diaminobenzidine (DAB) assay, ejaculates containing different gradients of spermatozoa with great amounts of mitochondrial activity were concurrently evaluated using DAB and JC-1 stains. The proportion of spermatozoa with abnormal DNA integrity when evaluated using the TB assessment correlated significantly with the expected percentages of UV-irradiated spermatozoa and with SCD results. A significant linear regression coefficient was also observed between expected amounts of acrosome-intact spermatozoa and FG-RB readings, and there was a significant correlation of the data when FG-RB and FITC-PSA were used. Likewise, the use of the DAB assay enabled for accurately ascertaining percentages of rooster spermatozoa with greater and lesser mitochondrial function, and results were highly correlated to results with staining with JC-1. Altogether, findings of the present study indicate acrosomal status, DNA integrity and mitochondrial activity in rooster spermatozoa can be easily and reliably determined using FG-RB, TB and DAB stains

  4. Effect of Number of Spermatozoa, Oviduct Condition and Timing of Artificial Insemination on Fertility and Fertile Period of Kampung Rooster Spermatozoa

    Directory of Open Access Journals (Sweden)

    DM Saleh

    2012-01-01

    Full Text Available Abstract. This study was carried out to determine the optimum fertility and fertile period using the number of spermatozoa, oviduct condition and timing of insemination of native rooster spermatozoa. Ninety six commercial Isa brown pullets and nine kampung roosters were used in this study in a 3×2×2 factorial arrangement with one bird in a cage constituting a unit. The factor levels were the number of spermatozoa (50, 100 and 150 million/0.1 ml, oviduct condition (hard-shelled eggs and free hard-shelled eggs, and timing of artificial insemination (in the morning, at 7 AM and in the afternoon, at 4 PM.  The results showed that among the treatments there was no significant interaction to fertility and fertile period. Insemination with 50 million sperm number seemed to be the same result with the other 2 treatments. Oviduct condition had a highly significant difference on fertility and fertile period percentage, and timing of insemination did not differ between morning and afternoon.  In conclusion, the only oviduct condition (free hard-shelled eggs was the best results for insemination in terms of fertility and fertile period of native roosters.  It is recommended that for the maximum fertility and fertile period, hens should be inseminated with 50 million spermatozoa, free of hard-shelled eggs and insemination performed in the morning or in the afternoon.   Keywords: timing of artificial insemination, fertility, fertile period, semen dose, oviduct condition Animal Production 14(1:32-36, January 2012

  5. Spermatozoa of the shrew, Suncus murinus, undergo the acrosome reaction and then selectively kill cells in penetrating the cumulus oophorus.

    Science.gov (United States)

    Kaneko, T; Iida, H; Bedford, J M; Mōri, T

    2001-08-01

    In the musk shrew, Suncus murinus (and other shrews), the cumulus oophorus is ovulated as a discrete, compact, matrix-free ball of cells linked by specialized junctions. In examining how they penetrate the cumulus, Suncus spermatozoa were observed to first bind consistently by the ventral face over the acrosomal region to the exposed smooth surface of a peripheral cumulus cell. This was apparently followed by point fusions between the plasma and outer acrosomal membranes. Thereafter, spermatozoa without acrosomes were observed within cumulus cells that displayed signs of necrosis, as did some radially neighboring cumulus cells linked by zona adherens and gap junctions. Eventually, penetration of spermatozoa as far as the perizonal space around the zona pellucida left linear tracks of locally necrotic cells flanked by normal cumulus cells. Based on these and previous observations, we conclude that the acrosome reaction in Suncus is always induced by cumulus cells, and that reacted spermatozoa penetrate the cumulus by selective invasion and killing of cumulus cells along a linear track. Loss of the acrosome also exposes an apical body/perforatorium that is covered with barbs that appear to assist reacted fertilizing spermatozoa in binding to the zona pellucida. Because fertilized eggs displayed no other spermatozoa within or bound to the zona, an efficient block to polyspermy must prevent such binding of additional spermatozoa.

  6. Radiolabeling of mammalian spermatozoa and their use to monitor sperm transport in females

    International Nuclear Information System (INIS)

    Lorton, S.P.; Gatley, S.J.; Lieberman, L.M.; First, N.L.

    1980-01-01

    Radiolabeling of mammalian spermatozoa with 131 I, 67 Ga, 111 In, and /sup 99m/Tc was investigated. Spermatozoa were labeled with /sup 99m/Tc for in vivo studies because of a high labeling yield (70 to 90%) combined with the lack of impairment of sperm motility. Ovariectomized sheep were brought into estrus by sequential administration of progesterone and estradiol cyprionate. Sheep were necropsied up to 6 hours after insemination with /sup 99m/Tc-labeled ram sperm and their reproductive tracts were resected and examined with a rectilinear scanner. Radioactivity was clearly observed in the fallopian tubes, with larger amounts in the vaginas, cervices, and uteri. In contrast, when /sup 99m/Tc-spermatozoa were replaced with /sup 99m/TcO 4 - , much less radioactivity remained in the reproductive tract at resection and this was evenly distributed. Some label left the /sup 99m/Tc-spermatozoa in vivo, but radioactivity remained on the cells long enough to consider attempting to monitor sperm transport in vivo in a suitable species

  7. FREQUENCY OF ANEUPLOID SPERMATOZOA STUDIED BY MULTICOLOR FISH IN SERIAL SEMEN SAMPLES

    Science.gov (United States)

    Frequency of aneuploid spermatozoa studied by multicolor FISH in serial semen samplesM. Vozdova1, S. D. Perreault2, O. Rezacova1, D. Zudova1 , Z. Zudova3, S. G. Selevan4, J. Rubes1,51Veterinary Research Institute, Brno, Czech Republic; 2U.S. Environmental Protection A...

  8. Effects of low dose gamma- and UV-radiation on sea urchin eggs and spermatozoa

    International Nuclear Information System (INIS)

    Czihak, G.K.

    1991-01-01

    The paper outlines the results of a study of the effects of low dose gamma-and UV-irradiation on sea urchin eggs and spermatozoa with particular reference to the effects on the stages of the mitotic cycle and individual susceptibility. (UK)

  9. [Comparison of reactive oxygen species production in neat semen and washed spermatozoa].

    Science.gov (United States)

    Svobodová, M; Oborná, I; Fingerová, H; Novotný, J; Brezinová, J; Radová, L; Vyslouzilová, J; Horáková, J; Grohmannová, J

    2009-12-01

    To determine Reactive Oxygen Species (ROS) production in neat semen and spermatozoa suspension using chemiluminescence and to examine correlation between both methods. Prospective laboratory study. Department of Obstetric and Gynecology, University Hospital, Olomouc. The study included fertile volunteers (FV, n = 17), men from infertile couples (NM, n = 19) and men with idiopathic infertility (NMI, n = 15). ROS levels were determined by the same method in neat and washed semen samples. The ROS production in neat semen was lower than that in spermatozoa suspension. There was no significant diference in ROS production between volunteers and males from infertile couples. There was a significant correlation between log ROS in neat semen and in spermatozoa suspension in studied groups (FV r = 0.85, p = 1.5 x 10(-5); NM r = 0.76, p neat semen is simpler, faster and better reflecting the actual level of oxidative stress than the same measurement in spermatozoa suspension. The implementation of this method can complement the algorithm of diagnostics and treatment of male infertility and be helpful in selection of patients for antioxidant or antibiotic treatment.

  10. Oviduct Binding and Elevated Environmental pH Induce Protein Tyrosine Phosphorylation in Stallion Spermatozoa

    NARCIS (Netherlands)

    Leemans, B.; Gadella, B.M.; Sostaric, E.; Nelis, H.; Stout, T.A.E.; Hoogewijs, M.; van Soom, A.

    2014-01-01

    Sperm-oviduct binding is an essential step in the capacitation process preparing the sperm for fertilization in several mammalian species. In many species, capacitation can be induced in vitro by exposing spermatozoa to bicarbonate, Ca2+, and albumin; however, these conditions are insufficient in

  11. FRAKSI ETANOL 96% BIJI KORO BENGUK (Mucuna pruriens L. SEBAGAI PENINGKAT KUALITAS SPERMATOZOA MENCIT (Mus musculus

    Directory of Open Access Journals (Sweden)

    Sri Winarni

    2012-11-01

    Full Text Available Background: The examination of sperm quality is the main priority for infertility diagnosis. Based on previous study with mice, active ingredient of Mucuna pruriens L. or koro benguk (Papilionaceae, the L-dopa, may affect the quality of spermatozoa.Objective: Research was to study the effect of 96% ethanol fraction Mucuna pruriens seed on spermatozoaquality of mice exposed to 2-Methoxy ethanol. L-dopa in 96% ethanol fraction of M. pruriens seed was 14.7%.Methode: This was an experimental study using complete randomized design. Subjects were BALB/C mice (Mus musculus. Five groups served as control, 3 groups received subcutaneos injection of 2-ME as muchas 100 mg/kg.bw/day for 12 days, followed with 96% ethanol fraction Mucuna pruriens seed starting from14 mg/kg.bw/day, 28 mg/kg.bw/day, and 56 mg/kg.bw/day for 51 days.Result: The 96% ethanol fraction of Mucuna pruriens seeds are significant increase motility (p<0,01 andthe percentage of normal spermatozoa morphology (p= 0,042.Conclusion: 96% ethanol fraction of Mucuna pruriens seeds are able to increase motility and the percentage of normal spermatozoa morphology in mice exposed to 2-ME. Keywords: Mucuna pruriens L., L-dopa, mouse spermatozoa

  12. Relationship between bovine fertility and the number of spermatozoa penetrating the cervical mucus within straws.

    Science.gov (United States)

    Taş, Muzaffer; Bacinoglu, Suleyman; Cirit, Umüt; Ozdaş, Ozen Banu; Ak, Kemal

    2007-09-01

    In this study, by using a recently developed test technique, the relationship between the total spermatozoa number penetrating determined sites of bovine cervical mucus in straws and potential fertility of bulls, and other spermatological characteristics were investigated. Furthermore, we aimed to determine the effect on the test results, of two different incubation temperatures (37 and 41 degrees C) and two sperm penetration distance ranges (PDRs). Frozen semen samples of six Holstein bulls were used in the study. The bulls were divided into two fertility groups (high and low fertility) according to the "non-return rates" (NRR). For the penetration test, cervical mucus was drawn into transparent plastic straws and incubated with semen at 37 and 41 degrees C for 15 min. After the incubation, straws were frozen in liquid nitrogen vapour and stored at -20 degrees C. On the evaluation day, concentrations of spermatozoa penetrated to the PDRs, each of which was 2.5 mm, between 32.5 and 35 mm (first penetration distance range, PDR1), and 50 and 52.5 mm (second penetration distance range, PDR2) distance in the straws from the open end, were measured. When compared with the low fertility group, bulls from the high fertility group showed a higher number of spermatozoa at the determined PDRs, and a significant positive correlation was found between the total number of spermatozoa at the penetration distances and the NRR scores of the bulls.

  13. Comparison of glycerol, lactamide, acetamide and dimethylsulfoxide as cryoprotectants of Japanese white rabbit spermatozoa.

    Science.gov (United States)

    Kashiwazaki, Naomi; Okuda, Yasushi; Seita, Yasunari; Hisamatsu, Shin; Sonoki, Shigenori; Shino, Masao; Masaoka, Toshio; Inomata, Tomo

    2006-08-01

    The rabbit is considered to be a valuable laboratory animal. We compared glycerol, lactamide, acetamide, and dimethylsulfoxide (DMSO) as cryoprotectants in egg-yolk diluent of ejaculated Japanese white rabbit spermatozoa for improvement of sperm cryopreservation methods. Rabbit semen was frozen with 1.0 M glycerol, lactamide, acetamide, or DMSO in plastic straws. Forward progressive motility and plasma membrane integrity of the post-thaw spermatozoa were examined. The rate of forward progressive motile spermatozoa in lactamide (37.8 +/- 3.0%) was significantly (P<0.05) higher than in glycerol (17.0 +/- 3.3%). In addition, the rates of sperm plasma membrane integrity in lactamide and acetamide (35.9 +/- 3.3% and 30.2 +/- 3.0%, respectively) were significantly (P<0.05) higher than in glycerol (17.0 +/- 2.6%). The results indicate that 1.0 M lactamide and acetamide have higher cryoprotective effects than 1.0 M glycerol for cryopreservation of Japanese white rabbit spermatozoa.

  14. Complete staining of human spermatozoa and immature germ cells combined with phase contrast microscopy

    DEFF Research Database (Denmark)

    Michael, A Y; Drejer, J O; Bagger, P V

    1987-01-01

    A method combining Janus green B and Thymol blue stains the anterior part of the head, the nuclear membrane, middle piece, and tail of spermatozoa light green and the nucleus deep purple. The method provides excellent stained preparations for the evaluation of sperm morphology by phase contrast...

  15. Epigenetic analysis of human spermatozoa after their injection into ovulated mouse oocytes

    Czech Academy of Sciences Publication Activity Database

    Fulka, Helena; Barnetová, I.; Moško, T.; Fulka, J.

    2008-01-01

    Roč. 23, č. 3 (2008), s. 627-634 ISSN 1355-4786 R&D Projects: GA ČR(CZ) GD204/05/H023 Institutional research plan: CEZ:AV0Z50520514 Keywords : spermatozoa * epigenetics * chromatin * acetylation * methylation Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 7.590, year: 2008

  16. Modelling of energy expended by free swimming spermatozoa in temperature-dependent viscous semen.

    Science.gov (United States)

    Foo, Jong Yong Abdiel

    2010-01-01

    Derived models of fertilization kinetics have relied upon estimates of the swimming velocity of spermatozoa from the insemination site to a fallopian tube. However, limited derivations are available describing the probability and energy expended when spermatozoa collide with one another. An analytic approach of spermatozoon motion in a linear viscoelastic fluid is adopted to simplify the derivation. The complex kinematics of motion of an inextensible flagellum is modelled as planar flagellar wave of small amplitude. In humans, a temperature difference is expected between the cooler tubal isthmus and the warmer tubal ampulla. Thus, fluidic characteristics of semen such as viscosity can vary along the female reproductive tract. The results suggest that the probability of spermatozoa colliding in relatively lower viscous semen increases by 64.87% for a 0.5 degrees C surge in temperature. Moreover, this increases for a denser concentration of spermatozoa due to the limited semen volume available to manoeuvre. In addition, the propulsive forces and shear stress were 39.35% lower in less viscous semen due to an increase in temperature of only 0.5 degrees C. Hence, the described derivations herein can assist in the understanding of work done by a normal motile spermatozoon in a pool of semen.

  17. RELATIONSHIP BETWEEN LEVEL OF COPPER IN BOVINE SEMINAL PLASMA AND SPERMATOZOA MOTILITY

    Directory of Open Access Journals (Sweden)

    Zuzana Kňažická

    2013-02-01

    Full Text Available The aim of this study was to evaluate relationship between copper (Cu concentration of bovine seminal plasma and spermatozoa motility. Semen samples were collected from 13 breeding bulls. The motility analysis was carried out using the Computer Assisted Sperm Analysis (CASA system. The mean value for the percentage of motile spermatozoa (MOT was 92.46±3.99% and the progressive motility of the spermatozoa (PROG as 90.23±4.02%. The seminal plasma Cu concentrations were analyzed by UV/VIS spectrophotometry. The total Cu concentration of the seminal plasma was 4.28±1.47 μM/L. The correlation analysis revealed a strong negative correlation between MOT and seminal plasma Cu concentration (rp=-0.781; P<0.01 as well as between PROG and Cu content in the seminal plasma (rp=-0.726; P<0.01. The data obtained from this study clearly indicated that concentration of copper in seminal plasma negatively affects the spermatozoa motility parameters and subsequently might cause reproductive alteration in male sexual functions.

  18. Trehalose in glycerol-free freezing extender enhances post-thaw survival of boar spermatozoa

    Science.gov (United States)

    ATHURUPANA, Rukmali; TAKAHASHI, Daisen; IOKI, Sumire; FUNAHASHI, Hiroaki

    2015-01-01

    Cryopreservation of boar semen is still considered suboptimal due to lower fertility as compared with fresh samples when glycerol, a permeating cryoprotectant, is used. Trehalose is a non-permeable cryoprotectant and nonreducing disaccharide known to stabilize proteins and biologic membranes. The aim of this study was to evaluate the cryosurvival and in vitro penetrability of boar spermatozoa when glycerol was replaced with trehalose in a freezing extender. Ejaculated Berkshire semen samples were diluted in egg yolk-based freezing extender containing glycerol (100 mM) or trehalose (0, 50, 100, 150, 200 and 250 mM) and cryopreserved using a straw freezing procedure. Thawed samples were analyzed for motility, viability, mitochondrial membrane potential (MMP), and acrosome integrity. In experiment 2, penetrability of spermatozoa cryopreserved with 100 mM glycerol or trehalose was examined. Replacement of cryoprotectant glycerol (100 mM) with trehalose had no effect on sperm viability, but replacing it with 100 mM trehalose improved motility, MMP and acrosome integrity significantly. Sperm motility and MMP were considerably higher in 100 mM trehalose, whereas the acrosome integrity was substantially higher in 100–250 mM trehalose. The in vitro penetration rate was also significantly higher in spermatozoa cryopreserved with trehalose (61.3%) than in those cryopreserved with glycerol (43.6%). In conclusion, 100 mM non-permeable trehalose can be used to replace glycerol, a permeating cryoprotectant, for maintenance of better post-thaw quality of boar spermatozoa. PMID:25754239

  19. THE PROPORTION OF X AND Y SPERM, VIABILITY AND MOTILITY OF RAM SPERMATOZOA AFTER SEPARATED WITH WHITE EGG ALBUMIN

    Directory of Open Access Journals (Sweden)

    Moh Takdir

    2017-02-01

    The aim of this research was to determine the proportion, viability and motility of X and Y ram spermatozoa separated with egg white albumin. Sperm samples derived from Garut ram, which was collected by using an artificial vagina. Observations were made on spermatozoa fraction above and below each medium fraction treatment. There are treatment egg white albumin as separation medium, each medium consisting of fractions top and bottom fraction with different concentration: 1 P0 = sperma before separation (control; 2 P1 = 10% above fraction + 30% lower fraction; P2 = 25% + 45%; P3 = 25% + 75%. Data proportion of X and Y, viability and motility were analyzed statistically by Completely Randomized Design patern in the direction followed by Duncan’s Multiple Range Test for data with a real difference. Separation with egg white albumin affect significantly increased the proportion of spermatozoa X and Y (P≤0.05, but tends to decrease the viability and motility of spermatozoa.The proportion of spermatozoa X and Y was highest in treatment P3,76.76% of spermatozoa X (fraction above 25% and 79.81% spermatozoa Y (75% lower fraction, with an average viability obtained respectively 68,9% (fraction above and 59,7% (bottom fraction, motility 77,5% (fraction above dan 84,0% (bottom fraction. It was concluded that the egg white albumin is very effective in changing the proportions of X and Y ram sperm with the quality of spermatozoa after separation feasible for applications insemination or processed into frozen semen.   (Keywords: Garut ram, White egg albumin, Spermatozoa X and Y

  20. Proteomic Markers of Functional Sperm Population in Bovines: Comparison of Low- and High-Density Spermatozoa Following Cryopreservation.

    Science.gov (United States)

    D'Amours, Olivier; Frenette, Gilles; Bourassa, Sylvie; Calvo, Ézéchiel; Blondin, Patrick; Sullivan, Robert

    2018-01-05

    Mammalian semen contains a heterogeneous population of sperm cells. This heterogeneity results from variability in the complex processes of cell differentiation in the testis, biochemical modifications undergone by spermatozoa during transit along the male reproductive tract, interactions with secretions from accessory sex glands at ejaculation, and, in the context of reproductive technologies, in the ability of ejaculated spermatozoa to resist damage associated with freeze-thaw procedures. When submitted to density gradient centrifugation, ejaculated spermatozoa distribute themselves into two distinct populations: a low-density population characterized by low motility parameters, and a high-density population with high motility characteristics. To understand the origin of ejaculated spermatozoa heterogeneity, cryopreserved semen samples from bulls used by the artificial insemination (A.I.) industry were submitted to Percoll gradient centrifugation. Proteins from low and high density spermatozoa were then extracted with sodium deoxycholate and submitted to proteomic analysis using iTRAQ (isobaric tag for relative and absolute quantitation) methodologies. Quantification of selected sperm proteins was confirmed by multiple reaction monitoring (MRM). Overall, 31 different proteins were more abundant in low-density spermatozoa, while 80 different proteins were more abundant in the high-density subpopulation. Proteins enriched in high-density spermatozoa were markers of sperm functionality such as the glycolytic process, binding to the egg zona pellucida, and motility. Low-density spermatozoa were not solely characterized by loss of proteins and their associated functions. Chaperonin-containing TCP1s and chaperones are hallmarks of the low-density subpopulation. iTRAQ analysis revealed that other proteins such as binder of sperm proteins, histone, GPX5, ELSPBP1, and clusterin are overexpressed in low-density spermatozoa suggesting that these proteins represent defects

  1. Pharmacological inhibition of arachidonate 15-lipoxygenase (ALOX15) protects human spermatozoa against oxidative stress.

    Science.gov (United States)

    Walters, Jessica L H; De Iuliis, Geoffry N; Dun, Matthew D; Aitken, Robert John; McLaughlin, Eileen A; Nixon, Brett; Bromfield, Elizabeth G

    2018-03-13

    One of the leading causes of male infertility is defective sperm function, a pathology that commonly arises from oxidative stress in the germline. Lipid peroxidation events in the sperm plasma membrane result in the generation of cytotoxic aldehydes such as 4-hydroxynonenal (4HNE), which accentuate the production of reactive oxygen species (ROS) and cause cellular damage. One of the key enzymes involved in the metabolism of polyunsaturated fatty acids to 4HNE in somatic cells is arachidonate 15-lipoxygenase (ALOX15). Although ALOX15 has yet to be characterized in human spermatozoa, our previous studies have revealed a strong link between ALOX15 activity and the levels of oxidative stress and 4HNE in mouse germ cell models. In view of these data, we sought to assess the function of ALOX15 in mature human spermatozoa and determine whether the pharmacological inhibition of this enzyme could influence the level of oxidative stress experienced by these cells. By driving oxidative stress in vitro with exogenous H2O2, our data reveal that 6,11-dihydro[1]benzothiopyrano[4,3-b]indole (PD146176; a selective ALOX15 inhibitor), was able to significantly reduce several deleterious, oxidative insults in spermatozoa. Indeed, PD146176 attenuated the production of ROS, as well as membrane lipid peroxidation and 4HNE production in human spermatozoa. Accordingly, ALOX15 inhibition also protected the functional competence of these cells to acrosome react and bind homologous human zonae pellucidae. Together, these results implicate ALOX15 in the propagation of an oxidative stress cascade within human spermatozoa and offer insight into potential therapeutic avenues to address male fertility that arises from oxidative stress.

  2. Estimate of oxygen consumption and intracellular zinc concentration of human spermatozoa in relation to motility.

    Science.gov (United States)

    Henkel, Ralf R; Defosse, Kerstin; Koyro, Hans-Wilhelm; Weissmann, Norbert; Schill, Wolf-Bernhard

    2003-03-01

    To investigate the human sperm oxygen/energy consumption and zinc content in relation to motility. In washed spermatozoa from 67 ejaculates, the oxygen consumption was determined. Following calculation of the total oxygen consumed by the Ideal Gas Law, the energy consumption of spermatozoa was calculated. In addition, the zinc content of the sperm was determined using an atomic absorption spectrometer. The resulting data were correlated to the vitality and motility. The oxygen consumption averaged 0.24 micromol/10(6) sperm x 24h, 0.28 micromol/10(6) live sperm x 24h and 0.85 micromol/10(6) live motile sperm x 24h. Further calculations revealed that sperm motility was the most energy consuming process (164.31 mJ/10(6) motile spermatozoa x 24h), while the oxygen consumption of the total spermatozoa was 46.06 mJ/10(6) spermatozoa x 24h. The correlation of the oxygen/energy consumption and zinc content with motility showed significant negative correlations (r= -0.759; P<0.0001 and r=-0.441; P<0.0001, respectively). However, when correlating sperm energy consumption with the zinc content, a significant positive relation (r=0.323; P=0.01) was observed. Poorly motile sperm are actually wasting the available energy. Moreover, our data clearly support the "Geometric Clutch Model" of the axoneme function and demonstrate the importance of the outer dense fibers for the generation of sperm motility, especially progressive motility.

  3. Association between exposure to persistent organohalogen pollutants and epididymal and accessory sex gland function: Multicentre study in Inuit and European populations

    DEFF Research Database (Denmark)

    Elzanaty, Saad; Rignell-Hydbom, Anna; Jönsson, Bo A.G.

    2006-01-01

    Exposure to persistent organochlorine pollutants (POPs) may have negative impact on male reproductive function. We, therefore, investigated the association between serum levels of POPs and epididymal and accessory sex gland function. Serum levels of CB-153, p,p′-DDE and seminal markers of epididy......Exposure to persistent organochlorine pollutants (POPs) may have negative impact on male reproductive function. We, therefore, investigated the association between serum levels of POPs and epididymal and accessory sex gland function. Serum levels of CB-153, p,p′-DDE and seminal markers...... with the activity of NAG were found among Greenlandic men (mean difference 7.0 mU/ejaculate, 95% CI 3.0, 34), and in the aggregated cohort (mean difference 4.0 mU/ejaculate, 95% CI -0.2, 8.0). A positive association was observed between CB-153 and PSA as well as zinc among Kharkiv men. In the Swedish cohort...

  4. Neonatal outcome and congenital malformations in children born after ICSI with testicular or epididymal sperm: a controlled national cohort study.

    Science.gov (United States)

    Fedder, J; Loft, A; Parner, E T; Rasmussen, S; Pinborg, A

    2013-01-01

    Does neonatal outcome including congenital malformations in children born after ICSI with epididymal and testicular sperm [testicular sperm extraction (TESE)/percutaneous epididymal sperm aspiration (PESA)/testicular sperm aspiration (TESA) (TPT)] differ from neonatal outcome in children born after ICSI with ejaculated sperm, IVF and natural conception (NC)? Children born after TPT have similar neonatal outcome, including total malformation rates, as have children born after ICSI and IVF with ejaculated sperm. Testing for variance over the four groups may indicate smaller differences in specific malformation rates with TPT as the highest risk group. Regarding neonatal outcome as well as congenital malformations in children born after TPT, studies are few, with limited sample size, heterogeneous and often performed without relevant control groups. Population-based cohort study including all Danish children born after TPT and fresh embryo transfer in Denmark from 1995 to 2009. Children born after transfer of frozen-thawed embryos were excluded. Control groups of children conceived by ICSI with ejaculated sperm, IVF and NC were identified by cross-linkage of the Danish IVF Register, Medical Birth Register (MBR) and National Hospital Discharge Register (HDR). The study group consisted of 466 children born after TPT, while the control groups consisted of 8967 (ICSI with ejaculated sperm), 17 592 (IVF) and 63 854 (NC) children. Neonatal outcomes and congenital malformations were analysed for singletons and twins separately. Risk estimates for low birthweight (LBW, congenital malformations in the TPT group was 7.7% and did not differ significantly from any of the control groups. However, singleton TPT boys showed an increased rate of cardiac malformations (3.6%) compared with singleton boys after IVF (1.4%; P = 0.04) and NC (1.1%; P = 0.02). Considering the level of male infertility as a continuum over the four groups, tests for variance in the rate of cardiac

  5. Maternal exposure to a mixture of persistent organic pollutants (POPs) affects testis histology, epididymal sperm count and induces sperm DNA fragmentation in mice.

    Science.gov (United States)

    Khezri, Abdolrahman; Lindeman, Birgitte; Krogenæs, Anette K; Berntsen, Hanne F; Zimmer, Karin E; Ropstad, Erik

    2017-08-15

    Persistent organic pollutants (POPs) are widespread throughout the environment and some are suspected to induce reproductive toxicity. As animals and humans are exposed to complex mixtures of POPs, it is reasonable to assess how such mixtures could interact with the reproductive system. Our aim is to investigate how maternal exposure to a mixture of 29 different persistent organic pollutants, formulated to mimic the relative POP levels in the food basket of the Scandinavian population, could alter reproductive endpoints. Female mice were exposed via feed from weaning, during pregnancy and lactation in 3 exposure groups (control (C), low (L) and high (H)). Testicular morphometric endpoints, epididymal sperm concentration and sperm DNA integrity were assessed in adult male offspring. We found that the number of tubules, proportion of tubule compartments and epididymal sperm concentration significantly decreased in both POP exposed groups. Epididymal sperm from both POP exposed groups showed increased DNA fragmentation. It is concluded that maternal exposure to a defined POP mixture relevant to human exposure can affect testicular development, sperm production and sperm chromatin integrity. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Comparative studies on testicular and epididymal morphology, and serum hormone concentrations in foxes and the hybrids during the breeding season.

    Science.gov (United States)

    Yang, T A; Yang, Y H; Peng, Y H; Cong, B; Diao, Y F; Bao, K; Hu, P F; Song, X C; Liu, L L; Yang, Y F; Xing, X M; Yang, F H

    2016-05-01

    The silver fox and the blue fox belong to different genera, and the hybrid males are fully or partially sterile. In the present study, the objective was to evaluate the causes of hybrid male sterility, and therefore analyze the differences in testicular, and epididymal morphology and serum hormone concentrations among silver foxes, blue foxes, and the hybrids during the breeding season. Samples were collected from 20 male silver foxes, 20 male blue foxes, 15 male HSBs (silver fox female × blue fox male hybrids) and 14 male HBSs (blue fox male × silver fox female hybrids), respectively. Seminal evaluation showed large numbers of sperm present in the semen of blue foxes and silver foxes, but no sperm present in the hybrids. Mean testicular volume and the diameter of seminiferous tubules in silver foxes and blue foxes were greater than in the hybrids; and there were many Sertoli cells, spermatogenic cells, and sperm in silver foxes and blue foxes, while spermatogenic cells decreased with no sperm in the hybrids. Mean serum LH and prolactin concentrations in silver foxes and blue foxes were less and testosterone was greater than in the hybrids (P<0.05). The results indicate that germ cell meioses in the hybrids were arrested at the prophase stage of meiosis, and that lesser concentrations of testosterone and greater concentrations of LH and prolactin can inhibit the completion of spermatogenesis. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Healthy offspring from freeze-dried mouse spermatozoa held on the International Space Station for 9 months.

    Science.gov (United States)

    Wakayama, Sayaka; Kamada, Yuko; Yamanaka, Kaori; Kohda, Takashi; Suzuki, Hiromi; Shimazu, Toru; Tada, Motoki N; Osada, Ikuko; Nagamatsu, Aiko; Kamimura, Satoshi; Nagatomo, Hiroaki; Mizutani, Eiji; Ishino, Fumitoshi; Yano, Sachiko; Wakayama, Teruhiko

    2017-06-06

    If humans ever start to live permanently in space, assisted reproductive technology using preserved spermatozoa will be important for producing offspring; however, radiation on the International Space Station (ISS) is more than 100 times stronger than that on Earth, and irradiation causes DNA damage in cells and gametes. Here we examined the effect of space radiation on freeze-dried mouse spermatozoa held on the ISS for 9 mo at -95 °C, with launch and recovery at room temperature. DNA damage to the spermatozoa and male pronuclei was slightly increased, but the fertilization and birth rates were similar to those of controls. Next-generation sequencing showed only minor genomic differences between offspring derived from space-preserved spermatozoa and controls, and all offspring grew to adulthood and had normal fertility. Thus, we demonstrate that although space radiation can damage sperm DNA, it does not affect the production of viable offspring after at least 9 mo of storage on the ISS.

  8. [Epididymis in an experimental model of DHT deficiency: immunolocalization of ERalpha and ERbeta in rat epididymal epithelial cells. In vivo and in vitro studies].

    Science.gov (United States)

    Kolasa, Agnieszka

    2006-01-01

    The aim of this study was to determine the effect of reduced availability of dihydrotestosterone (DHT) on the expression of estrogen receptors alpha and beta (ERalpha and ERbeta) in the epididymis in vivo and in vitro. Expression of estrogen receptors (ERs) is interesting because of the fact that the male reproductive system is controlled not only by androgens but also, in a far-reaching and complex manner, by estrogens. Control by estrogens is exercised through activation of ERs widely distributed in the epididymal epithelium. Epididymal epithelial cells contain a 5alpha-reductase (5alpha-red) which catalyzes the irreversible conversion of testosterone (T) into the most potent and chief androgen of the epididymis, dihydrotestosterone, known to maintain and regulate the structure and functions of the epididymis. Two isoforms of the 5alpha-red were identified: type 1 (5alpha-redl) and type 2 (5alpha-red2). 5alpha-reductase type 2 is more widely expressed in the epididymis than 5alpha-redl. DHT deficit was produced by inhibition of 5alpha-red2 using finasteride (Proscar, MSD Sweden), a steroid inhibitor of this enzyme. The study was performed in the adult, male Wistar rats randomly divided into control (K) and study (Fin56) groups (5 animals in each). Animals in the study group received 5mg finasteride/kg b.w., orally during 56 days (duration of one spermatogenesis). Immunoexpression of ERs was also studied in epididymal epithelial cells cultured with or without finasteride. It was shown that DHT deficiency, both in vivo and in vitro condition, modulated ERs expression in comparison to the epididymis from control rats and to epididymal cells cultured without finasteride. Distribution of ERalpha and ERbeta in epididymal cells changed (from nucleus to cytoplasm) and the level of ERs expression was markedly decreased. The present findings show that the DHT deficiency caused by finasteride altered the expression of ERalpha and ERbeta in the epididymis and possibly may

  9. Dioxin-induced changes in epididymal sperm count and spermatogenesis Mudanças induzidas por dioxina na contagem epididimal de esperma e espermatogênese

    Directory of Open Access Journals (Sweden)

    Warren G. Foster

    2011-06-01

    Full Text Available A single in utero exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD on gestation day 15 decreased epididymal sperm count in adult rats and thus was used to establish a tolerable daily intake for TCDD. However, several laboratories have been unable to replicate these findings. Moreover, conflicting reports of TCDD effects on daily sperm production suggest that spermatogenesis may not be as sensitive to the adverse effects of TCDD as previously thought. We performed a PubMed search using relevant search terms linking dioxin exposure with adverse effects on reproduction and spermatogenesis. Developmental exposure to TCDD is consistently linked with decreased cauda epididymal sperm counts in animal studies, although at higher dose levels than those used in some earlier studies. However, the evidence linking in utero TCDD exposure and spermatogenesis is not convincing. Animal studies provide clear evidence of an adverse effect of in utero TCDD exposure on epididymal sperm count but do not support the conclusion that spermatogenesis is adversely affected. The mechanisms underlying decreased epididymal sperm count are unknown; however, we postulate that epididymal function is the key target for the adverse effects of TCDD.Uma única exposição in utero a 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD no 15º dia de gestação diminuiu a contagem de esperma epididimal em ratos adultos e por isso foi utilizada para estabelecer uma dosagem diária tolerável para TCDD. No entanto, diversos laboratórios não conseguiram reproduzir esses resultados. Além disso, relatórios conflitantes dos efeitos de TCDD na produção diária de esperma sugere que espermatogênese pode não ser tão sensível aos efeitos adversos do TCDD como antes se pensava. Foi feita uma pesquisa no PubMed usando termos de pesquisa relevantes, relacionados à exposição à dioxina com efeitos adversos na reprodução e na espermatogênese. Exposição em desenvolvimento ao TCDD

  10. Efficacy of hyaluronic acid binding assay in selecting motile spermatozoa with normal morphology at high magnification

    Directory of Open Access Journals (Sweden)

    Mauri Ana L

    2010-12-01

    Full Text Available Abstract Background The present study aimed to evaluate the efficacy of the hyaluronic acid (HA binding assay in the selection of motile spermatozoa with normal morphology at high magnification (8400x. Methods A total of 16592 prepared spermatozoa were selected and classified into two groups: Group I, spermatozoa which presented their head attached to an HA substance (HA-bound sperm, and Group II, those spermatozoa that did not attach to the HA substance (HA-unbound sperm. HA-bound and HA-unbound spermatozoa were evaluated according to the following sperm forms: 1-Normal morphology: normal nucleus (smooth, symmetric and oval configuration, length: 4.75+/-2.8 μm and width: 3.28+/-0.20 μm, no extrusion or invagination and no vacuoles occupied more than 4% of the nuclear area as well as acrosome, post-acrosomal lamina, neck, tail, besides not presenting a cytoplasmic droplet or cytoplasm around the head; 2-Abnormalities of nuclear form (a-Large/small; b-Wide/narrow; c-Regional disorder; 3-Abnormalities of nuclear chromatin content (a-Vacuoles: occupy >4% to 50% of the nuclear area and b-Large vacuoles: occupy >50% of the nuclear area using a high magnification (8400x microscopy system. Results No significant differences were obtained with respect to sperm morphological forms and the groups HA-bound and HA-unbound. 1-Normal morphology: HA-bound 2.7% and HA-unbound 2.5% (P = 0.56. 2-Abnormalities of nuclear form: a-Large/small: HA-bound 1.6% vs. HA-unbound 1.6% (P = 0.63; b-Wide/narrow: HA-bound 3.1% vs. HA-unbound 2.7% (P = 0.13; c-Regional disorders: HA-bound 4.7% vs. HA-unbound 4.4% (P = 0.34. 3. Abnormalities of nuclear chromatin content: a-Vacuoles >4% to 50%: HA-bound 72.2% vs. HA-unbound 72.5% (P = 0.74; b-Large vacuoles: HA-bound 15.7% vs. HA-unbound 16.3% (P = 0.36. Conclusions The findings suggest that HA binding assay has limited efficacy in selecting motile spermatozoa with normal morphology at high magnification.

  11. AMP-activated kinase in human spermatozoa: identification, intracellular localization, and key function in the regulation of sperm motility

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    Violeta Calle-Guisado

    2017-01-01

    Full Text Available AMP-activated kinase (AMPK, a protein that regulates energy balance and metabolism, has recently been identified in boar spermatozoa where regulates key functional sperm processes essential for fertilization. This work′s aims are AMPK identification, intracellular localization, and their role in human spermatozoa function. Semen was obtained from healthy human donors. Sperm AMPK and phospho-Thr172-AMPK were analyzed by Western blotting and indirect immunofluorescence. High- and low-quality sperm populations were separated by a 40%-80% density gradient. Human spermatozoa motility was evaluated by an Integrated Semen Analysis System (ISAS in the presence or absence of the AMPK inhibitor compound C (CC. AMPK is localized along the human spermatozoa, at the entire acrosome, midpiece and tail with variable intensity, whereas its active form, phospho-Thr172-AMPK, shows a prominent staining at the acrosome and sperm tail with a weaker staining in the midpiece and the postacrosomal region. Interestingly, spermatozoa bearing an excess residual cytoplasm show strong AMPK staining in this subcellular compartment. Both AMPK and phospho-Thr172-AMPK human spermatozoa contents exhibit important individual variations. Moreover, active AMPK is predominant in the high motility sperm population, where shows a stronger intensity compared with the low motility sperm population. Inhibition of AMPK activity in human spermatozoa by CC treatment leads to a significant reduction in any sperm motility parameter analyzed: percent of motile sperm, sperm velocities, progressivity, and other motility coefficients. This work identifies and points out AMPK as a new molecular mechanism involved in human spermatozoa motility. Further AMPK implications in the clinical efficiency of assisted reproduction and in other reproductive areas need to be studied.

  12. AMP-activated kinase in human spermatozoa: identification, intracellular localization, and key function in the regulation of sperm motility

    Science.gov (United States)

    Calle-Guisado, Violeta; de Llera, Ana Hurtado; Martin-Hidalgo, David; Mijares, Jose; Gil, Maria C; Alvarez, Ignacio S; Bragado, Maria J; Garcia-Marin, Luis J

    2017-01-01

    AMP-activated kinase (AMPK), a protein that regulates energy balance and metabolism, has recently been identified in boar spermatozoa where regulates key functional sperm processes essential for fertilization. This work's aims are AMPK identification, intracellular localization, and their role in human spermatozoa function. Semen was obtained from healthy human donors. Sperm AMPK and phospho-Thr172-AMPK were analyzed by Western blotting and indirect immunofluorescence. High- and low-quality sperm populations were separated by a 40%–80% density gradient. Human spermatozoa motility was evaluated by an Integrated Semen Analysis System (ISAS) in the presence or absence of the AMPK inhibitor compound C (CC). AMPK is localized along the human spermatozoa, at the entire acrosome, midpiece and tail with variable intensity, whereas its active form, phospho-Thr172-AMPK, shows a prominent staining at the acrosome and sperm tail with a weaker staining in the midpiece and the postacrosomal region. Interestingly, spermatozoa bearing an excess residual cytoplasm show strong AMPK staining in this subcellular compartment. Both AMPK and phospho-Thr172-AMPK human spermatozoa contents exhibit important individual variations. Moreover, active AMPK is predominant in the high motility sperm population, where shows a stronger intensity compared with the low motility sperm population. Inhibition of AMPK activity in human spermatozoa by CC treatment leads to a significant reduction in any sperm motility parameter analyzed: percent of motile sperm, sperm velocities, progressivity, and other motility coefficients. This work identifies and points out AMPK as a new molecular mechanism involved in human spermatozoa motility. Further AMPK implications in the clinical efficiency of assisted reproduction and in other reproductive areas need to be studied. PMID:27678462

  13. [Size of testes and epididymes in boys up to 17 years of life assessed by ultrasound method and method of external linear measurements].

    Science.gov (United States)

    Osemlak, Paweł

    2011-01-01

    1. Determination of the size of testes and epididymes on the right and left side, in healthy boys in various age groups with use of non-invasive ultrasound examination method and the method of external linear measurements. 2. Determination of age, when intensive growth of testicular and epididymal size starts. 3. Determination whether there are statistically significant differences between the size of the right and the left testis, as well as between the right and left epididymis. 4. Evaluation of the ultrasound method and method of external linear measurements in their use for scientific investigations. 309 boys, aged from 1 day to 17 years of life, treated in the Clinical Department of Paediatric Surgery and Traumatology of the Medical University in Lublin from 2009 to 2010 due to diseases needed to be treated surgically, but not the scrotum, were examined in this study. No pathologies influencing the development of genital organs were found in these boys. Dimension of the testes was studied with ultrasound method and with method of external linear measurements. Dimension of epididymes was only examined with ultrasound method. In every age group the author calculated mean arithmetical values for: testiscular length, thickness, width and volume, as well as epididymal depth and basis. With consideration of standard deviation (X+/-1 SD) it was possible to define the range of dimension of healthy testes and epididymes and their change with age. Final dimensions of the right and left testis as well as of the right and left epididymis were compared. Dimensions of the testis on the same side of body acquired with the ultrasound method and acquired with the method of external linear measurements were compared. Statistical work-up with Wilcoxon test for two dependent groups was implemented. Ultrasound evaluation pointed to intensive 2.5-times increase in testicular length and width, and 2-times increase in testicular thickness in boys aged 10 to 17 years. Mean volume of

  14. Improved exogenous DNA uptake in bovine spermatozoa and gene expression in embryos using membrane destabilizing agents in ICSI-SMGT.

    Science.gov (United States)

    Sánchez-Villalba, Esther; Arias, María Elena; Zambrano, Fabiola; Loren, Pía; Felmer, Ricardo

    2018-02-01

    Sperm-mediated gene transfer (SMGT) is a simple, fast, and economical biotechnological tool for producing transgenic animals. However, transgene expression with this technique in bovine embryos is still inefficient due to low uptake and binding of exogenous DNA in spermatozoa. The present study evaluated the effects of sperm membrane destabilization on the binding capacity, location and quantity of bound exogenous DNA in cryopreserved bovine spermatozoa using Triton X-100 (TX-100), lysolecithin (LL) and sodium hydroxide (NaOH). Effects of these treatments were also evaluated by intracytoplasmic sperm injection (ICSI)-SMGT. Results showed that all treatments bound exogenous DNA to spermatozoa including the control. Spermatozoa treated with different membrane destabilizing agents bound the exogenous DNA throughout the head and tail of spermatozoa, compared with the control, in which binding occurred mainly in the post-acrosomal region and tail. The amount of exogenous DNA bound to spermatozoa was much higher for the different sperm treatments than the control (P Exogenous gene expression in embryos was also improved by these treatments. These results demonstrated that sperm membrane destabilization could be a novel strategy in bovine SMGT protocols for the generation of transgenic embryos by ICSI.

  15. Effect of albumin and polyvinyl alcohol on the vitality, motility and acrosomal integrity of canine spermatozoa incubated in vitro.

    Science.gov (United States)

    Risopatrón, J; Catalán, S; Miska, W; Schill, W-B; Sánchez, R

    2002-12-01

    Sperm culture media used for in vitro fertilization (IVF) procedures are important factors concerning the viability, motility and acrosomal integrity of spermatozoa. The aim of this study was to investigate the effects of three different sperm diluting media, tissue culture medium (TCM-199), sperm culture medium (Sp-TALP) and human tubular fluid (HTF) supplemented with varying concentrations of bovine serum albumin (1, 4 and 6%) or polyvinyl alcohol (0.8%) on the acrosomal integrity, motility and viability of canine spermatozoa. Ejaculates collected from four dogs were diluted in all media and spermatozoa were separated from seminal plasma by the swim-up technique. Sperm progressive motility was assessed using a phase contrast microscope. Viability and acrosomal integrity were evaluated using a dual stain technique (Giemsa-Trypan blue). The results demonstrated that the number of live canine spermatozoa was similar in culture media supplemented or not supplemented with macromolecules. A minimal concentration of albumin (1%) in the three media showed similar effects on vitality, motility and acrosomal integrity, as had higher concentrations (4 and 6%). The percentage of acrosome-intact spermatozoa was markedly higher after HTF (94.1%) than after TCM-199 (70.1%) or Sp-TALP (71.0%) without supplementation. It is concluded that serum bovine albumin, irrespective of the concentration, preserved sperm viability and function, and HTF is the most suitable medium for preserving the acrosome in canine spermatozoa prepared for in vitro manipulation through short incubation.

  16. Improved recovery of post-thaw motility and vitality of human spermatozoa cryopreserved in the presence of dithiothreitol.

    Science.gov (United States)

    Rao, B; David, G

    1984-10-01

    Semen was collected in the laboratory from nine healthy donors. The concentrations and the percentages of live and motile spermatozoa in all semen samples were within the normal range. Each sample was diluted with citrate-egg yolk-glycerol medium with and without 5 mM dithiothreitol (DTT). Samples were frozen in liquid nitrogen vapor (-70 degrees C) for 7 min and subsequently stored in liquid nitrogen. The effect of DTT in cryopreservation of sperm was determined by comparing percentage of motile and live spermatozoa between controls and DTT-treated post-thaw samples. Percentage of motile spermatozoa was determined by two techniques, laser Doppler velocimetry (LDV) and light microscopy. The percentage of live spermatozoa was measured by microscopic evaluation after staining with eosin-nigrosin. It was shown that the addition of DTT to the freezing medium significantly improved the recovery of motile and live spermatozoa in the post-thaw samples. The mean motility recovery, as measured by LDV, was 44.9% in the controls as compared to 73.9% in the DTT-treated samples. Similarly the mean recovery of live spermatozoa in the controls and DTT-treated samples was 66.5 and 86.6%, respectively. Based on these results, a new hypothesis implicating lipid peroxidation in cryoinjury is proposed. It is also suggested that the use of DTT in the freezing medium may offer an advantage over the commonly used techniques of human sperm cryopreservation.

  17. Metabolic incorporation of unsaturated fatty acids into boar spermatozoa lipids and de novo formation of diacylglycerols

    DEFF Research Database (Denmark)

    Svetlichnyy, V.; Müller, P.; Günther-Pomorski, Thomas

    2014-01-01

    Lipids play an important role in the maturation, viability and function of sperm cells. In this study, we examined the neutral and polar lipid composition of boar spermatozoa by thin-layer chromatography/mass spectrometry. Main representatives of the neutral lipid classes were diacylglycerols...... containing saturated (myristoyl, palmitoyl and stearoyl) fatty acyl residues. Glycerophosphatidylcholine and glycerophosphatidylethanolamine with alk(en)yl ether residues in the sn-1 position and unsaturated long chained fatty acyl residues in sn-2 position were identified as the most prominent polar lipids....... The only glycoglycerolipid was sulfogalactosylglycerolipid carrying 16:0-alkyl- and 16:0-acyl chains. Using stable isotope-labelling, the metabolic incorporation of exogenously supplied fatty acids was analysed. Boar spermatozoa incorporated hexadecenoic (16:1), octadecenoic (18:1), octadecadienoic (18...

  18. PENGARUH EKSTRAK DAUN KEMANGI (Ocimum sanctum L. TERHADAP MOTILITAS DAN KONSENTRASI SPERMATOZOA MENCIT JANTAN (Mus musculus

    Directory of Open Access Journals (Sweden)

    Safwan

    2016-10-01

    Full Text Available We have conducted a research on the effect of the extract of leaves basil (Ocimum sanctum L on male mice to investigate the consentration and motility spermatozoa mice. Twoty male mice which has age about 2 months with wight 20 – 25 g were divided into four groups. The extract was provided orally daily until the 20 th day. At 21th day were sacrificed and determine of consentration and motility. The result of the research showed that the extract of Ocimum sanctum L leaves that being given during 20 days with the dosage of 50 mg/g body weight, 100 mg/g bw, and 250 mg/g bw could signifi increase the consentration and motility spermatozoa mice.

  19. Organization of the cysts in bee (Hymenoptera, Apidae testis: number of spermatozoa per cyst

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    Cruz-Landim Carminda da

    2001-01-01

    Full Text Available The morphology of the cyst cells in Apis mellifera Linné, 1758, Scaptotrigona postica Latreille, 1804, and Melipona bicolor bicolor Lepeletier, 1836 testis, as well as the average number of spermatic cells are reported. The data indicates a supporting and nourrishing role of the cyst cells to the developing cystocytes. The counts of immature spermatozoa in the cysts show an average of 202.8 ± 21.2 spermatozoa for A. mellifera, 117.4 ± 8.68 for S. postica and 88.8 ± 15.57 for M. bicolor, which predict the occurrence of 8 mitotic cycles in the cystocytes of A. mellifera and 7 in the meliponines, considering that only one spermatozoom originates of each final spermatogonium.

  20. The efficiency of conventional microscopic selection is comparable to the hyaluronic acid binding method in selecting spermatozoa for male infertility patients

    OpenAIRE

    Meng-Ting Huang; Robert Kuo-Kuang Lee; Chung-Hao Lu; Ying-Jie Chen; Sheng-Hsiang Li; Yuh-Ming Hwu

    2015-01-01

    Objective: To evaluate if hyaluronic acid (HA)-bound spermatozoa surpassed conventional microscopy-selected spermatozoa in the status of sperm DNA integrity by acridine orange (AO) fluorescence staining. Materials and methods: Spermatozoa obtained from couples with indication for the intracytoplasmic sperm injection (ICSI) procedure due to male infertility (n = 34) and control males with normal sperm parameters (n = 12) were analyzed using AO fluorescence staining after density-gradient ce...

  1. Dysregulation of long noncoding RNAs in mouse testes and spermatozoa after exposure to cadmium

    International Nuclear Information System (INIS)

    Gao, Fengxin; Zhang, Peng; Zhang, Hongyan; Zhang, Yunhui; Zhang, Yunwen; Hao, Qingyun; Zhang, Xiaoning

    2017-01-01

    There is increasing evidence that cadmium (Cd) exposure can cause male subfertility and even complete infertility in mammals. Long noncoding (lnc) RNAs are critical for spermatogenesis, and their dysregulation might lead to male infertility. However, whether they are involved in Cd-induced subfertility is unknown. Here we found that intraperitoneal exposure to Cd in mice led to male subfertility indicated by reductions in testicular sperm production and motility, and by abnormal morphology. Testicular and sperm RNAs were used to investigate lncRNA expression profiles by strand-specific RNA sequencing at the transcriptome level to help determine any RNA-related mechanisms in Cd-induced subfertility. The Cd-treated testes and spermatozoa exhibited aberrant expression profiles for lncRNAs and mRNAs. Of the lncRNAs, there were 139 with upregulated expression and 174 with downregulated expression in testes; in contrast, 685 were upregulated and 375 were downregulated in spermatozoa. For mRNA expression, 214 were upregulated and 226 were downregulated in testes; 272 were upregulated and 111 were downregulated in spermatozoa. Gene ontology and pathway analyses showed that the functions of differentially expressed lncRNA targets and mRNAs were closely linked with many processes involved in spermatogenesis. Additionally, many newly identified lncRNAs showed inducible expression, suggesting that they might be good candidate markers for Cd-induced male reproductive toxicity. This study provides a preliminary database for further exploring lncRNA-related mechnisms in male infertility induced by Cd. - Highlights: • LncRNA profiles were changed by Cd exposure in mouse testes and spermatozoa. • Differentially expressed lncRNA targets and mRNAs were linked with spermatogenesis. • Providing a database for exploring lncRNA-related mechnisms in male infertility. • LncRNA might be candidate markers for Cd-induced male reproductive toxicity.

  2. Sneaker Male Squid Produce Long-lived Spermatozoa by Modulating Their Energy Metabolism

    OpenAIRE

    Hirohashi, Noritaka; Tamura-Nakano, Miwa; Nakaya, Fumio; Iida, Tomohiro; Iwata, Yoko

    2016-01-01

    Spermatozoa released by males should remain viable until fertilization. Hence, sperm longevity is governed by intrinsic and environmental factors in accordance with the male mating strategy. However, whether intraspecific variation of insemination modes can impact sperm longevity remains to be elucidated. In the squid Heterololigo bleekeri, male dimorphism (consort and sneaker) is linked to two discontinuous insemination modes that differ in place and time. Notably, only sneaker male spermato...

  3. Water contaminated with Didymosphenia geminata generates changes in Salmo salar spermatozoa activation times.

    Science.gov (United States)

    Olivares, Pamela; Orellana, Paola; Guerra, Guillermo; Peredo-Parada, Matías; Chavez, Viviana; Ramirez, Alfredo; Parodi, Jorge

    2015-06-01

    Didimosphenia geminata ("didymo"), has become a powerful and devastating river plague in Chile. A system was developed in D. geminata channels with the purpose evaluating the effects of water polluted with didymo on the activation of Atlantic salmon (Salmo salar) spermatozoa. Results indicate that semen, when activated with uncontaminated river water had an average time of 60±21s. When using Powermilt, (a commercial activator), times of 240±21s are achieved, while rivers contaminated with D. geminata achieve a motility time of 30±12s. Interestingly enough, the kinetic parameters of VSL, VCL and VAP showed no significant changes under all of the conditions. Furthermore, the presence of D. geminata reduces activation time of the samples as the cells age, indicating increased effects in spermatozoa that are conserved for more than 5 days. D. geminata has antioxidant content, represented by polyphenols; 200ppm of polyphenol were obtained in this study per 10g of microalgae. Spermatozoa exposed to these extracts showed a reduction in mobility time in a dose dependent manner, showing an IC50 of 15ppm. The results suggest an effect on spermatozoa activation, possibly due to the release of polyphenols present in contaminated rivers, facilitating the alteration of sperm motility times, without affecting the viability or kinetics of the cells. These findings have important implications for current policy regarding the control of the algae. Current control measures focus on the number of visible species, and not on the compounds that they release, which this study shows, also have a problematic effect on salmon production. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. GABAA [gamma-aminobutyric acid] type binding sites on membranes of spermatozoa

    International Nuclear Information System (INIS)

    Erdoe, S.L.; Wekerle, L.

    1990-01-01

    The binding of [ 3 H] gamma-aminobutyric acid (GABA) to seminal membranes of swines and rams was examined. Specific, GABA binding was demonstrated in both species, which showed the features of GABA A type receptors. The affinity of binding was similar in both species, whereas the density of seminal GABA binding sites was 5 times higher in swine. Our findings suggest that GABA may have a direct effect on spermatozoa

  5. Methyl-parathion decreases sperm function and fertilization capacity after targeting spermatocytes and maturing spermatozoa

    Energy Technology Data Exchange (ETDEWEB)

    Pina-Guzman, Belem; Sanchez-Gutierrez, M.; Marchetti, Francesco; Hernandez-Ochoa, I.; Solis-Heredia, M.J .; Quintanilla-Vega, B.

    2009-05-03

    Paternal germline exposure to organophosphorous pesticides (OP) has been associated with reproductive failures and adverse effects in the offspring. Methyl parathion (Me-Pa), a worldwide-used OP, has reproductive adverse effects and is genotoxic to sperm. Oxidative damage has been involved in the genotoxic and reproductive effects of OP. The purpose of this study was to determine the effects of Me-Pa on spermatozoa function and ability to fertilize. Male mice were exposed to Me-Pa (20 mg/kg bw, i.p.) and spermatozoa from epididymis-vas deferens were collected at 7 or 28 days post-treatment (dpt) to assess the effects on maturing spermatozoa and spermatocytes, respectively. DNA damage was evaluated by nick translation (NT-positive cells) and SCSA (percentDFI); lipoperoxidation (LPO) by malondialdehyde production; sperm function by spontaneous- and induced-acrosome reactions (AR); mitochondrial membrane potential (MMP) by using the JC-1 flurochrome; and, fertilization ability by an in vitro assay and in vivo mating. Results showed alterations in DNA integrity (percentDFI and NT-positive cells) at 7 and 28 dpt, in addition to decreased sperm quality and a decrease in induced-AR; reduced MMP and LPO was observed only at 7 dpt. We found negative correlations between LPO and all sperm alterations. Altered sperm functional parameters were associated with reduced fertilization rates at both times, evaluated either in vitro or in vivo. These results show that Me-Pa exposure of maturing spermatozoa and spermatocytes affects many sperm functional parameters that result in a decreased fertilizing capacity. Oxidative stress seems to be a likely mechanism ofthe detrimental effects of Me-Pa in male germ cells.

  6. Effect of heterologous seminal plasma and semen extenders on motility of frozen-thawed ram spermatozoa

    Directory of Open Access Journals (Sweden)

    G.A. Mataveia

    2010-05-01

    Full Text Available Ram seminal plasma increases the fertility of frozen-thawed ram spermatozoa deposited into the cervix. The aim of the current study was to compare the effect of ram seminal plasma to that of bull seminal plasma, dog prostatic fluid, protein-free TALP, TrilEq (Triladyl with 0.5 mℓ of Equex STM paste added to each 100 mℓ and heat-treated skim milk on longevity and percentages of progressively motile and aberrantly motile frozen-thawed ram spermatozoa. Three ejaculates from each of 6 rams were extended in TrilEq, pooled and frozen in straws as a single batch per ram. One hundred and eight straws (3 straws from each ram for each fluid were thawed in random order. Once thawed, a straw was emptied into a tube with 0.85mℓ of the appropriate fluid at 37 °C and kept at that temperature for 6 h. Motility was assessed at x200 magnification immediately (time zero and 2, 4 and 6 h after thawing. Progressive motility decreased from each time to the next (P < 0.05 and was 39.0% (0 h, 26.0% (2 h, 19.6% (4 h and 12.6% (6 h; SEM 1.24, n=108 for each group. Ram seminal plasma resulted in higher progressive motility than bull seminal plasma, lower than milk, and similar to the other fluids. Ram seminal plasma resulted in lower aberrant motility than protein-free TALP and similar aberrant motility to other fluids. The effect of ram seminal plasma and dog prostatic fluid was very similar. The effect of ram seminal plasma on the fertility of frozen-thawed ram spermatozoa deposited into the cervix is not due an exceptionally beneficial effect on the motility of spermatozoa.

  7. Feasibility of refreezing human spermatozoa through the technique of liquid nitrogen vapor

    Directory of Open Access Journals (Sweden)

    Sidney Verza Jr

    2004-12-01

    Full Text Available OBJECTIVE: To assess the feasibility of refreezing human semen using the technique of liquid nitrogen vapor with static phases. MATERIALS AND METHODS: Twenty samples from 16 subjects who required disposal of their cryopreserved semen were thawed, corresponding to 6 cancer patients and 10 participants in the assisted reproduction (AR program. Samples were refrozen using the technique of liquid nitrogen vapor with static phases, identical to the one used for the initial freezing, and thawed again after 72 hours. We assessed the concentration of motile spermatozoa, total and progressive percent motility and spermatic vitality, according to criteria of the World Health Organization (WHO, as well as spermatic morphology according to the strict Kruger criterion, after the first and after the second thawing. RESULTS: We observed a significant decrease in all the parameters evaluated between the first and the second thawing. Median values for the concentration of motile spermatozoa decreased from 2.0x10(6/mL to 0.1x10(6/mL (p < 0.01; total percent motility from 42% to 22.5% (p < 0.01; progressive percent motility from 34% to 9.5% (p < 0.01; vitality from 45% to 20% (p < 0.01; and morphology from 5% to 5% (p = 0.03. There was no significant difference in the spermatic parameters between the cancer and assisted reproduction groups, both after the first and after the second thawing. We observed that in 100% of cases there was retrieval of motile spermatozoa after the second thawing. CONCLUSIONS: Refreezing of human semen by the technique of liquid nitrogen vapor allows the retrieval of viable spermatozoa after thawing.

  8. The effect of tetrabromobisphenol A on protamine content and DNA integrity in mouse spermatozoa

    Czech Academy of Sciences Publication Activity Database

    Žatecká, Eva; Castillo, J.; Elzeinová, Fatima; Kubátová, Alena; Děd, Lukáš; Pěknicová, Jana; Oliva, R.

    2014-01-01

    Roč. 2, č. 6 (2014), s. 910-917 ISSN 2047-2927 R&D Projects: GA ČR(CZ) GAP503/12/1834; GA MŠk(CZ) ED1.1.00/02.0109 Institutional research plan: CEZ:AV0Z50520701 Institutional support: RVO:86652036 Keywords : protamines * spermatozoa * tetrabromobisphenol A * TUNAL assay Subject RIV: EB - Genetics ; Molecular Biology

  9. Study of pentoxifylline effects on motility and viability of spermatozoa from infertile asthenozoospermic males

    OpenAIRE

    Ghasemzadeh, Aliye; Karkon-Shayan, Farid; Yousefzadeh, Solmaz; Naghavi-Behzad, Mohammad; Hamdi, Kobra

    2016-01-01

    Background: The quality of semen is one of the major parameters in male infertility. Pentoxifylline, a methylxanthine derivative, is an agent primarily used in the treatment of intermittent claudication and other vascular disorders. Studies have shown that pentoxifylline enhances the quality and quantity of sperms. In this study, we have investigated the in vitro effects of pentoxifylline on viability and motility of spermatozoa in samples of infertile oligoasthenozoospermic males. Materials ...

  10. the effect of freezing method on the survival of ram spermatozoa

    African Journals Online (AJOL)

    Sci.2l, 351. SALAIIiON, S., 1970. The survival of ram spermatozoa following pellet freezing below-79oC. Aust- .1. hiol..n'r. 13. 459. SALAMON, S. & BRANDON, M.R., l97l . Effect of compcition of thawing solution on sunrival of ram spernntozoa iroznn by the pellet method. Aust. J. biol. Sci. 24,355. SALAMON, S. & VISSER, D., ...

  11. Localization of porcine seminal plasma (PSP) proteins in the boar reproductive tract and spermatozoa

    Czech Academy of Sciences Publication Activity Database

    Maňásková, Pavla; Jonáková, Věra

    2008-01-01

    Roč. 78, č. 1 (2008), s. 40-48 ISSN 0165-0378 R&D Projects: GA ČR GA303/06/0895; GA MŠk 1M06011 Institutional research plan: CEZ:AV0Z50520514; CEZ:AV0Z50520701 Keywords : porcine seminal plasma proteins * boar reproductive tract * spermatozoa Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.778, year: 2008

  12. Differential expression of a novel seven transmembrane domain protein in epididymal fat from aged and diabetic mice.

    Science.gov (United States)

    Yang, H; Egan, J M; Rodgers, B D; Bernier, M; Montrose-Rafizadeh, C

    1999-06-01

    To identify novel seven transmembrane domain proteins from 3T3-L1 adipocytes, we used PCR to amplify 3T3-L1 adipocyte complementary DNA (cDNA) with primers homologous to the N- and C-termini of pancreatic glucagon-like peptide-1 (GLP-1) receptor. We screened a cDNA library prepared from fully differentiated 3T3-L1 adipocytes using a 500-bp cDNA PCR product probe. Herein describes the isolation and characterization of a 1.6-kb cDNA clone that encodes a novel 298-amino acid protein that we termed TPRA40 (transmembrane domain protein of 40 kDa regulated in adipocytes). TPRA40 has seven putative transmembrane domains and shows little homology with the known GLP-1 receptor or with other G protein-coupled receptors. The levels of TPRA40 mRNA and protein were higher in 3T3-L1 adipocytes than in 3T3-L1 fibroblasts. TPRA40 is present in a number of mouse and human tissues. Interestingly, TPRA40 mRNA levels were significantly increased by 2- to 3-fold in epididymal fat of 24-month-old mice vs. young controls as well as in db/db and ob/ob mice vs. nondiabetic control littermates. No difference in TPRA40 mRNA levels was observed in brain, heart, skeletal muscle, liver, or kidney. Furthermore, no difference in TPRA40 expression was detected in brown fat of ob/ob mice when compared with age-matched controls. Taken together, these data suggest that TPRA40 represents a novel membrane-associated protein whose expression in white adipose tissue is altered with aging and type 2 diabetes.

  13. Effectiveness of human spermatozoa biomarkers as indicators of structural damage during cryopreservation.

    Science.gov (United States)

    Gómez-Torres, María José; Medrano, Llanos; Romero, Alejandro; Fernández-Colom, Pedro José; Aizpurúa, Jon

    2017-10-01

    Human spermatozoa cryopreservation techniques are used to maintain and protect male fertility in cases such as infertility and malignancy treatments. However, during cryopreservation, the spermatozoa's metabolic rate is reduced and they undergo dramatic functional and structural changes owing to exposure to cryoprotectants and freezing-thawing procedures. While the effects of cryopreservation on cells are documented, to date the induced cryodamage on structural and/or functional sperm biomarkers is not well established at multivariate scale. To address this question, we performed basic sperm analysis, sperm DNA fragmentation assessment, spontaneous acrosome reaction measurement, and cytoskeleton evaluation after thawing samples from subjects with normal and low-quality semen. A cryodamage rate was used to determine the effects of the freeze-thaw process on spermatozoa. In addition, a Principal Component Analysis (PCA) was used for data reduction and to evaluate sperm-specific patterns during the cryopreservation process. We found that the vitality, progressive motility and sperm count from low-quality samples after cryopreservation show higher damage rates (≥40%) than in normal sperm samples. However, cytoskeleton, DNA, tail and mid-piece and acrosome display the highest cryodamage rates (∼50-99%) and are equally susceptible to cryopreservation-induced damage in both low- and normal-quality semen samples. Overall, the evaluation of these parameters provides meaningful information about different aspects of sperm functionality after cryopreservation. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Mitochondrial permeability transition pore (MPTP) desensitization increases sea urchin spermatozoa fertilization rate.

    Science.gov (United States)

    Torrezan-Nitao, Elis; Boni, Raianna; Marques-Santos, Luis Fernando

    2016-10-01

    Mitochondrial permeability transition pore (MPTP) is a protein complex whose opening promotes an abrupt increase in mitochondrial inner membrane permeability. Calcium signaling pathways are described in gametes and are involved in the fertilization process. Although mitochondria may act as Ca(2+) store and have a fast calcium-releasing mechanism through MPTP, its contribution to fertilization remains unclear. The work aimed to investigate the MPTP phenomenon in sea urchin spermatozoa and its role on the fertilization. Several pharmacological tools were used to evaluate the MPTP's physiology. Our results demonstrated that MPTP occurs in male gametes in a Ca(2+) - and voltage-dependent manner and it is sensitive to cyclosporine A. Additionally, our data show that MPTP opening does not alter ROS generation in sperm cells. Inhibition of MPTP in spermatozoa strongly improved the fertilization rate, which may involve mechanisms that increase the spermatozoa lifespan. The present work is the first report of the presence of a voltage- and Ca(2+) -dependent MPTP in gametes of invertebrates and indicates MPTP opening as another evolutionary feature shared by sea urchins and mammals. Studies about MPTP in sea urchin male gametes may contribute to the elucidation of several mechanisms involved in sperm infertility. © 2016 International Federation for Cell Biology.

  15. Tolerance of brown bear spermatozoa to conditions of pre-freezing cooling rate and equilibration time.

    Science.gov (United States)

    López-Urueña, E; Alvarez, M; Gomes-Alves, S; Martínez-Rodríguez, C; Borragan, S; Anel-López, L; de Paz, P; Anel, L

    2014-06-01

    Specific protocols for the cryopreservation of endangered Cantabrian brown bear spermatozoa are critical to create a genetic resource bank. The aim of this study was to assess the effect of cooling rates and equilibration time before freezing on post-thawed brown bear spermatozoa quality. Electroejaculates from 11 mature bears were extended to 100 × 10(6) spermatozoa/mL in a TES-Tris-Fructose-based extender, cryopreserved following performance of the respective cooling/equilibration protocol each sample was assigned to, and stored at -196 °C for further assessment. Before freezing, after thawing, and after 1 hour's incubation post-thawing at 37 °C (thermal stress test), the quality of the samples was assessed for motility by computer-assisted semen analysis, and for viability (SYBR-14/propidium iodide), acrosomal status (peanut agglutinin-fluorescein isothiocyanate /propidium iodide), and sperm chromatin stability (SCSA) by flow cytometry. In experiment 1, three cooling rates (0.25 °C/min, 1 °C/min, and 4 °C/min) to 5 °C were assessed. After thawing, total motility (%TM) was higher and percentage of damaged acrosomes (%dACR) was lower (P bear sperm. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. EFFECT OF STORAGE TEMPERATURE ON THE MOTILITY CHARACTERISTICS OF ROOSTER SPERMATOZOA

    Directory of Open Access Journals (Sweden)

    Jaromír Vašíček

    2013-02-01

    Full Text Available The objective of our study was to evaluate the influence of different storage temperature on rooster sperm motility. Semen was collected once a week from Lohmann Light breeder males into prepared sterile tube. The heterospermic pool was diluted at the ratio of 1:100 in a commercial avian extender, divided into two aliquots and incubated at 8°C or 37°C. The quality of semen samples were evaluated using CASA system (Sperm Vision™ after 30, 60 and 120 min of incubation. We observed significantly (P<0.05 the highest progressive motility of rooster sperm after 60 min of spermatozoa incubation at 8°C, that was also significantly (P<0.05 higher in comparison to spermatozoa incubated for 60 min at 37°C. Basing on the observed results, we hypothesized that law temperatures (about 8°C would be better for long-term storage of rooster spermatozoa. Further experiments with different semen diluents and storage temperatures and intervals are required in order to prove our hypothesis.

  17. Dietary omega-3 polyunsaturated fatty acids induce plasminogen activator activity and DNA damage in rabbit spermatozoa.

    Science.gov (United States)

    Kokoli, A N; Lavrentiadou, S N; Zervos, I A; Tsantarliotou, M P; Georgiadis, M P; Nikolaidis, E A; Botsoglou, N; Boscos, C M; Taitzoglou, I A

    2017-12-01

    The aim of this study was to determine the effect(s) of dietary omega-3 polyunsaturated fatty acids (ω-3 PUFA) on rabbit semen. Adult rabbit bucks were assigned to two groups that were given two diets, a standard diet (control) and a diet supplemented with ω-3 PUFA. Sperm samples were collected from all bucks with the use of an artificial vagina in 20-day intervals, for a total period of 120 days. The enrichment of membranes in ω-3 PUFA was manifested by the elevation of the 22:5 ω-3 (docosapentaenoic acid [DPA]) levels within 40 days. This increase in DPA content did not affect semen characteristics (i.e., concentration, motility and viability). However, it was associated with the induction of lipid peroxidation in spermatozoa, as determined on the basis of the malondialdehyde content. Lipid peroxidation was associated with DNA fragmentation in ω-3 PUFA-enriched spermatozoa and a concomitant increase in plasminogen activator (PA) activity. The effects of ω-3 PUFA on sperm cells were evident within 40 days of ω-3 PUFA dietary intake and exhibited peack values on day 120. Our findings suggest that an ω-3 PUFA-rich diet may not affect semen characteristics; however, it may have a negative impact on the oxidative status and DNA integrity of the spermatozoa, which was associated with an induction of PAs activity. © 2017 Blackwell Verlag GmbH.

  18. Effect of Freezing on Spermatozoa from Tigaie Rams Belonging to the Mountain Ecotype

    Directory of Open Access Journals (Sweden)

    Vasile Miclea

    2011-05-01

    Full Text Available Our aim was to study the influence of freezing on the viability and frequency of abnormalities in frozen ram spermatozoa. Sperm was collected form 20 rams belonging to the mountain ecotype of the Tigaie breed using the artificial vagina technique and volume and motility were assessed. Afterward it was diluted with Tryladil (1:4 supplemented with 20% egg yolk and heated at 37°C. Subsequently the temperature decreased at a rate of 0.2°C/minute until reaching 4°C and an equilibration time of 2 hours followed. During this time the diluted sperm was packaged in 0.25 ml straws. After sealing these were kept 6 cm above liquid nitrogen level for 13 minutes (- 120°C and then plunged into nitrogen. Volume, motility and concentration were assessed before freezing. After thawing sperm morphology was assessed using Hancock’s method and at the same time the endurance (at 10, 30 and 60 minutes and HOST tests were performed. The highest motility (0.40 was graded at 30 minutes. It could be correlated with the increased percentage of HOST positive spermatozoa, 27.78%. The percentage of abnormal spermatozoa was also high (47.89%, 38.44% of them having acrosome flaws. Cryopreservation has a negative effect on the characteristics of sperm cells from Tigaie rams belonging to the mountain ecotype.

  19. Infertile spermatozoa in a human carrier of robertsonian translocation 14;22.

    Science.gov (United States)

    Baccetti, Baccio; Capitani, Serena; Collodel, Giulia; Estenoz, Mariela; Gambera, Laura; Piomboni, Paola

    2002-11-01

    To present the ultrastructural, functional, and chromosomal analyses of spermatozoa from an infertile man with normal phenotype and chromosomal translocation 14;22. Case report. Regional Reference Center for Male Infertility in Siena, Italy. A 36-year-old man with primary infertility for 3 years and his parents. Family history and lymphocytic karyotypes, physical and hormonal assays, and semen analysis. Morphological sperm evaluation was performed by light, fluorescent, and electron microscopy; chromosomal constitution was examined by the fluorescence in situ hybridization (FISH) technique. The penetration ability of spermatozoa was checked by the hamster test. The spermatozoa of the patient showed unusual ultrastructural defects. The nuclei were large, spheroidal, and generally uncondensed; the acrosomes were frequently absent or reduced; and the axonemes were often devoid of dynein arms or central singlet tubules. These characteristics are related to immaturity. The lymphocytic karyotype revealed a robertsonian translocation 14;22 in the sterile patient and his mother. FISH sperm analysis demonstrated a high frequency of diploidy for the chromosome 18,XY. The hamster penetration test gave negative results. The unusual structural sperm immaturity is associated with the translocation 14;22. This chromosomal anomaly may therefore negatively influence the spermatogenesis; an interchromosomal effect on meiosis segregation is also suggested.

  20. Types of structural chromosome aberrations and their incidences in human spermatozoa X-irradiated in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Kamiguchi, Yujiroh; Tateno, Hiroyuki; Mikamo, Kazuya (Asahikawa Medical College (Japan). Department of Biological Sciences)

    1990-02-01

    The authors studied the effects of in vitro X-irradiation on human sperm chromosomes, using our interspecific in vitro fertilization system between human spermatozoa and zona-free hamster oocytes. 28 semen samples from 5 healthy men were exposed to 0.23, 0.45, 0.91 and 1.82 Gy of X-rays. Totals of 2098 and 2862 spermatozoa were karyotyped in the control and the irradiated groups, respectively. The indicence of spermatozoa with X-ray-induced structural chromosome aberrations (Y) increased linearly with increasing dosage (D), being best expressed by the equation, Y = 0.08 + 34.52 D. The incidence of breakage-type aberrations was moe than 9 times higher than that of exchange-type aberrations. Both of them showed linear dose-dependent increases, which were expressed by the regression lines, Y = -0.014 + 0.478 D and Y -0.010 + 0.057 D, respectively. The incidence of chromosome-ltype aberrations was about 6 times higher than that of chromatid-type aberrations. Their dose-dependent increases were expressed by the regression lines, Y = -0.015 + 0.462 D and Y = -0.006 + 0.079 D, respectively. These results are discussed in relation to the previous data obtained with {gamma}-rays. The repair mechanism of X-ray-induced sperm DNA lesions is also discussed. (author). 21 refs.; 4 figs.; 4 tabs.

  1. Levels of semenogelin in human spermatozoa decrease during capacitation: involvement of reactive oxygen species and zinc.

    Science.gov (United States)

    de Lamirande, E; Lamothe, G

    2010-07-01

    Semenogelin (Sg), the main protein of human semen coagulum, prevents sperm capacitation. The objective of this study was to examine the role of Sg and its mechanism of action. Sg blocked sperm capacitation triggered by various stimuli, via inhibition of superoxide anion (O(2)*-; luminescence assay) and nitric oxide (NO*; tested using diaminofluorescein) generation. Triton-soluble and -insoluble sperm fractions contained Sg and Sg peptides (immunoblotting), the level of which decreased with initiation of capacitation. This drop was prevented by superoxide dismutase and NO* synthase inhibitor and was reproduced by addition of O(2)*- and NO*. Zinc (Zn(2+)) blocked and a zinc chelator (TPEN) promoted the decline in Sg levels. There was a decreased labelling of Sg on the head in capacitating spermatozoa with the two fixation techniques tested (immunocytochemistry). Reactive oxygen species (ROS) (O(2)*- and NO*) caused, these changes, and zinc prevented them. Spermatozoa quickly internalized Sg upon incubation and Sg was then rapidly degraded in a zinc-inhibitable manner. Sg blocked capacitation mainly via inhibition of ROS generation. Spermatozoa appeared permeable to Sg and processed Sg in a zinc-inhibitable fashion. ROS themselves could promote sperm disposal of Sg which maybe one of the mechanisms that allows initiation of capacitation.

  2. Positioning of chromosomes in human spermatozoa is determined by ordered centromere arrangement.

    Directory of Open Access Journals (Sweden)

    Olga S Mudrak

    Full Text Available The intranuclear positioning of chromosomes (CHRs is a well-documented fact; however, mechanisms directing such ordering remain unclear. Unlike somatic cells, human spermatozoa contain distinct spatial markers and have asymmetric nuclei which make them a unique model for localizing CHR territories and matching peri-centromere domains. In this study, we established statistically preferential longitudinal and lateral positioning for eight CHRs. Both parameters demonstrated a correlation with the CHR gene densities but not with their sizes. Intranuclear non-random positioning of the CHRs was found to be driven by a specific linear order of centromeres physically interconnected in continuous arrays. In diploid spermatozoa, linear order of peri-centromeres was identical in two genome sets and essentially matched the arrangement established for haploid cells. We propose that the non-random longitudinal order of CHRs in human spermatozoa is generated during meiotic stages of spermatogenesis. The specific arrangement of sperm CHRs may serve as an epigenetic basis for differential transcription/replication and direct spatial CHR organization during early embryogenesis.

  3. The Antarctic holothurian genus Echinopsolus Gutt, 1990 (Dendrochirotida, Cucumariidae): brood pouches, spermatozoa, spermatozeugmata and taxonomic implications.

    Science.gov (United States)

    Bohn, Jens Michael; Heß, Martin

    2014-07-29

    An examination of seven Antarctic brooding cucumariid and psolid holothurian species revealed a variety of characters all of them have in common: (1) All are gonochoric. (2) A genital papilla is present on the oral disc (permanent and digitiform in males). (3) Females brood their offspring in five anterior interradial brood pouches that are situated at the transition of body to introvert. (4) Multiple spermatozoa are always bundled to bunch-like spermato-zeugmata. (5) The spermatozoa have a fusiform head and a hollow cylinder-like mid-piece encircling the anterior end of the flagellum. This combination of characters so far is unique, and indicates a close relationship based on common origin. As a consequence, we unite all species sharing this set of synapomorphies in the genus Echinopsolus Gutt, 1990. The herewith included species are: E. acanthocola Gutt, 1990, E. acutus (Massin, 1992) comb. nov., E. charcoti (Vaney, 1906) comb. nov., E. koehleri (Vaney, 1914) comb. nov., E. mollis (Ludwig & Heding, 1935) comb. nov., E. parvipes Massin, 1992 and E. splendidus (Gutt, 1990) comb. nov.. Because the current assignment of Echinopsolus to the family Psolidae can not be retained, the genus is tranferred to the family Cucumariidae, as relationships to taxa within this family are obvious. The peculiar spermatozoa and spermato-zeugmata of all Echinopsolus species are described using light- and electron-microscopical techniques and the results are evaluated and discussed concerning their taxonomy and phylogeny. 

  4. Effects of reduced glutathione on acrosin activity in frozen-thawed boar spermatozoa.

    Science.gov (United States)

    Estrada, Efrén; Rodríguez-Gil, Joan E; Rivera Del Álamo, Maria M; Peña, Alejandro; Yeste, Marc

    2017-02-01

    In pigs, acrosin activity in extended semen is correlated with reproductive performance and has recently been identified as a freezability marker. Reduced glutathione (GSH) is known to decrease sperm cryodamage and increase the reproductive performance of frozen-thawed boar spermatozoa. However, the effects of GSH on the acrosin activity of good and poor freezability ejaculates (GFE and PFE, respectively) is yet to be examined. The present study investigated how supplementing cryopreservation media with GSH affected acrosin activity in GFE and PFE, as well as the relationship between acrosin activity and reproductive performance in frozen-thawed boar spermatozoa. In addition, we examined whether the increase in fertility rates and litter sizes observed after the addition of 2mM GSH to cryopreservation extenders was related to acrosin activity. Supplementing freezing media with 2mM GSH partially counteracted the cryopreservation-related decrease in acrosin activity in GFE but not PFE. Acrosin activity was found to be significantly correlated with in vivo reproductive performance of frozen-thawed boar semen. In conclusion, the effects of adding GSH to freezing extenders on the acrosin activity of frozen-thawed boar spermatozoa rely on the intrinsic freezability of the ejaculate. Furthermore, the maintenance of proper acrosin activity could contribute to the increase in reproductive performance mediated by GSH.

  5. Labelling of living mammalian spermatozoa with the fluorescent thiol alkylating agent, monobromobimane (MB): immobilization upon exposure to ultraviolet light and analysis of acrosomal status

    International Nuclear Information System (INIS)

    Cummins, J.M.; Fleming, A.D.; Crozet, N.; Kuehl, T.J.; Kosower, N.S.; Yanagimachi, R.

    1986-01-01

    Living spermatozoa of seven mammalian species were treated with the thiol-alkylating fluorescent labelling compound, monobromobimane (MBBR). MB-labelling alone had no effect on sperm motility, nor on the time course or ability of golden hamster spermatozoa to undergo the acrosome reaction when capacitated in vitro. Exposure of MB-labelled spermatozoa to ultraviolet (UV) light and excitation of the MB fluorochrome resulted in virtually immediate immobilization of the spermatozoa without affecting acrosomal status. UV exposure of unlabelled spermatozoa for up to 30 sec had no effect upon motility. Immobilization of MB-labelled spermatozoa depended on the midpiece being irradiated, as irradiation of the head alone, or of the more distal parts of the principal piece, had little or no effect upon motility. Labelling with MB followed by immobilization of individually selected spermatozoa was most useful for detailing the course and site of occurrence of the acrosome reaction during penetration of the cumulus oophorus by golden hamster spermatozoa in vitro. In these often hyperactivated spermatozoa, precise determination of the acrosomal status could not often otherwise be made due to the difficulty in visualizing the acrosomal region of a vigorously thrashing, hyperactivated spermatozoon. This technique should prove valuable in a variety of studies on sperm motility, capacitation and fertilization, and could also be extended to other cell systems

  6. Adjusting cryodiluent composition for improved post-thaw quality of rabbit spermatozoa.

    Directory of Open Access Journals (Sweden)

    Sally E Hall

    Full Text Available Improved fertility following artificial insemination with frozen-thawed spermatozoa would offer rabbit producers faster genetic improvement. Previous work investigating cryoprotectants for rabbit spermatozoa have reported inconsistent results. Semen was collected from three rabbit bucks by artificial vagina and frozen using a standard procedure with varied cryodiluent components. Post-thaw analysis encompassed motility, sperm kinematic parameters and acrosome and membrane integrity. Spermatozoa were evaluated at 0, 2 and 4 h after thawing. Experiment 1 compared diluents with 3.5% dimethyl sulfoxide (DMSO, 1.5% acetamide, 1.75% DMSO + 0.75% acetamide or 3.5% DMSO + 1.5% acetamide. The treatment that resulted in the highest post-thaw motility (P<0.001 and acrosome integrity (P<0.001 was DMSO alone. Experiment 2 compared 3.5, 7 and 10% DMSO in the cryodiluent. The best post-thaw sperm motility (P<0.001 and linearity (P = .002 was in 3.5% DMSO, while 10% DMSO afforded higher acrosome/membrane integrity at this last time point (P<0.05. Experiment 3 varied the cryodiluent to contain either 9 or 17% egg yolk or 9 or 17% low density lipoproteins extracted from whole egg yolk. The treatment with the best post-thaw result was 17% egg yolk (motility, P = 0.01; acrosome/membrane integrity, P<0.001. Experiment 4 compared different carbohydrates in the cryodiluent; 50 mM glucose (TCG, 25 mM glucose with 25 mM sucrose (TCGS low, or 50 mM glucose with 50 mM sucrose (TCGS high. When data were pooled across time points, TCG had significantly higher motility than TCGS high (P = 0.021, but was not different from TCGS low. However, TCG had fewer spermatozoa with intact acrosomes and membranes than both TCGS low and TCGS high (P = .002. Put together, these results indicate that the best cryodiluent for rabbit spermatozoa frozen under the conditions used in this paper is with 7% DMSO and 17% egg yolk in a base medium containing 25 mM glucose and 25 mM sucrose.

  7. Spermatogenesis and spermatozoa ultrastructure of two Dipolydora species (Annelida: Spionidae) from the Sea of Japan.

    Science.gov (United States)

    Radashevsky, Vasily I; Yurchenko, Olga V; Tyurin, Sergey A; Alexandrova, Yana N

    2015-02-01

    Spermatogenesis and the structure of the spermatozoa of two spionid polychaetes Dipolydora bidentata and Dipolydora carunculata are described by light and transmission electron microscopy. Both species are gonochoristic borers in shells of various molluscs. Proliferation of spermatogonia occurs in paired testes regularly arranged in fertile segments, and the rest of spermatogenesis occurs in the coelomic cavity. Early spermatogenesis occurs quite similarly in the two species but results in formation of tetrads of interconnected spermatids in D. bidentata and octads of spermatids in D. carunculata. Three consecutive stages of spermiogenesis are recognized according to the condensation of chromatin in nucleus: (1) early spermatids with heterogeneous, partly clumped chromatin, (2) middle spermatids with homogeneous, coarsely granular chromatin, and (3) late spermatids with homogeneous fibrillar chromatin. Moreover, late stage of spermatids is further classified into two stages, I and II, according to the position of the acrosome and shape of the nucleus. In late spermatids I, the acrosome is situated in the anterior invagination of the funnel-shaped to oval nucleus, whereas in late spermatids II the acrosome is situated on top of the elongated nucleus. Ultrastructural composition of cells at each stage of spermatogenesis is described and illustrated. The possible process of morphogenesis of organelles during spermato- and spermiogenesis is reconstructed for both species. The proacrosomal vesicle first appears in early spermatids near the Golgi complex and then migrates anteriorly; in the middle spermatids, the acrosome comes to lie in a deep anterior nuclear fossa. In late spermatids I, this fossa evaginates and a posterior fossa develops in the nucleus housing basal body and the anterior part of the axoneme. In late spermatids II, the mitochondria elongate and probably reduce in number due to fusion of some of them. The mature spermatozoa in both species are

  8. Gestational Exposure to Bisphenol A Affects the Function and Proteome Profile of F1 Spermatozoa in Adult Mice.

    Science.gov (United States)

    Rahman, Md Saidur; Kwon, Woo-Sung; Karmakar, Polash Chandra; Yoon, Sung-Jae; Ryu, Buom-Yong; Pang, Myung-Geol

    2017-02-01

    Maternal exposure to the endocrine disruptor bisphenol A (BPA) has been linked to offspring reproductive abnormalities. However, exactly how BPA affects offspring fertility remains poorly understood. The aim of the present study was to evaluate the effects of gestational BPA exposure on sperm function, fertility, and proteome profile of F1 spermatozoa in adult mice. Pregnant CD-1 mice (F0) were gavaged with BPA at three different doses (50 μg/kg bw/day, 5 mg/kg bw/day, and 50 mg/kg bw/day) on embryonic days 7 to 14. We investigated the function, fertility, and related processes of F1 spermatozoa at postnatal day 120. We also evaluated protein profiles of F1 spermatozoa to monitor their functional affiliation to disease. BPA inhibited sperm count, motility parameters, and intracellular ATP levels in a dose-dependent manner. These effects appeared to be caused by reduced numbers of stage VIII seminiferous epithelial cells in testis and decreased protein kinase A (PKA) activity and tyrosine phosphorylation in spermatozoa. We also found that BPA compromised average litter size. Proteins differentially expressed in spermatozoa from BPA treatment groups are known to play a critical role in ATP generation, oxidative stress response, fertility, and in the pathogenesis of several diseases. Our study provides mechanistic support for the hypothesis that gestational exposure to BPA alters sperm function and fertility via down-regulation of tyrosine phosphorylation through a PKA-dependent mechanism. In addition, we anticipate that the BPA-induced changes in the sperm proteome might be partly responsible for the observed effects in spermatozoa. Citation: Rahman MS, Kwon WS, Karmakar PC, Yoon SJ, Ryu BY, Pang MG. 2017. Gestational exposure to bisphenol-A affects the function and proteome profile of F1 spermatozoa in adult mice. Environ Health Perspect 125:238-245; http://dx.doi.org/10.1289/EHP378.

  9. Is there a relationship between the chromatin status and DNA fragmentation of boar spermatozoa following freezing-thawing?

    Science.gov (United States)

    Fraser, L; Strzezek, J

    2007-07-15

    In this study a radioisotope method, which is based on the quantitative measurements of tritiated-labeled actinomycin D ((3)H-AMD) incorporation into the sperm nuclei ((3)H-AMD incorporation assay), was used to assess the chromatin status of frozen-thawed boar spermatozoa. This study also tested the hypothesis that frozen-thawed spermatozoa with altered chromatin were susceptible to DNA fragmentation measured with the neutral comet assay (NCA). Boar semen was diluted in lactose-hen egg yolk-glycerol extender (L-HEY) or lactose ostrich egg yolk lipoprotein fractions-glycerol extender (L-LPFo), packaged into aluminum tubes or plastic straws and frozen in a controlled programmable freezer. In Experiment 1, the chromatin status and DNA fragmentation were measured in fresh and frozen-thawed spermatozoa from the same ejaculates. There was a significant increase in sperm chromatin destabilization and DNA fragmentation in frozen-thawed semen as compared with fresh semen. The proportions of spermatozoa labeled with (3)H-AMD were concurrent with elevated levels of sperm DNA fragmentation in K-3 extender, without cryoprotective substances, compared with L-HEY or L-LPFo extender. Regression analysis revealed that the results of the (3)H-AMD incorporation assay and NCA for frozen-thawed spermatozoa were correlated. Boars differed significantly in terms of post-thaw sperm DNA damage. In Experiment 2, the susceptibility of sperm chromatin to decondensation was assessed using a low concentration of heparin. Treatment of frozen-thawed spermatozoa with heparin revealed enhanced (3)H-AMD binding, suggesting nuclear chromatin decondensation. The deterioration in post-thaw sperm viability, such as motility, mitochondrial function and plasma membrane integrity, was concurrent with increased chromatin instability and DNA fragmentation. This is the first report to show that freezing-thawing procedure facilitated destabilization in the chromatin structure of boar spermatozoa, resulting in

  10. Different patterns of metabolic cryo-damage in domestic cat (Felis catus) and cheetah (Acinonyx jubatus) spermatozoa.

    Science.gov (United States)

    Terrell, Kimberly A; Wildt, David E; Anthony, Nicola M; Bavister, Barry D; Leibo, S P; Penfold, Linda M; Marker, Laurie L; Crosier, Adrienne E

    2012-04-01

    Felid spermatozoa are sensitive to cryopreservation-induced damage, but functional losses can be mitigated by post-thaw swim-up or density gradient processing methods that selectively recover motile or structurally-normal spermatozoa, respectively. Despite the importance of sperm energy production to achieving fertilization, there is little knowledge about the influence of cryopreservation or post-thaw processing on felid sperm metabolism. We conducted a comparative study of domestic cat and cheetah sperm metabolism after cryopreservation and post-thaw processing. We hypothesized that freezing/thawing impairs sperm metabolism and that swim-up, but not density gradient centrifugation, recovers metabolically-normal spermatozoa. Ejaculates were cryopreserved, thawed, and processed by swim-up, Accudenz gradient centrifugation, or conventional washing (representing the 'control'). Sperm glucose and pyruvate uptake, lactate production, motility, and acrosomal integrity were assessed. Mitochondrial membrane potential (MMP) was measured in cat spermatozoa. In both species, lactate production, motility, and acrosomal integrity were reduced in post-thaw, washed samples compared to freshly-collected ejaculates. Glucose uptake was minimal pre- and post-cryopreservation, whereas pyruvate uptake was similar between treatments due to high coefficients of variation. In the cat, swim-up, but not Accudenz processing, recovered spermatozoa with increased lactate production, pyruvate uptake, and motility compared to controls. Although confounded by differences in non-specific fluorescence among processing methods, MMP values within treatments were positively correlated to sperm motility and acrosomal integrity. Cheetah spermatozoa isolated by either selection method exhibited improved motility and/or acrosomal integrity, but remained metabolically compromised. Collectively, findings revealed a metabolically-robust subpopulation of cryopreserved cat, but not cheetah, spermatozoa

  11. pKI values of prazosin and idazoxan for receptors stimulated by neuronally released transmitter in the epididymal portion of rat isolated vas deferens.

    OpenAIRE

    Mackay, D.; Kengatharan, M.

    1994-01-01

    1. A new method has been used to measure pKI values of prazosin and idazoxan against neuronally-released transmitter in the epididymal portion of the rat isolated vas deferens. The most reproducible results were obtained with a prolonged antagonist equilibration time (1 h). 2. Under these conditions the pKI of prazosin was practically unaffected by addition of alpha, beta-methylene-adenosine-5'-triphosphate (10 microM) to desensitize purinoceptors. Addition of desmethylimipramine (DMI) (0.3 m...

  12. The efficiency of conventional microscopic selection is comparable to the hyaluronic acid binding method in selecting spermatozoa for male infertility patients.

    Science.gov (United States)

    Huang, Meng-Ting; Kuo-Kuang Lee, Robert; Lu, Chung-Hao; Chen, Ying-Jie; Li, Sheng-Hsiang; Hwu, Yuh-Ming

    2015-02-01

    To evaluate if hyaluronic acid (HA)-bound spermatozoa surpassed conventional microscopy-selected spermatozoa in the status of sperm DNA integrity by acridine orange (AO) fluorescence staining. Spermatozoa obtained from couples with indication for the intracytoplasmic sperm injection (ICSI) procedure due to male infertility (n = 34) and control males with normal sperm parameters (n = 12) were analyzed using AO fluorescence staining after density-gradient centrifugation (DGC), polyvinylpyrrolidone (PVP)-microscopic selection, and HA-binding selection to determine sperm DNA integrity. Percentages of DNA intact spermatozoa with green fluorescence were significantly higher in both PVP-microscopic selected spermatozoa (82.1 ± 24.0%) and HA-bound spermatozoa (83.9 ± 21.1%) than in spermatozoa prepared by DGC (66.8 ± 24.0%). However, there was no significant difference between the PVP-sperm and HA-sperm groups. When the percentage of green fluorescent spermatozoa prepared by DGC fell initially below 68%, both PVP-microscopic and HA-binding selection failed to select over 90% spermatozoa with intact DNA for ICSI in the male infertility group. Compared to control males with normal sperm parameters (99.3 ± 1.8%), the proportion of green fluorescence sperm after HA-binding selection from couples with male infertility (83.9 ± 21.1%) did not reach the range of > 99% reported by Yagci et al. The percentages of DNA intact spermatozoa between the PVP-sperm and HA-sperm groups were not significantly different. In an ICSI procedure, a well-trained embryologist will have the same ability to choose sperm with intact DNA by conventional microscopic selection as with HA-bound spermatozoa selection. Copyright © 2014. Published by Elsevier B.V.

  13. Influence of environmentally relevant concentrations of vinclozolin on quality, DNA integrity, and antioxidant responses of sterlet Acipenser ruthenus spermatozoa.

    Science.gov (United States)

    Gazo, Ievgeniia; Linhartova, Pavla; Shaliutina, Anna; Hulak, Martin

    2013-04-25

    The effects of vinclozolin (VIN), an anti-androgenic fungicide, on quality, oxidative stress, DNA integrity, and ATP level of sterlet (Acipenser ruthenus) spermatozoa were investigated in vitro. Fish spermatozoa were incubated with different concentrations of vinclozolin (0.5, 2, 10, 15, 20 and 50 μg/l) for 2 h. A dose-dependent reduction in spermatozoa motility and velocity was observed at concentrations of 2-50 μg/l. A dramatic increase in DNA fragmentation was recorded at concentrations 10 μg/l and above. After 2 h exposure at higher test concentrations (10-50 μg/l), oxidative stress was apparent, as reflected by significantly higher levels of protein and lipid oxidation and significantly greater superoxide dismutase activity. Intracellular ATP content of spermatozoa decreased with increasing concentrations of VIN. The results demonstrated that VIN can induce reactive oxygen species stress in fish spermatozoa, which could impair the sperm quality, DNA integrity, ATP content, and the antioxidant defense system. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  14. Spermatozoa isolated from cat testes retain their structural integrity as well as a developmental potential after refrigeration for up to 7 days.

    Science.gov (United States)

    Buarpung, Sirirak; Tharasanit, Theerawat; Thongkittidilok, Chommanart; Comizzoli, Pierre; Techakumphu, Mongkol

    2015-10-01

    The objective of this study was to compare the efficiency of preservation media for isolated feline testicular spermatozoa as well as the concentrations of bovine serum albumin (BSA) on: (1) the membrane (sperm membrane integrity (SMI)) and DNA integrity of spermatozoa; and (2) the developmental potential of spermatozoa after intracytoplasmic sperm injection (ICSI). Isolated cat spermatozoa were stored in HEPES-M199 medium (HM) or Dulbecco's phosphate-buffered saline (DPBS) at 4°C for up to 7 days. Results indicated that HM maintained a better SMI than DPBS throughout the storage periods (P > 0.05). When spermatozoa were stored in HM supplemented with BSA at different concentrations (4, 8 or 16 mg/ml), SMI obtained from HM containing 8 and 16 mg/ml BSA was higher than with 4 mg/ml BSA (P 0.05). In summary, cat spermatozoa immediately isolated from testicular tissue can be stored as a suspension in basic buffered medium at 4°C for up to 7 days. BSA supplementation into the medium improves membrane integrity of the spermatozoa during cold storage. Testicular spermatozoa stored in HM containing 16 mg/ml BSA retained full in vitro developmental potential after ICSI, similar to that of fresh controls even though DNA integrity had slightly declined.

  15. The efficiency of conventional microscopic selection is comparable to the hyaluronic acid binding method in selecting spermatozoa for male infertility patients

    Directory of Open Access Journals (Sweden)

    Meng-Ting Huang

    2015-02-01

    Conclusion: The percentages of DNA intact spermatozoa between the PVP-sperm and HA-sperm groups were not significantly different. In an ICSI procedure, a well-trained embryologist will have the same ability to choose sperm with intact DNA by conventional microscopic selection as with HA-bound spermatozoa selection.

  16. Protective effect of pumpkin seed extract on sperm characteristics, biochemical parameters and epididymal histology in adult male rats treated with cyclophosphamide.

    Science.gov (United States)

    Aghaei, S; Nikzad, H; Taghizadeh, M; Tameh, A A; Taherian, A; Moravveji, A

    2014-10-01

    Cancer treatment with cyclophosphamide (CP) may result in reproductive toxicity as one of its side effects. The pumpkin seed is a rich natural source of antioxidant. We have assessed the possible protective efficacy of pumpkin seed extract on sperm characteristics, biochemical parameters and epididymal histology of CP-treated rats. Male adult Wistar rats were categorised into four groups. Group 1 served as control and received intraperitoneal (IP) injection of isotonic saline solution. Group 2 rats were treated with CP by IP injection in a single dose of 100 mg/kg body weight, only once. Group 3 and 4 received CP plus 300 and 600 mg/kg pumpkin seed extract respectively. Six weeks after treatment, sperm characteristics, biochemical parameters and histopathological changes were examined. Results showed that, sperm characteristics in CP-treated rats were significantly decreased. Biochemical analysis results showed that the co-administration of 300 mg pumpkin seed extract could increase the total antioxidant capacity (TAC) level significantly. In CP-treated rats, histopathological changes such as vacuolisation, disorganisation and separation of epididymal epithelium were observed as well. Interestingly, pumpkin seed extract could improve the above-mentioned parameters remarkably in CP-treated rats. Our findings indicated that pumpkin seed extract might be used as protective agent against CP-induced reproductive toxicity. © 2013 Blackwell Verlag GmbH.

  17. Probe-based confocal laser endomicroscopy (pCLE) - a new imaging technique for in situ localization of spermatozoa.

    Science.gov (United States)

    Trottmann, Matthias; Stepp, Herbert; Sroka, Ronald; Heide, Michael; Liedl, Bernhard; Reese, Sven; Becker, Armin J; Stief, Christian G; Kölle, Sabine

    2015-05-01

    In azoospermic patients, spermatozoa are routinely obtained by testicular sperm extraction (TESE). However, success rates of this technique are moderate, because the site of excision of testicular tissue is determined arbitrarily. Therefore the aim of this study was to establish probe-based laser endomicroscopy (pCLE) a noval biomedical imaging technique, which provides the opportunity of non-invasive, real-time visualisation of tissue at histological resolution. Using pCLE we clearly visualized longitudinal and horizontal views of the tubuli seminiferi contorti and localized vital spermatozoa. Obtained images and real-time videos were subsequently compared with confocal laser scanning microscopy (CLSM) of spermatozoa and tissues, respectively. Comparative visualization of single native Confocal laser scanning microscopy (CLSM, left) and probe-based laser endomicroscopy (pCLE, right) using Pro Flex(TM) UltraMini O after staining with acriflavine. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Evaluation of motility, membrane status and DNA integrity of frozen-thawed bottlenose dolphin (Tursiops truncatus) spermatozoa after sex-sorting and recryopreservation.

    Science.gov (United States)

    Montano, G A; Kraemer, D C; Love, C C; Robeck, T R; O'Brien, J K

    2012-06-01

    Artificial insemination (AI) with sex-sorted frozen-thawed spermatozoa has led to enhanced management of ex situ bottlenose dolphin populations. Extended distance of animals from the sorting facility can be overcome by the use of frozen-thawed, sorted and recryopreserved spermatozoa. Although one bottlenose dolphin calf had been born using sexed frozen-thawed spermatozoa derived from frozen semen, a critical evaluation of in vitro sperm quality is needed to justify the routine use of such samples in AI programs. Sperm motility parameters and plasma membrane integrity were influenced by stage of the sex-sorting process, sperm type (non-sorted and sorted) and freezing method (straw and directional) (P0.05) at 24 h. The viability of sorted spermatozoa was higher (Pdolphin spermatozoa undergoing cryopreservation, sorting and recryopreservation are of adequate quality for use in AI.

  19. Can the rapid identification of mature spermatozoa during microdissection testicular sperm extraction guide operative planning?

    Science.gov (United States)

    Alrabeeah, K; Doucet, R; Boulet, E; Phillips, S; Al-Hathal, N; Bissonnette, F; Kadoch, I J; Zini, A

    2015-05-01

    The minimum sperm count and quality that must be identified during microdissection testicular sperm extraction (micro-TESE) to deem the procedure successful remains to be established. We conducted a retrospective study of 81 consecutive men with non-obstructive azoospermia who underwent a primary (first) micro-TESE between March 2007 and October 2013. Final assessment of sperm recovery [reported on the day of (intracytoplasmic sperm injection) ICSI] was recorded as (i) successful (available spermatozoa for ICSI) or (ii) unsuccessful (no spermatozoa for ICSI). The decision to perform a unilateral (with limited or complete microdissection) or bilateral micro-TESE was guided by the intra-operative identification of sperm recovery (≥5 motile or non-motile sperm) from the first testicle. Overall, sperm recovery was successful in 56% (45/81) of the men. A unilateral micro-TESE was performed in 47% (38/81) of the men (based on intra-operative identification of sperm) and in 100% (38/38) of these men, spermatozoa was found on final assessment. In 42% (16/38) of the unilateral cases, a limited microdissection was performed (owing to the rapid intra-operative identification of sperm). The remaining 43 men underwent a bilateral micro-TESE and 16% (7/43) of these men had sperm identified on final assessment. The cumulative ICSI pregnancy rates (per cycle started and per embryo transfer) were 47% (21/45) and 60% (21/35), respectively, with a mean (±SD) of 1.9 ± 1.0 embryos transferred. The data demonstrate that intra-operative assessment of sperm recovery can correctly identify those men that require a unilateral micro-TESE. Moreover, the rapid identification of sperm recovery can allow some men to undergo a limited unilateral micro-TESE and avoid the need for complete testicular microdissection. © 2015 American Society of Andrology and European Academy of Andrology.

  20. Regulatory properties of 6-phosphofructokinase and control of glycolysis in boar spermatozoa.

    Science.gov (United States)

    Kamp, G; Schmidt, H; Stypa, H; Feiden, S; Mahling, C; Wegener, G

    2007-01-01

    Glycolysis is crucial for sperm functions (motility and fertilization), but how this pathway is regulated in spermatozoa is not clear. This prompted to study the location and the regulatory properties of 6-phosphofructokinase (PFK, EC 2.7.1.11), the most important element for control of glycolytic flux. Unlike some other glycolytic enzymes, PFK showed no tight binding to sperm structures. It could readily be extracted from ejaculated boar spermatozoa by sonication and was then chromatographically purified. At physiological pH, the enzyme was allosterically inhibited by near-physiological concentrations of its co-substrate ATP, which induced co-operativity, i.e. reduced the affinity for the substrate fructose 6-phosphate. Inhibition by ATP was reinforced by citrate and H+. Above pH 8, PFK lost all its regulatory properties and showed maximum activity. However, in the physiological pH range, PFK activity was very sensitive to small changes in effectors. At near-physiological substrate concentrations, PFK activity requires activators (de-inhibitors) of which the combination of AMP and fructose 2,6-bisphosphate (F2,6P2) was most efficient as a result of synergistic effects. The kinetics of PFK suggest AMP, F2,6P2, H+, and citrate as allosteric effectors controlling PFK activity in boar spermatozoa. Using immunogold labeling, PFK was localized in the mid-piece and principal piece of the flagellum as well as in the acrosomal area at the top of the head and in the cytoplasmic droplets released from the mid-piece after ejaculation.

  1. CURCUMIN IN MALE FERTILITY: EFFECTS ON SPERMATOZOA VITALITY AND OXIDATIVE BALANCE

    Directory of Open Access Journals (Sweden)

    Eva Tvrdá

    2015-02-01

    Full Text Available The aim of this study was to assess the dose- and time-dependent effects of curcumin on bovine spermatozoa during short-term (0h, 2h, 6h and long-term (12h, 24h in vitro culture periods. Semen samples were collected from 20 adult breeding bulls, and diluted in physiological saline solution containing 0.5% DMSO together with 0, 1, 5, 10, 50 and 100 μM/L of curcumin. Spermatozoa motion parameters were determined using the SpermVisionTM and CASA (Computer Assisted Semen Analyzer system. Cell viability was measured using the metabolic activity MTT assay, and the nitroblue-tetrazolium (NBT test was used to assess the intracellular superoxide formation. The CASA analysis revealed that concentrations of 50 μM/L and 10 μM/L of curcumin were able to significantly prevent the decrease of motility and progressive motility (P<0.001 in case of group B and P<0.01 in case of group C over all time periods of the in vitro incubation. At the same time, supplementation of concentrations ranging from 50 μM/L to 5 μM/L of curcumin led to a significant preservation of the cell viability in comparison to the control (P<0.001 in case of groups B and C; P<0.05 in case of group D. Concentrations in between 50 μM/L and 5 μM/L of curcumin demonstrated antioxidant properties, translated in a significant reduction of the intracellular superoxide production throughout the in vitro culture (P<0.001. The results indicate that the addition of curcumin, especially in concentrations between 50 μM and 10 μM to the culture medium could be beneficial for a complex enhancement of spermatozoa activity and protection against complications resulting from in vitro culture.

  2. Characterizing the glycocalyx of poultry spermatozoa: I. Identification and distribution of carbohydrate residues using flow cytometry and epifluorescence microscopy.

    Science.gov (United States)

    Peláez, Jesús; Long, Julie A

    2007-01-01

    The aim of the present work was to use a battery of lectins to 1) delineate the carbohydrate content of sperm glycocalyx in the turkey and chicken using flow cytometry analysis, and 2) evaluate the distribution of existing sugars over the sperm plasma membrane surface with epifluorescent microscopy. Carbohydrate groups (corresponding lectins) that were investigated included galactose (GS-I, Jacalin, RCA-I, PNA), glucose and/or mannose (Con A, PSA, GNA), N-acetyl-glucosamine (GS-II, s-WGA, STA), N-acetyl-galactosamine (SBA, WFA), fucose (Lotus, UEA-I), sialic acid (LFA, LPA), and N-acetyl-lactosamine (ECA). Spermatozoa were assessed before and after treatment with neuraminidase to remove sialic acid. Mean fluorescence intensity (MnFI) was used as indicator of lectin binding for flow cytometry analysis. Nontreated spermatozoa from both species showed high MnFI when incubated with RCA-I, Con A, LFA, and LPA, as did chicken spermatozoa incubated with s-WGA. Neuraminidase treatment increased the MnFI for most lectins except LFA and LPA, as expected. Differences in MnFI between species included higher values for s-WGA and ECA in chicken spermatozoa and for WFA in turkey spermatozoa. Microscopy revealed segregation of some sugar residues into membrane-specific domains; however, the 2 staining techniques (cell suspension vs fixed preparation) differed in identifying lectin binding patterns, with fixed preparations yielding a high degree of nonspecific binding. We conclude that 1) the glycocalyx of turkey and chicken spermatozoa contains a diversity of carbohydrate groups, 2) these residues are extensively masked by sialic acid, 3) the glycocalyx composition is species-specific, and 4) some glycoconjugates appear to be segregated into membrane-specific domains. Characterization of the poultry sperm glycocalyx is the first step in identifying the physiological impact of semen storage on sperm function.

  3. Fraksi Etanol 96% Biji Koro Benguk (Mucuna Pruriens L.) Sebagai Peningkat Kualitas Spermatozoa Mencit (Mus Musculus)

    OpenAIRE

    Winarni, Sri; Judiwati, Rina; Prajogo, Bambang; Hayati, Alfiah

    2011-01-01

    Background: The examination of sperm quality is the main priority for infertility diagnosis. Based on previous study with mice, active ingredient of Mucuna pruriens L. or koro benguk (Papilionaceae), the L-dopa, may affect the quality of spermatozoa.Objective: Research was to study the effect of 96% ethanol fraction Mucuna pruriens seed on spermatozoaquality of mice exposed to 2-Methoxy ethanol. L-dopa in 96% ethanol fraction of M. pruriens seed was 14.7%.Methode: This was an experimental stu...

  4. Obesity and Bariatric Surgery Drive Epigenetic Variation of Spermatozoa in Humans

    DEFF Research Database (Denmark)

    Donkin, Ida; Versteyhe, Soetkin; Ingerslev, Lars R.

    2016-01-01

    Obesity is a heritable disorder, with children of obese fathers at higher risk of developing obesity. Environmental factors epigenetically influence somatic tissues, but the contribution of these factors to the establishment of epigenetic patterns in human gametes is unknown. Here, we hypothesized...... of morbidly obese men, surgery-induced weight loss was associated with a dramatic remodeling of sperm DNA methylation, notably at genetic locations implicated in the central control of appetite. Our data provide evidence that the epigenome of human spermatozoa dynamically changes under environmental pressure...

  5. Binding of antibodies to dsDNA by UVB-irradiated spermatozoa in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Wollina, U; Schaarschmidt, H; Thiel, W; Knopf, B

    1986-08-01

    Human spermatozoa were exposed to monochromatic UVB light (300 nm) at a dosage range from 1-10/sup 3/ J/m/sup 2/ (xenon-mercury short-arc lamp) and 10/sup -2/-10/sup -1/ J/m/sup 2/ (xenon lamp). UVB irradiation >= 10 J/m/sup 2/ resulted in a subsequent binding of antibodies to double-stranded (ds) DNA. A strong homogeneous head fluorescence was visible. There was no immunoglobulin class restriction. The results were discussed in regards to lupus antigen expression by UV light.

  6. Mobile phone radiation induces reactive oxygen species production and DNA damage in human spermatozoa in vitro.

    Directory of Open Access Journals (Sweden)

    Geoffry N De Iuliis

    Full Text Available BACKGROUND: In recent times there has been some controversy over the impact of electromagnetic radiation on human health. The significance of mobile phone radiation on male reproduction is a key element of this debate since several studies have suggested a relationship between mobile phone use and semen quality. The potential mechanisms involved have not been established, however, human spermatozoa are known to be particularly vulnerable to oxidative stress by virtue of the abundant availability of substrates for free radical attack and the lack of cytoplasmic space to accommodate antioxidant enzymes. Moreover, the induction of oxidative stress in these cells not only perturbs their capacity for fertilization but also contributes to sperm DNA damage. The latter has, in turn, been linked with poor fertility, an increased incidence of miscarriage and morbidity in the offspring, including childhood cancer. In light of these associations, we have analyzed the influence of RF-EMR on the cell biology of human spermatozoa in vitro. PRINCIPAL FINDINGS: Purified human spermatozoa were exposed to radio-frequency electromagnetic radiation (RF-EMR tuned to 1.8 GHz and covering a range of specific absorption rates (SAR from 0.4 W/kg to 27.5 W/kg. In step with increasing SAR, motility and vitality were significantly reduced after RF-EMR exposure, while the mitochondrial generation of reactive oxygen species and DNA fragmentation were significantly elevated (P<0.001. Furthermore, we also observed highly significant relationships between SAR, the oxidative DNA damage bio-marker, 8-OH-dG, and DNA fragmentation after RF-EMR exposure. CONCLUSIONS: RF-EMR in both the power density and frequency range of mobile phones enhances mitochondrial reactive oxygen species generation by human spermatozoa, decreasing the motility and vitality of these cells while stimulating DNA base adduct formation and, ultimately DNA fragmentation. These findings have clear implications

  7. Are there optimal numbers of oocytes, spermatozoa and embryos in assisted reproduction?

    Science.gov (United States)

    Milachich, Tanya; Shterev, Atanas

    2016-08-01

    The aim of this overview is to discuss the current information about the search for the optimum yield of gametes in assisted reproduction, as one of the major pillars of IVF success. The first topic is focused on the number of male gametes and the possible impact of some genetic traits on these parameters. The number of spermatozoa did not seem to be crucial when there is no severe male factor of infertility. Genetic testing prior to using those sperm cells is very important. Different methods were applied in order to elect the "best" spermatozoa according to specific indications. The next problem discussed is the importance of the number of oocytes collected. Several studies have agreed that "15 oocytes is the perfect number," as the number of mature oocytes is more important. However, if elective single embryo transfer is performed, the optimal number of oocytes will enable a proper embryo selection. The third problem discussed concerns fertility preservation. Many educational programs promote and encourage procreation at maternal ages between 20-35 years, since assisted reproduction is unable to fully overcome the effects of female aging and fertility loss after that age. It is also strongly recommended to ensure a reasonable number of cryopreserved mature oocytes, preferably in younger ages (average of two stimulation cycles are likely required. For embryo cryopreservation, the "freeze all" strategy suggests the vitrification of good embryos, therefore quality is prior to number and patient recruitment for this strategy should be performed cautiously.

  8. Soybean lecithin-based extender preserves spermatozoa membrane integrity and fertilizing potential during goat semen cryopreservation.

    Science.gov (United States)

    Chelucci, Sara; Pasciu, Valeria; Succu, Sara; Addis, Daniela; Leoni, Giovanni G; Manca, Maria E; Naitana, Salvatore; Berlinguer, Fiammetta

    2015-04-01

    Soybean lecithin may represent a suitable alternative to egg yolk for semen cryopreservation in livestock species. However, additional studies are needed to elucidate its effects on spermatozoa functional properties. Semen collected from five Sarda bucks was cryopreserved in Tris-based extender and glycerol (4% v:v) with different supplementations. In a preliminary experiment, different soybean lecithin concentrations were tested (1%-6% wt/vol) and results in terms of viability, percentages of progressive motile and rapid spermatozoa, and DNA integrity after thawing showed that the most effective concentration was 1%. In the second experiment, semen was frozen in a Tris-based extender with no supplementation (EXT), with 1% lecithin (EXT LC), and 20% egg yolk (EXT EY). The effectiveness of these extenders was also compared with a commercial extender. The EXT EY led to the highest viability and motility parameters after freezing and thawing (P lecithin can be considered as a suitable alternative to egg yolk in goat semen cryopreservation, because it ensures higher fertilization rates and a better protection from membrane damage by cold shock. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. The Assessment of DNA Damage in Poultry Spermatozoa after Exposure to Low Doses of Ionising Radiation

    International Nuclear Information System (INIS)

    Kasuba, V.; Milic, M.; Pejakovic Hlede, J.; Gottstein, Z.; Karadjole, M.; Miljanic, S.

    2013-01-01

    The existence of dose-related induction of DNA strand breaks in spermatozoa following in vitro exposure to ionising radiation represents sperm DNA integrity as an important parameter in the evaluation of semen functionality. Maintaining of normal sperm becomes even more important when it is known that DNA in semen samples is already fragmentated in certain amount in human and turkey semen and that it lacks DNA repair mechanisms making DNA damage irreversible. The aim of this paper was to provide an insight in the amount of DNA damage detected in chicken spermatozoa (5 cocks, 45 weeks old) of heavy line after radiation with doses of 0.3, 0.5, 1 and 2 Gy gamma radiation and to address the question of the potential ecological consequences of the damage that was measured with comet assay. Scored parameters included tail intensity, tail length and tail moment. Results showed sensitivity of comet assay technique that detected significant DNA damage even after exposure to 0.3 Gy, but also showed no dose-related responses after 0.5, 1 and 2 Gy. Distribution of damaged cells was widely spread for the higher doses, showing the influence of possible adaptive response, but for further conclusions, larger studies are needed to answer that question.(author)

  10. Expression, Localization of SUMO-1, and Analyses of Potential SUMOylated Proteins in Bubalus bubalis Spermatozoa.

    Science.gov (United States)

    Brohi, Rahim Dad; Wang, Li; Hassine, Najla Ben; Cao, Jing; Talpur, Hira Sajjad; Wu, Di; Huang, Chun-Jie; Rehman, Zia-Ur; Bhattarai, Dinesh; Huo, Li-Jun

    2017-01-01

    Mature spermatozoa have highly condensed DNA that is essentially silent both transcriptionally and translationally. Therefore, post translational modifications are very important for regulating sperm motility, morphology, and for male fertility in general. Protein sumoylation was recently demonstrated in human and rodent spermatozoa, with potential consequences for sperm motility and DNA integrity. We examined the expression and localization of small ubiquitin-related modifier-1 (SUMO-1) in the sperm of water buffalo ( Bubalus bubalis ) using immunofluorescence analysis. We confirmed the expression of SUMO-1 in the acrosome. We further found that SUMO-1 was lost if the acrosome reaction was induced by calcium ionophore A23187. Proteins modified or conjugated by SUMO-1 in water buffalo sperm were pulled down and analyzed by mass spectrometry. Sixty proteins were identified, including proteins important for sperm morphology and motility, such as relaxin receptors and cytoskeletal proteins, including tubulin chains, actins, and dyneins. Forty-six proteins were predicted as potential sumoylation targets. The expression of SUMO-1 in the acrosome region of water buffalo sperm and the identification of potentially SUMOylated proteins important for sperm function implicates sumoylation as a crucial PTM related to sperm function.

  11. Expression, Localization of SUMO-1, and Analyses of Potential SUMOylated Proteins in Bubalus bubalis Spermatozoa

    Directory of Open Access Journals (Sweden)

    Rahim Dad Brohi

    2017-06-01

    Full Text Available Mature spermatozoa have highly condensed DNA that is essentially silent both transcriptionally and translationally. Therefore, post translational modifications are very important for regulating sperm motility, morphology, and for male fertility in general. Protein sumoylation was recently demonstrated in human and rodent spermatozoa, with potential consequences for sperm motility and DNA integrity. We examined the expression and localization of small ubiquitin-related modifier-1 (SUMO-1 in the sperm of water buffalo (Bubalus bubalis using immunofluorescence analysis. We confirmed the expression of SUMO-1 in the acrosome. We further found that SUMO-1 was lost if the acrosome reaction was induced by calcium ionophore A23187. Proteins modified or conjugated by SUMO-1 in water buffalo sperm were pulled down and analyzed by mass spectrometry. Sixty proteins were identified, including proteins important for sperm morphology and motility, such as relaxin receptors and cytoskeletal proteins, including tubulin chains, actins, and dyneins. Forty-six proteins were predicted as potential sumoylation targets. The expression of SUMO-1 in the acrosome region of water buffalo sperm and the identification of potentially SUMOylated proteins important for sperm function implicates sumoylation as a crucial PTM related to sperm function.

  12. Freezability of water buffalo spermatozoa is improved with the addition of catalase in cryodiluent.

    Science.gov (United States)

    Ali, L; Hassan Andrabi, S M; Ahmed, H; Hussain Shah, A A

    Catalase enzyme is usually distributed in mammalian seminal plasma, where it decomposes hydrogen peroxide into water and oxygen and enhances sperm survivability. To evaluate the effect of catalase (0, 100, 200 or 300 IU/ml) added in tris-citric acid (TCA) based extender on motion characteristics, viability and DNA integrity of bubaline spermatozoa at post dilution (PD) and post thawing (PT) stages of cryopreservation. Collection of semen was done in four Nili-Ravi bulls with an artificial vagina (42 degree C). Qualified semen samples from each bull were further subdivided into four aliquots for dilution with the experimental TCA extender containing either 0.0 (T1), 100 IU (T2), 200 IU (T3) or 300 IU (T4) catalase (activity12660 U/mg). At PT, mean computer progressive motility, average path velocity, straight line velocity, curvilinear velocity, visual motility and DNA integrity were higher (P catalase fortified treatment groups as compared with control. Regarding plasma membrane integrity and supra-vital plasma membrane integrity, at PT the mean values were higher (P catalase at a concentration of 300IU/ml in TCA cryodiluent improved the freezability of water buffalo spermatozoa.

  13. Lipsome-mediated uptake of exogenous DNA by Sahiwal cattle spermatozoa

    Directory of Open Access Journals (Sweden)

    Thomas V. Babu

    Full Text Available Aim: To investigate the influence of lipofection treatment and exogenous DNA uptake on the quality of sahiwal cattle spermatozoa. Materials and Methods: Semen collected from sahiwal bulls (n=7 were evaluated separately for color, volume, mass activity, concentration, motility and viability using standard procedures. Pooled sperm samples from selected bulls (n=3 were transfected with a model gene construct enhanced green fluorescent protein (p-EGFP via lipofection method and confirmed the genome integration by PCR technique. Furthermore the effect of transfection on spermatozoa was assessed based on apotosis, viability and motility. Results: In the current investigation sahiwal bulls were selected based on their breeding records and better semen characteristics. Although the transfected sperm samples failed to show florescence under fluorescence microscope, PCR studies confirmed the successful uptake of the p-EGFP gene in to the host sperm cell genome. Moreover transfected samples showed a significant reduction in the viability and motility without causing any DNA damage induced apoptosis as demonstrated by DNA Ladder assay. [Vet World 2012; 5(10.000: 621-627

  14. Efektivitas Penambahan Vitamin E (alfa-Tokoferol dalam Medium Pencucian Sperma dengan Sentrifugasi terhadap Kualitas Spermatozoa Sapi Brahman

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    Dasrul Dasrul

    2012-10-01

    Full Text Available Effect of vitamin E addition (alfa-tokoferol into sperm washing medium by centrifuge on the quality of Brahman cattle spermatozoa  ABSTRACT. The aims of study to determine the effectiveness of the addition of vitamin E in the washing medium by centrifugation on sperm quality Brahman cattle. frozen semen of Brahman cattle, divided into 4 treatment groups addition of vitamin E in the washing medium: 0.0gr/100 ml medium (K0, 0.1gr /100 ml medium (K1; 0.2gr/100 ml medium (K2 and 0.3 g / 100 ml medium (K4, each group was repeated 5 times. Examination of motility, viability and integrity of sperm membrane performed according to WHO standards. The data obtained were analyzed with one-way ANOVA and Duncan test. The average percentage of motility, viability and membrane integrity of spermatozoa in the addition of vitamin E were significantly different (P 0.05 compared with the group K0. The addition of vitamin E in the medium on the process of washing spermatozoa Brahman cattle. The addition of vitamin E 0.2gr/100ml better than vitamin E 0.1gr/100ml and 0.3gr/100ml in maintaining the percentage of motility and live spermatozoa Brahman cattle.

  15. Radiation-induced DNA single-strand scission and its rejoining in spermatogonia and spermatozoa of mouse

    International Nuclear Information System (INIS)

    Ono, T.; Okada, S.

    1977-01-01

    Gamma-ray-induced DNA single-strand scissions and the ability to repair the scissions in spermatogonia from young mice and in spermatozoa from adult mice were studied quantitatively by an alkaline sucrose density-gradient centrifugation method. The average size of DNAs in non-irradiated spermatogonia was 2.6-3.0xx10 8 daltons, similar to those of a spermatid-rich population, and the size of DNA in non-irradiated spermatozoa was 1.2x10 8 daltons. In spermatogonia, the radiosensitivity of DNA was 0.42 single-strand breaks/10 12 daltons of DNA/rad in oxic conditions and only 0.24 under anoxic conditions. In spermatozoa the break efficiency of DNA was 0.22 single-strand breaks/10 12 daltons of DNA/rad under oxic conditions and altered little under anoxic irradiation. The DNA scissions were efficiently repaired in spermatogonia within 10 min, whereas the breaks in spermatozoa were not rejoined at all even after two days of post-irradiation time. The radiosensitivities of DNA, repair capability and non- and/or slowreparable DNA scissions were compared in spermatogonium-rich, spermatid-rich and spermatozoanrich populations

  16. THE EFFECT OF IN VITRO SEMEN STORAGE TEMPERATURE AND AGE OF MALES ON SPERMATOZOA MOTILITY PARAMETERS OF TURKEYS SEMEN

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    Joanna Paluch

    2013-02-01

    Full Text Available This work was to evaluate the effect of in vitro storage temperature and age of males on turkey spermatozoa motility. For this purpose samples were collected from British United Turkeys (BUT Big 6 line and semen quality was assessed by using Computer Assisted Semen Analyzer (CASA system. After 60 minutes of storage spermatozoa motility, progressive motility and amplitude of lateral head displacement decreased significantly both in 4° and 41°C regardless of birds age. However the lowest values of all parameters were noted after storage in thermostat. Spermatozoa motility after 0 and 60 minutes in 4°C was higher in samples collected from turkeys of 35 – 42 weeks of age (60.94% and 53.33% respectively. Whereas the value of that parameter in semen stored in 41°C was lower in that age group. The same tendency was found in progressive motility. The results showed that higher temperature of in vitro storage (even that similar to animal body temperature, in this case 41ºC has more negative effect on spermatozoa motility parameters than lower temperature.

  17. Spermiogenesis and spermatozoa ultrastructure in five species of the Curimatidae with some considerations on spermatozoal ultrastructure in the Characiformes

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    Irani Quagio-Grassiotto

    Full Text Available Spermiogenesis in the curimatid species, Steindachnerina insculpta, Cyphocharax gillii, C. modestus, C. spilotus, and Potamorhina altamazonica, is characterized by lateral development of the flagellum, nuclear rotation, eccentric nuclear fossa formation, and chromatin compacted into thick fibers. These spermatozoa exhibit a spherical head containing a nucleus with the chromatin highly condensed into thick fibers with small electron-lucent areas, and no acrosome. The nuclear fossa is of the moderate type and eccentric and penetrated by the centriolar complex. The midpiece is small, has many elongate vesicles, and a short cytoplasmic channel. Mitochondria may be elongate, branched or C-shaped, and are separated from the initial segment of the axoneme by the cytoplasmic channel. The flagellum contains the classical axoneme structure (9+2 and has a membranous compartment in the initial region; it does not have lateral fins. Only small differences were observed among the analyzed species and genera of the Curimatidae. Spermiogenesis and spermatozoa in the Curimatidae have many of the characteristics found in almost all other characiform species. On the other hand, the presence of a membranous compartment in the initial region of curimatid flagella, a structure common in many Cypriniformes spermatozoa, is unknown in other characiforms. Spermiogenesis and the spermatozoa of the Characiformes are discussed.

  18. The in vitro effect of Lambda-cyhalothrin on quality and antioxidant responses of rainbow trout Oncorhynchus mykiss spermatozoa.

    Science.gov (United States)

    Kutluyer, Filiz; Erişir, Mine; Benzer, Fulya; Öğretmen, Fatih; İnanan, Burak Evren

    2015-11-01

    There is little information in the scientific literature about effect of in vitro exposure of fish spermatozoa to pesticides. In vitro effect of Lambda-cyhalothrin (LCT) on sperm quality and oxidative stress has not been fully explored yet. The effects of LCT, which is a synthetic pyrethroid insecticide, on quality and oxidative stress of spermatozoa were investigated in vitro due to extensively use to control a wide range of insect pests in agriculture, public health, and homes and gardens. To explore the potential in vitro toxicity of LCT, fish spermatozoa were incubated with different concentrations of LCT (0.6, 1.2 and 2.4 μg/L) for 2h. Reduced glutathione (GSH), glutathione peroxidase (GSH-Px), catalase (CAT) and malondialdehyde (MDA) in spermatozoa were analyzed for determination of oxidant and antioxidant balance. Our results indicated that the percentage and duration of sperm motility significantly decreased with exposure to LCT. Activity of GSH-Px and MDA (P<0.05) and GSH levels (P<0.05) increased in a concentration-dependent manner while CAT activity decreased (P<0.05). In conclusion, the oxidant and antioxidant status and sperm quality were affected by increasing concentrations of LCT. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Localisation and quantification of alkali-labile sites in human spermatozoa by DNA breakage detection-fluorescence in situ hybridisation.

    Science.gov (United States)

    Cortés-Gutiérrez, E I; Dávila-Rodríguez, M I; Cerda-Flores, R M; Fernández, J L; López-Fernández, C; Aragón Tovar, A R; Gosálvez, J

    2015-03-01

    The localisation and quantification of constitutive alkali-labile sites (ALSs) were investigated using a protocol of DNA breakage detection plus fluorescence in situ hybridisation (DBD-FISH) and alkaline single-cell gel electrophoresis (SCGE or comet assay), in spermatozoa of infertile and fertile men. Semen samples from 10 normozoospermic patients undergoing infertility treatment and 10 fertile men were included in this study. ALSs were localised and quantified by DBD-FISH. The region most sensitive to alkali treatment in human spermatozoa was located in the basal region of the head. ALSs were more frequent in spermatozoa of infertile men than in those of fertile men. These results were confirmed by SCGE comet assays. In conclusion, the most intense localisation of hybridisation signals in human spermatozoa, representing the highest density of constitutive ALSs, was not randomly distributed and was predominantly located in the base of the head. Moreover, infertile men presented with an increase in ALS frequency. Further studies are necessary to determine the association between ALS, sperm chromatin organisation and infertility. © 2014 Blackwell Verlag GmbH.

  20. Protein involvement in the fusion between the equatorial segment of acrosome-reacted human spermatozoa and liposomes

    NARCIS (Netherlands)

    Arts, EGJM; Wijchman, JG; Jager, S; Hoekstra, D

    1997-01-01

    Artificial membranes (liposomes) can interact with the equatorial segment (ES) of human spermatozoa, provided that the acrosome reaction (AR) has occurred [Arts, Kuiken, Jager and Hoekstra (1993) fur. J. Biochem. 217, 1001-1009]. Using fluorescently labelled liposomes, this interaction can be seen

  1. Cholesterol loaded cyclodextrin increases freezability of buffalo bull (Bubalus bubalis spermatozoa by increasing cholesterol to phospholipid ratio

    Directory of Open Access Journals (Sweden)

    J. S. Rajoriya

    2014-09-01

    Full Text Available Aim: The study was conducted to investigate the effect of cholesterol loaded cyclodextrin (CLC on freezability of buffalo spermatozoa. Materials and Methods: Murrah buffalo bull semen samples with progressive motility of 70% and greater were used. After the evaluation of motility and livability, four equal fractions of semen samples were made. Group I was kept as control and diluted with Tris, whereas Group II, III and IV were treated with CLC solution at the rate of 2.0, 3.0 and 4.0 mg/ml respectively to obtain 120 × 106 sperm/ml as final spermatozoa concentration. The aliquots of all the groups were incubated for action of CLC, followed by dilution and freezing. Evaluation at pre-freeze and post-thaw stage of progressive motility, viability and level of cholesterol and phospholipid was done. Results: The mean cholesterol content (μg/100 × 106 spermatozoa of Group I, II, III and IV at pre-freeze stage was 21.55±0.63, 49.56±1.38, 55.67±0.45 and 47.79±1.01 and at post-thaw stage were 13.18±0.45, 34.27±0.71, 36.21±0.48 and 33.68±0.56, respectively. At pre-freeze stage, cholesterol content was significantly (p<0.01 higher in Group III in comparison to other groups. The mean cholesterol and phospholipids content of fresh sperm was 24.14±0.58 and 51.13±0.66 μg/100 × 106 sperm cells, respectively, and C/P ratio of spermatozoa at fresh stage was 0.47±0.067. Conclusion: CLC treatment maintains the C/P ratio and plays an important role in maintaining membrane architecture of spermatozoa. Hence, addition of CLC may be helpful in increasing freezability of buffalo spermatozoa by increasing the C/P ratio of spermatozoa.

  2. Eicosapentaenoic acid (EPA) vs. Docosahexaenoic acid (DHA): Effects in epididymal white adipose tissue of mice fed a high-fructose diet.

    Science.gov (United States)

    Bargut, Thereza Cristina Lonzetti; Santos, Larissa Pereira; Machado, Daiana Guimarães Lopes; Aguila, Marcia Barbosa; Mandarim-de-Lacerda, Carlos Alberto

    2017-08-01

    Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been demonstrated to be beneficial for many diseases, including those associated with the metabolic syndrome (e.g. insulin resistance and hypertension). Nevertheless, not only their actions are not entirely understood, but also their only effects were not yet elucidated. Therefore, we aimed to compare the effects of EPA and DHA, alone or in combination, on the epididymal white adipose tissue (WAT) metabolism in mice fed a high-fructose diet. 3-mo-old C57Bl/6 mice were fed a control diet (C) or a high-fructose diet (HFru). After three weeks on the diets, the HFru group was subdivided into four new groups for another five weeks: HFru, HFru+EPA, HFru+DHA, and HFru-EPA+DHA (n=10/group). Besides evaluating biometric and metabolic parameters of the animals, we measured the adipocyte area and performed molecular analyses (inflammation and lipolysis) in the epididymal WAT. The HFru group showed adipocyte hypertrophy, inflammation, and uncontrolled lipolysis. The treated animals showed a reversion of adipocyte hypertrophy, inhibition of inflammation with activation of anti-inflammatory mediators, and regularization of lipolysis. Overall, the beneficial effects were more marked with DHA than EPA. Although the whole-body metabolic effects were similar between EPA and DHA, DHA appeared to be the central actor in WAT metabolism, modulating pro and anti-inflammatory pathways and alleviating adipocytes abnormalities. Therefore, when considering fructose-induced adverse effects in WAT, the most prominent actions were observed with DHA. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Effect modification by apoptosis-related gene polymorphisms on the associations of phthalate exposure with spermatozoa apoptosis and semen quality

    International Nuclear Information System (INIS)

    Yang, Pan; Gong, Ya-Jie; Wang, Yi-Xin; Liang, Xin-Xiu; Liu, Qing; Liu, Chong; Chen, Ying-Jun; Sun, Li; Lu, Wen-Qing

    2017-01-01

    Background: Human studies indicate that phthalate exposure is associated with adverse male reproductive health, and this association may be modified by genetic polymorphisms. Objectives: We investigated whether apoptosis-related gene polymorphisms modified the associations of phthalate exposure with spermatozoa apoptosis and semen quality. Methods: In this Chinese population who sought for semen examination in an infertility clinic, we measured 8 phthalate metabolites in two urine samples to assess the individual's exposure levels. Apoptosis-related gene (Fas, FasL, and caspase3) polymorphisms were performed by real-time PCR. Spermatozoa apoptosis and semen quality parameters were evaluated by Annexin V/PI assay and computer-aided semen analysis, respectively. Results: We found that Fas rs2234767, FasL rs763110, and caspase3 rs12108497 gene polymorphisms significantly modified the associations between urinary phthalate metabolites and spermatozoa apoptosis. For example, urinary monobutyl phthalate (MBP) associated with an increased percentage of Annexin V + /PI − spermatozoa of 25.11% (95% CI: 4.08%, 50.53%) were only observed among men with CT/TT genotype of FasL rs763110. In addition, we found that caspase3 rs12108497 gene polymorphisms significantly modified the associations of urinary mono (2-ethyl-5-hydroxyhexyl) phthalate (MEHHP) with decreased sperm concentration and sperm count (both p-values for interactions = 0.02). Conclusion: Our results provided the first evidence that apoptosis-related gene polymorphisms might contribute to the effects of phthalate exposure on male reproductive health. - Highlights: • We used two urine samples to assess the individual's phthalate exposure levels. • Fas, FasL, and caspase3 variants modified the association between phthalate exposure and spermatozoa apoptosis. • Caspase3 variants modified the association between phthalate exposure and semen quality. • Gene-environment interaction effects should be

  4. Development and evaluation of deep intra-uterine artificial insemination using cryopreserved sexed spermatozoa in bottlenose dolphins (Tursiops truncatus).

    Science.gov (United States)

    Robeck, Todd R; Montano, G A; Steinman, K J; Smolensky, P; Sweeney, J; Osborn, S; O'Brien, J K

    2013-06-01

    Since its development in bottlenose dolphins, widespread application of AI with sex-selected, frozen-thawed (FT) spermatozoa has been limited by the significant expense of the sorting process. Reducing the total number of progressively motile sperm (PMS) required for an AI would reduce the sorting cost. As such, this research compared the efficacy of small-dose deep uterine AI with sexed FT spermatozoa (SEXED-SMALL; ~50×10(6)PMS, n=20), to a moderate dose deposited mid-horn (SEXED-STD, ~200×10(6)PMS; n=20), and a large dose of FT non-sexed spermatozoa deposited in the uterine body (NONSEXED-LARGE, 660×10(6)PMS, n=9). Ten of the 11 calves resulting from use of sexed spermatozoa were of the predetermined sex. Similar rates of conception (NONSEXED-LARGE: 78%, SEXED-STD: 60%, SEXED-SMALL: 57%) and total pregnancy loss (TPL: NONSEXED-LARGE: 28.6%; SEXED-STD: 41.0%; SEXED-SMALL: 63.6%) were observed across groups, but early pregnancy loss (EPL, spermatozoa are achieved after mid-horn deposition of 200×10(6) PMS, when used with females aged less than 25 y. Copyright © 2013 Elsevier B.V. All rights reserved.

  5. Evaluation of intracellular anion superoxide level, heat shock protein A2 and protamine positive spermatozoa percentages in teratoasthenozoospermia

    Directory of Open Access Journals (Sweden)

    Parvin Sabeti

    2017-09-01

    Full Text Available Background: Teratoasthenozoospermia (TA is a severe form of male infertility with no clear etiology. Objective: To compare the level of intracellular anion superoxide (O2–, heat shock protein A2 (HSPA2 and protamine deficiencies in ejaculated spermatozoa between teratoasthenozoospermic and normozoospermic men. Materials and Methods: In this case- control study, semen samples of 20 infertile men, with TA (with normal morphology lower than 4%_ and total motility lower than 40% as the case group and 20 normozoospermic fertile men as the control group were evaluated for intracellular O2 – and HSPA2 by flow cytometry and protamine deficiency by Chromomycin A3 (CMA3 test. Results: The rate of CMA3+ spermatozoa in the case group was higher than controls (p=0.001. The percentages of HSPA2+ spermatozoa in the cases were significantly lower than controls (p=0.001. Also, intracellular O2 – levels in the case group were significantly higher than controls (p=0.001 and had positive correlations with sperm apoptosis (r=0.79, p=0.01 and CMA3 positive sperm (r=0.76, p=0.01, but negative correlations with normal morphology (r=-0.81, p=0.01 and motility (r=-0.81, p=0.01. There was no significant correlation between intracellular O2 – and HSPA2 in the case group (r=0.041, p=0.79. Conclusion: We suggest that the increase in intracellular O2 –, decrease in spermatozoa HSPA2+, and high percentages of spermatozoa with immature chromatin might be considered as etiologies of infertility in TA patients

  6. Impact of supplementation of semen extender with antioxidants on the quality of chilled or cryopreserved Arabian stallion spermatozoa.

    Science.gov (United States)

    Ghallab, AbdelRaouf M; Shahat, Abdallah M; Fadl, Aya M; Ayoub, Mohamed M; Moawad, Adel R

    2017-12-01

    The aim of the present study was to evaluate the effects of supplementation of semen extender with various non-enzymatic antioxidants on the quality of cooled or cryopreserved Arabian stallion spermatozoa. Semen collected from four pure Arabian stallions was centrifuged at 600g for 15 min. Spermatozoa were then diluted in INRA-82 extender supplemented with bovine serum albumin (BSA; 0, 10, 15 and 20 mg/mL) or trehalose (0, 75, 100 and 150 mM) or zinc sulphate (0, 100, 150 and 200 μM). The diluted semen was then either cooled at 5 °C or cryopreserved in 0.5-ml plastic straws. After cooling or thawing, sperm motility, viability, sperm abnormalities, viability index, and plasma membrane integrity were evaluated. The results showed that supplementation of semen extender with 150 mM trehalose or with 200 μM zinc sulphate significantly (P < 0.05) improved motility, viability, sperm membrane integrity and acrosome status in Arabian stallion spermatozoa after cooling or after freezing and thawing compared with controls (non-supplemented media) or with those supplemented with other concentrations of trehalose or zinc sulphate. Supplementation of semen extender with BSA did not improve sperm motility or cryosurvival of Arabian stallion spermatozoa after cooling or after freezing and thawing. In conclusion, supplementation of semen extender with non-enzymatic antioxidants (trehalose or zinc sulphate) improved the quality of chilled and frozen/thawed Arabian stallion spermatozoa. The most beneficial effects occur when semen diluent was supplemented with 150 mM trehalose or 200 μM zinc sulphate. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Comparative characteristics of spermatozoa harvested and cryopreserved in culture and cryoprotectant media with or without donor serum proteins.

    Science.gov (United States)

    Ata'Allah, Ghofraan A; Adenan, Noor Azmi Mat; Razali, Nuguelis; Palaniappan, Kannappan; Saad, Rosliza; Idris, Siti Khadijah; Kanniah, Krishnan; Ali, Jaffar

    2017-06-01

    The objectives of this study is to evaluate the efficacy of protein-free media in the preparation, holding and crypreservation of spermatazoa for use in ART. Normozoospermic semen samples (N=71) were used to compare the effects of media on the survival and quality of spermatozoa when washed and cultured with different media with and without added proteins at 4°C, 15°C, 22°C and 37°C for 0, 4-7 and 24h. Survival and quality of spermatozoa were assessed after freeze-thaw with synthetic cryoprotectant with and without proteins. Ethics/IRB approval was obtained (Ref. 1073.52). Spermatozoa parameters were similar in all media after washing and culture for 24h. Post-thaw survival and quality of spermatozoa was not significantly different 24h after thawing of samples frozen in all cryoprotectant medium. In conclusion synthetic protein-free culture and cryoprotectant media are equal in efficacy to protein-containing media in culture and cryopreservation of spermatozoa . Use of these synthetic media are anticipated to significantly reduce the risk, potentially associated with conventional protein-containing media, of transmission of disease and possibly harmful undeclared proteins to the patient, baby and the healtcare worker. Synthetic media also ensure consistency of quality between batches of media. Copyright © 2017 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  8. Assessment of bovine spermatozoa viability using different cooling protocols prior to cryopreservation

    Directory of Open Access Journals (Sweden)

    Paulo B.D. Gonçalves

    2010-11-01

    Full Text Available The aim of our study was to evaluate the effect of different cooling rates on the post-thawing quality of bovine spermatozoa. Ejaculated semen from a 24-month-old Jersey bull was collected using an artificial vagina and diluted in a commercial extender to evaluate spermatozoan concentration and motility subjectively before cooling and freezing and after thawing. Straws were allocated to four cooling curves: rapid (RD, semi-rapid (SRD, semi-slow (SSLW and slow (SLW. The temperature was decreased from 25ºC to 4ºC in 10, 50, 110 and 135 min, which represents a cooling rate of 2.06, 0.40, 0.18 and 0.15ºC/min, respectively. Then straws were frozen and stored at -196ºC. After thawing, one aliquot of each straw was used for evaluation. Spermatozoan integrity and mitochondrial function were evaluated using a combination of fluorescent probes containing 100 mg/mL FITC-PSA, 0.5 μg/mL PI and 153 μM JC-1. At the end of cooling, spermatozoan motility did not differ among RD (63.3%, SRD (66.7%, SSLW (66.7% and SLW (80.0%. However, normal spermatozoan morphology was lower in SRD (84.8% compared to RD (91.7%, SSLW (91.7% and SLW (90.3% (P<0.05. In thawed semen, spermatozoan motility and normal morphology did not differ among RD (40.0%; 88.8%, SRD (43.3%; 82.5%, SSLW (40.0%; 87.2% and SLW (36.7%; 88.0%. The percentage of damaged spermatozoa, including plasma and acrosome membrane damage and low mitochondrial potential, was higher in RD compared to the others (P<0.05. In conclusion, a rapid cooling curve is detrimental to the spermatozoa and affects the post-thaw spermatozoan integrity of bovine frozen semen.

  9. Milt quality and spermatozoa morphology of captive Brycon siebenthalae (Eigenmann) broodstock

    DEFF Research Database (Denmark)

    Cruz-Casallas, Pablo E.; Lombo-Rodríguez, Dora A.; Velasco-Santamaría, Yohana M.

    2005-01-01

    collected for each male was 1.8 mL and the sperm concentration was 13.99 spermatozoa/mL. Spermatocrit (41.5%) was positively associated (r2: 0.30) with sperm density calculated using a corpuscle counting chamber. Sperm motility was 88% and the average duration of forward motility was 41 s. Fertilization......In order to develop artificial reproduction in freshwater fish for potential species to be developed in South American aquaculture, milt quality and sperm morphology were studied in yamu (Brycon siebenthalae) under captive conditions during the natural middle spermiation period. The volume of milt...... rate was 84% and there was no association between this trait and sperm motility (r2: 0.009) or with sperm density (r2: 0.073). These results suggest that captive B. siebenthalae broodstock can be reproduced successfully....

  10. The ultrastructure of the mullet Mugil curema Valenciennes (Teleostei, Mugilidae spermatozoa

    Directory of Open Access Journals (Sweden)

    Daura Regina Eiras-Stofella

    1993-01-01

    Full Text Available The structure of the spermatozoon of Mugil curema Valenciennes, 1836 was studied using scanning and transmission electron microscopy. The spermatic head is rounded and formed by the nucleus containing granular chromatin, firmly packed resulting in a mass extremely electron dense. The acrossome is absent. The midpiece is characterized by the presence of two centrioles, a plasmatic canal, very few vesicles, and several mitochondria (9-10 with aproximately 0.50µm in diameter. The head and the midpiece are aproximately 1.56µm in diameter. The flagellum conforms to the 9 + 0 flagellar pattern near the transition region in its midpiece and is 9 + 2 from there on up to the distal region of the axoneme. The electron density in the A tubules 1, 2, 5 and 6 shows the asymetry of this spermatozoa. Its spermatic cell differs ultrastructuraly from those of other Mugilidae species mainly because it has the highest number of mitochondria.

  11. Hypoosmotic swelling test in alpaca (Vicugna pacos spermatozoa recovered the vas deferens

    Directory of Open Access Journals (Sweden)

    Quispe HA

    2015-08-01

    Full Text Available The study objective was to evaluate the endosmotic reaction in 5 hypoosmotic solutions 100, 125, 150, 175 and 200 mOsm/l (HOS test incubated 5 and 15 minutes using alpaca sperm recovered the vas deferens. We used 5 male alpacas with the vas deferens surgically diverted, recovering sperm 10 times for each animal; the microscopic characteristics were evaluated with ISAS®. Information’s hipoosmotic test was analyzed with a factorial arrangement in a random one-way model, the correlation between endosmosis and percentages of motility and vitality was analyzed with Spearman correlation. There were no significant differences (p>0,05 in incubation times; highly significant differences (p 0,05was found to endosmosis with percentages of sperm motility and vitality; in conclusion, a 125 mOsm/l solution with 5 minutes of incubation is sufficient to perform the hipoosmotic test in alpaca spermatozoa recovered the vas deferens.

  12. Sperm-attractant peptide influences the spermatozoa swimming behavior in internal fertilization in Octopus vulgaris.

    Science.gov (United States)

    De Lisa, Emilia; Salzano, Anna Maria; Moccia, Francesco; Scaloni, Andrea; Di Cosmo, Anna

    2013-06-15

    Marine invertebrates exhibit both chemokinesis and chemotaxis phenomena, induced in most cases by the release of water-borne peptides or pheromones. In mollusks, several peptides released during egg-laying improve both male attraction and mating. Unlike other cephalopods, Octopus vulgaris adopts an indirect internal fertilization strategy. We here report on the identification and characterization of a chemoattractant peptide isolated from mature eggs of octopus females. Using two-chamber and time-lapse microscopy assays, we demonstrate that this bioactive peptide is able to increase sperm motility and induce chemotaxis by changing the octopus spermatozoa swimming behavior in a dose-dependent manner. We also provide evidence that chemotaxis in the octopus requires the presence of extracellular calcium and membrane protein phophorylation at tyrosine. This study is the first report on a sperm-activating factor in a non-free-spawning marine animal.

  13. COMPUTER-ASSISTED SEMEN ANALYSIS OF RAT SPERMATOZOA AFTER AN INTRAPERITONEAL ADMINISTRATION OF INSECTICIDE DIAZINON

    Directory of Open Access Journals (Sweden)

    R. TOMAN

    2008-05-01

    Full Text Available The aim of this study was to reveal the effect of diazinon on the rat spermatozoa motility characteristics using the computer-assisted semen analysis (CASA. Motility, progressive motility, DAP, DCL, DSL, VAP, VCL, VSL, STR, LIN, WOB, ALH, and BCF after the diazinon i.p. administration of 20 mg/kg b.w. were evaluated. 36 hours after the diazinon administration, only slight decrease in VCL, DCL and increase in percentage of progressive motility in the diazinon-treated group. Significant decrease (P<0.01 was only observed in BCF in diazinon-treated group. Computer-assisted semen analysis (CASA of rat sperm motility showed that acute diazinon administration slightly affected the rat sperm motility which can be the first step in the decreased fertilization capacity caused by pesticides. Further investigation of reproductive effects of diazinon is needed.

  14. CELL DEATH DIFFERENTIATION IN BLACK HEADED RAMS SPERMATOZOA, USING FLUORESCENT LABELED ANNEXIN V

    Directory of Open Access Journals (Sweden)

    M Ivanova-Kicheva

    2008-11-01

    Full Text Available Double staining kit of Annexin V Cy3.18/6-CFDA was used to investigate the changes in phospholipide asymmetry after treating sperm cells with dexamethasone. The % of spermatozoa with registered translocation of PS in treated with dexamethazone groups at the 10-th min and in control no treated varied from 2.74%±0.65 to 2.30%±0.89. After the 5 hour of incubation these % increased to 39.83±3.33 for the treated group and 23.44±1.12 for the control. It was concluded that Annexin V binding assay is more sensitive in the detection of deterioration in membrane function than other conventional methods such as motility analysis and supravital techniques.

  15. Effects of Hormones on the Expression of Matrix Metalloproteinases and Their Inhibitors in Bovine Spermatozoa

    Directory of Open Access Journals (Sweden)

    Sang-Hwan Kim

    2013-03-01

    Full Text Available Proteases and protease inhibitors play key roles in most physiological processes, including cell migration, cell signaling, and cell surface and tissue remodeling. Among these, the matrix metalloproteinase (MMPs pathway is one of the most efficient biosynthetic pathways for controlling the activation of enzymes responsible for protein degradation. This also indicates the association of MMPs with the maturation of spermatozoa. In an attempt to investigate the effect of MMP activation and inhibitors in cultures with various hormones during sperm capacitation, we examined and monitored the localization and expression of MMPs (MMP-2 and MMP-9, tissue inhibitors of metalloproteinases (TIMP-2 and TIMP-3, as well as their expression profiles. Matured spermatozoa were collected from cultures with follicle-stimulating hormone (FSH, luteinizing hormone (LH, and Lutalyse at 1 h, 6 h, 18 h, and 24 h. ELISA detected the expression of MMP-2, MMP-9, TIMP-2, and TIMP-3 in all culture media, regardless of medium type (FSH-supplemented fertilization Brackett-Oliphant medium (FFBO, LH-supplemented FBO (LFBO, or Lutalyse-supplemented FBO (LuFBO. TIMP-2 and TIMP-3 expression patterns decreased in LFBO and LuFBO. MMP-2 and MMP-9 activity in FBO and FFBO progressively increased from 1 h to 24 h but was not detected in LFBO and LuFBO. The localization and expression of TIMP-2 and TIMP-3 in sperm heads was also measured by immunofluorescence analysis. However, MMPs were not detected in the sperm heads. MMP and TIMP expression patterns differed according to the effect of various hormones. These findings suggest that MMPs have a role in sperm viability during capacitation. In conjunction with hormones, MMPs play a role in maintaining capacitation and fertilization by controlling extracellular matrix inhibitors of sperm.

  16. Study of pentoxifylline effects on motility and viability of spermatozoa from infertile asthenozoospermic males.

    Science.gov (United States)

    Ghasemzadeh, Aliye; Karkon-Shayan, Farid; Yousefzadeh, Solmaz; Naghavi-Behzad, Mohammad; Hamdi, Kobra

    2016-01-01

    The quality of semen is one of the major parameters in male infertility. Pentoxifylline, a methylxanthine derivative, is an agent primarily used in the treatment of intermittent claudication and other vascular disorders. Studies have shown that pentoxifylline enhances the quality and quantity of sperms. In this study, we have investigated the in vitro effects of pentoxifylline on viability and motility of spermatozoa in samples of infertile oligoasthenozoospermic males. In this observer-blinded clinical trial, semen samples of 25 infertile oligoasthenozoospermic males were collected in Alzahra Educational Medical Center of Tabriz University of Medical Sciences from August 2010 to August 2012. After the isolation of spermatozoa by the swim-up method, they were randomized into four groups in ISM1 environment: The controls treated normally: Group 1 treated by pentoxifylline at a dose of 50 μg/ml, Group 2 treated by pentoxifylline at a dose of 100 μg/ml, and Group 3 treated by pentoxifylline at a dose of 200 μg/ml. Sperm viability and motility were compared among the groups on 45 min, 24 h, 36 h, and 48 h intervals. Mean percentages of live sperms were 98.40%, 51.40%, 20.60%, and 6.00% in control group and 98.40%, 69.20%, 38.60%, and 14.60% in Group 3 on the mentioned intervals, respectively. This mean percentage decrease of live sperms was significantly lower in Group 3 comparing with that of other groups ( P = 0.01). Mean percentages of motile sperms were 54%, 8.40%, 2.80%, and 0% in control group; and 54%, 16%, 4.80%, and 1.40% in Group 3 on the mentioned intervals, respectively. There was not a significant difference between the four groups in this regard ( P = 0.19). Pentoxifylline can enhance the viability of sperm of infertile oligoasthenozoospermic males with no significant effect on its motility.

  17. DNA profiling of spermatozoa by laser capture microdissection and low volume-PCR.

    Directory of Open Access Journals (Sweden)

    Cai-xia Li

    Full Text Available Genetic profiling of sperm from complex biological mixtures such as sexual assault casework samples requires isolation of a pure sperm population and the ability to analyze low abundant samples. Current standard procedure for sperm isolation includes preferential lysis of epithelial contaminants followed by collection of intact sperm by centrifugation. While effective for samples where sperm are abundant, this method is less effective when samples contain few spermatozoa. Laser capture microdissection (LCM is a proven method for the isolation of cells biological mixtures, even when found in low abundance. Here, we demonstrate the efficacy of LCM coupled with on-chip low volume PCR (LV-PCR for the isolation and genotyping of low abundance sperm samples. Our results indicate that this method can obtain complete profiles (13-16 loci from as few as 15 sperm cells with 80% reproducibility, whereas at least 40 sperm cells are required to profile 13-16 loci by standard 'in-tube' PCR. Further, LCM and LV-PCR of a sexual assault casework sample generated a DNA genotype that was consistent with that of the suspect. This method was unable, however, to analyze a casework sample from a gang rape case in which two or more sperm contributors were in a mixed population. The results indicate that LCM and LV-PCR is sensitive and effective for genotyping sperm from sperm/epithelial cell mixtures when epithelial lysis may be insufficient due to low abundance of sperm; LCM and LV-PCR, however, failed in a casework sample when spermatozoa from multiple donors was present, indicating that further study is necessitated.

  18. High rates of de novo 15q11q13 inversions in human spermatozoa

    Directory of Open Access Journals (Sweden)

    Molina Òscar

    2012-02-01

    Full Text Available Abstract Low-Copy Repeats predispose the 15q11-q13 region to non-allelic homologous recombination. We have already demonstrated that a significant percentage of Prader-Willi syndrome (PWS fathers have an increased susceptibility to generate 15q11q13 deletions in spermatozoa, suggesting the participation of intrachromatid exchanges. This work has been focused on assessing the incidence of de novo 15q11q13 inversions in spermatozoa of control donors and PWS fathers in order to determine the basal rates of inversions and to confirm the intrachromatid mechanism as the main cause of 15q11q13 anomalies. Semen samples from 10 control donors and 16 PWS fathers were processed and analyzed by triple-color FISH. Three differentially labeled BAC-clones were used: one proximal and two distal of the 15q11-q13 region. Signal associations allowed the discrimination between normal and inverted haplotypes, which were confirmed by laser-scanning confocal microscopy. Two types of inversions were detected which correspond to the segments involved in Class I and II PWS deletions. No significant differences were observed in the mean frequencies of inversions between controls and PWS fathers (3.59% ± 0.46 and 9.51% ± 0.87 vs 3.06% ± 0.33 and 10.07% ± 0.74. Individual comparisons showed significant increases of inversions in four PWS fathers (P Results suggest that the incidence of heterozygous inversion carriers in the general population could reach significant values. This situation could have important implications, as they have been described as predisposing haplotypes for genomic disorders. As a whole, results confirm the high instability of the 15q11-q13 region, which is prone to different types of de novo reorganizations by intrachromatid NAHR.

  19. Interacting proteins on human spermatozoa: adaptive evolution of the binding of semenogelin I to EPPIN.

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    Erick J R Silva

    Full Text Available Semenogelin I (SEMG1 is found in human semen coagulum and on the surface of spermatozoa bound to EPPIN. The physiological significance of the SEMG1/EPPIN interaction on the surface of spermatozoa is its capacity to modulate sperm progressive motility. The present study investigates the hypothesis that the interacting surface of SEMG1 and EPPIN co-evolved within the Hominoidea time scale, as a result of adaptive pressures applied by their roles in sperm protection and reproductive fitness. Our results indicate that some amino acid residues of SEMG1 and EPPIN possess a remarkable deficiency of variation among hominoid primates. We observe a distinct residue change unique to humans within the EPPIN sequence containing a SEMG1 interacting surface, namely His92. In addition, Bayes Empirical Bayes analysis for positive selection indicates that the SEMG1 Cys239 residue underwent positive selection in humans, probably as a consequence of its role in increasing the binding affinity of these interacting proteins. We confirm the critical role of Cys239 residue for SEMG1 binding to EPPIN and inhibition of sperm motility by showing that recombinant SEMG1 mutants in which Cys239 residue was changed to glycine, aspartic acid, histidine, serine or arginine have reduced capacity to interact to EPPIN and to inhibit human sperm motility in vitro. In conclusion, our results indicate that EPPIN and SEMG1 rapidly co-evolved in primates due to their critical role in the modulation of sperm motility in the semen coagulum, providing unique insights into the molecular co-evolution of sperm surface interacting proteins.

  20. Improved quality of cryopreserved cheetah (Acinonyx jubatus) spermatozoa after centrifugation through Accudenz.

    Science.gov (United States)

    Crosier, Adrienne E; Henghali, Josephine N; Howard, Jogayle; Pukazhenthi, Budhan S; Terrell, Kimberly A; Marker, Laurie L; Wildt, David E

    2009-01-01

    Sperm cryopreservation, in combination with assisted reproductive techniques, is a valuable tool for the genetic management of endangered felids. However, the acrosome of the cheetah spermatozoon is especially sensitive to cryopreservation, with approximately 40% of spermatozoa experiencing acrosomal damage immediately after thawing and then another approximately 15% loss during the next 4 hours in vitro. Additionally, thawing causes a reduction in sperm motility by approximately 20% with another decrease of approximately 12% during subsequent incubation in vitro. We hypothesized that slow removal of glycerol from cryopreserved cheetah spermatozoa using an Accudenz gradient would improve acrosomal integrity, sperm motility longevity, and structural morphology. Accudenz was compared with traditional cheetah sperm processing methods for glycerol removal that involves washing, multistep resuspension, and swim-up processing. Electroejaculates (n = 21 total from 8 males) were washed in Ham F10 medium, and sperm pellets were resuspended in TEST-yolk buffer with 0% glycerol. Samples were cryopreserved in straws in 4% final glycerol, thawed, and assessed for percent intact acrosomes (% IA), percent motility (% M), and forward progressive status (FPS; scale, 0-5). Sperm motility index (SMI) was calculated as (% M + [FPS x 20]) / 2. In study 1, glycerol removal by centrifugation through an Accudenz gradient (4%, 10%) was compared with traditional sperm washing (control) and multistep resuspension protocols. At each time after centrifugation (hourly for 4 hours), % IA was improved (P cheetah sperm mitigates the significant loss in sperm quality that occurs after freeze-thawing. This alleviation of cellular damage resulting from cryopreservation contributes to a more than 10% improvement in overall sperm motility and, more importantly, allows retention of 40% or more of sperm with intact acrosomes.

  1. Is prnt a pseudogene? Identification of ram Prt in testis and ejaculated spermatozoa.

    Directory of Open Access Journals (Sweden)

    Jorge Pimenta

    Full Text Available A hallmark of prion diseases or transmissible spongiform encephalopaties is the conversion of the cellular prion protein (PrP(C, expressed by the prion gene (prnp, into an abnormally folded isoform (PrP(Sc with amyloid-like features that causes scrapie in sheep among other diseases. prnp together with prnd (which encodes a prion-like protein designated as Doppel, and prnt (that encodes the prion protein testis specific--Prt with sprn (shadow of prion protein gene, that encodes Shadoo or Sho genes, constitute the "prion gene complex". Whereas a role for prnd in the proper functioning of male reproductive system has been confirmed, the function of prnt, a recently discovered prion family gene, comprises a conundrum leading to the assumption that ruminant prnt is a pseudogene with no protein expression. The main objective of the present study was to identify Prt localization in the ram reproductive system and simultaneously to elucidate if ovine prnt gene is transcribed into protein-coding RNA. Moreover, as Prt is a prnp-related protein, the amyloid propensity was also tested for ovine and caprine Prt. Recombinant Prt was used to immunize BALB/c mice, and the anti-Prt polyclonal antibody (APPA immune response was evaluated by ELISA and Western Blot. When tested by indirect immunofluorescence, APPA showed high avidity to the ram sperm head apical ridge subdomain, before and after induced capacitation, but did not show the same behavior against goat spermatozoa, suggesting high antibody specificity against ovine-Prt. Prt was also found in the testis when assayed by immunohistochemistry during ram spermatogenesis, where spermatogonia, spermatocytes, spermatids and spermatozoa, stained positive. These observations strongly suggest ovine prnt to be a translated protein-coding gene, pointing to a role for Prt protein in the ram reproductive physiology. Besides, caprine Prt appears to exhibit a higher amyloid propensity than ovine Prt, mostly associated

  2. Spermatozoa motility and short-term sperm storage of colourful orfe (Leuciscus idus aberr orfus

    Directory of Open Access Journals (Sweden)

    Beata I. Cejko

    2012-07-01

    Full Text Available In this study the effect of six activating buffers on the movement parameters of sperm was determined and short-term storage of semen in TLP buffer was attempted (0.292g NaCl; 0.012g KCl; 0.011g CaCl2; 0.004g MgCl2; 0.105g NaHCO3; 0.002g NaH2PO4; 50ml; pH 8.6. Sperm was collected from five orfe individual, and spermiation was stimulated by means of an intraperitoneal Ovopel injection. The basic parameters of spermatozoa motility were measured after the semen was diluted with six different activating solutions, previously used successfully in other fish species. The motility analysis was conducted on a Crismas apparatus. Additionally, short-term storage of semen in TLP buffer was attempted. Subsequently, motility parameters were verified after 0 (Control, 24 and 120 h of storage at 4°C. It has been found that Lahnsteiner’s buffer (100 mM NaCl, 10 mM Tris, 0.5% albumin, 199 mOsmkg-1 was found to be the most effective in sperm activation. In this paper, the spermatozoa motility of colourful ide is indicated for the first time. Finally, there was a successful attempt at short-term sperm storage for five days. For artificial insemination, it is very important to select the most effective solution to stimulate sperm motility. Data regarding sperm manipulation of orfe are scarce, so the aim of the study was to determine the basic sperm quality parameters of the colour ide form, i.e. Leuciscus idus aberr orfus.

  3. Protective features of resveratrol on human spermatozoa cryopreservation may be mediated through 5' AMP-activated protein kinase activation.

    Science.gov (United States)

    Shabani Nashtaei, M; Amidi, F; Sedighi Gilani, M A; Aleyasin, A; Bakhshalizadeh, Sh; Naji, M; Nekoonam, S

    2017-03-01

    Biochemical and physical modifications during the freeze-thaw process adversely influence the restoration of energy-dependent sperm functions required for fertilization. Resveratrol, a phytoalexin, has been introduced to activate 5' AMP-activated protein kinase which is a cell energy sensor and a cell metabolism regulator. The cryoprotection of resveratrol on sperm cryoinjury via activation of AMP-activated protein kinase also remains to be elucidated. Our aim, thus, was to investigate: (i) the presence and intracellular localization of AMP-activated protein kinase protein; (ii) whether resveratrol may exert a protective effect on certain functional properties of fresh and post-thaw human spermatozoa through modulation of AMP-activated protein kinase. Spermatozoa from normozoospermic men were incubated with or without different concentrations of Compound C as an AMP-activated protein kinase inhibitor or resveratrol as an AMP-activated protein kinase activator for different lengths of time and were then cryopreserved. AMP-activated protein kinase is expressed essentially in the entire flagellum and the post-equatorial region. Viability of fresh spermatozoa was not significantly affected by the presence of Compound C or resveratrol. However, although Compound C caused a potent inhibition of spermatozoa motility parameters, resveratrol did not induce negative effect, except a significant reduction in motility at 25 μm for 1 h. Furthermore, resveratrol significantly increased AMP-activated protein kinase phosphorylation and mitochondrial membrane potential and decreased reactive oxygen species and apoptosis-like changes in frozen-thawed spermatozoa. Nevertheless, it was not able to compensate decreased sperm viability and motility parameters following cryopreservation. In contrast, Compound C showed opposite effects to resveratrol on AMP-activated protein kinase phosphorylation, reactive oxygen species, apoptosis-like changes, mitochondrial membrane potential, and

  4. Use of boar spermatozoa for artificial insemination. III. Fecundity of boar spermatozoa stored in Beltsville liquid and Kiev extenders for three days at 18 C.

    Science.gov (United States)

    Johnson, L A; Aalbers, J G; Willems, C M; Rademaker, J H; Rexroad, C E

    1982-01-01

    A field trial was conducted on several hundred farms in The Netherlands to compare the fertilizing capacity of boar spermatozoa stored for 1, 2, or 3 d at 18 C in either Kiev or Beltsville liquid extender (BL-1). Two storage volumes (25 and 100 ml) were used. Twelve Dutch Landrace boars, all in routine commercial artificial insemination service, provided the semen in twice-a-week collections. Over an 8-wk period in March and April, 1979, 2,849 sows and gilts were inseminated. Farrowing rates were higher (P less than .0001) with Kiev extender than with BL-1 extender (69.3 vs 60.5%) when semen was used over a 3 d period. The volume at which the semen was extended and stored had no effect on any parameter. Farrowing rate declined significantly when semen stored at 18 C for more than 2 d was used (74.5% vs 64.7% with semen stored for 1 and 3 in Kiev extender; 65.9% vs 52.7% with semen stored for 1 and 3 d in BL-1 extender). The farrowing rate, total number of pigs/litter and live pigs/litter were significantly higher for sows than for gilts inseminated with similar extended semen. On the basis of the results of this trial, Kiev extender appears to be more suitable as a diluent for liquid semen than does BL-1. AI with semen extended and stored for 3 d at 18 C can be expected to result in a lower farrowing rate than AI with semen stored for 1 or 2 d.

  5. The test about blood serum capabilities in maintaining the quality of bull spermatozoa during storage in cep diluent at refrigerator temperature

    Science.gov (United States)

    Ducha, Nur

    2018-03-01

    The storage of spermatozoa requires a protective material from cold shock events and the presence of free radicals.In CEP diluent contain BSA, that was used as spermatozoa protection. This study aim was to examine the ability of cow blood serum in replacing BSA as spermatozoa protective in CEP diluent. Fresh semen from Limousin bull was diluted with CEP diluent + BSA as control, in the treatment group were CEP without BSA, but replaced with 3%, 5%, and 7% serum from fresh blood. Spermatozoa quality tests included motility and viability. The motility of spermatozoa was observed by two people using a light microscope with 200 X magnification at temperature of 37°C. The method of viability observation was eosin nigrosin staining, and observed under a light microscope with 400 X magnification. The results showed that the replacement of cow blood serum with various concentrations gave different effects on the quality of spermatozoa. The best motility and viability of the treatment group was at serum concentrations of 5% after eight days storage and was not significantly different from the controls. The conclusion in this study was cow blood serum can replace BSA in CEP diluents.

  6. Ultrastructural characters of the spermatozoa in Digeneans of the genus Lecithochirium Lühe, 1901 (Digenea, Hemiuridae, parasites of fishes: comparative study of L. microstomum and L. musculus

    Directory of Open Access Journals (Sweden)

    Ndiaye Papa Ibnou

    2014-01-01

    Full Text Available This study provides the first ultrastructural data of spermatozoa in the genus Lecithochirium. The spermatozoa of L. microstomum (from Trichiurus lepturus in Senegal and L. musculus (from Anguilla anguilla in Corsica exhibit the general pattern described in the great majority of the Digenea, namely two axonemes with the 9 + “1” pattern typical of the Trepaxonemata, one mitochondrion, a nucleus, parallel cortical microtubules and external ornamentation of the plasma membrane. Spermatozoa of L. microstomum and L. musculus have some specific features such as the presence of a reduced number of cortical microtubules arranged on only one side of the spermatozoon, the lack of spine-like bodies and expansion of the plasma membrane. The external ornamentation of the plasma membrane entirely covers the anterior extremity of the spermatozoa. The ultrastructure of the posterior extremity of the spermatozoa corresponds to the pattern previously described in the Hemiuridae, characterized by only singlets of the second axoneme. A particularity of these spermatozoa is the organization of the microtubule doublets of the second axoneme around the nucleus in the posterior part of the spermatozoon.

  7. PENGARUH SUBSTITUSI AIR KELAPA MUDA DENGAN PENGENCER SITRAT KUNING TELUR TERHADAP MOTILITAS DAN PERSENTASE HIDUP SPERMATOZOA ANJING

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    I Nyoman Sulabda

    2012-11-01

    Full Text Available The purpose of this study was to determine the effect of tender coconut water substitutionon egg yolk citrate diluent with different doses on local breeds dog sperm motility and livespermatozoa. Semen was manually collected. Progressive motility and percentage of livespermatozoa were evaluated under a microscope utilizing a drop of semen between awarmed glass slide and coverslip, both at a temperature of 38 0C. The percentage of motileand live spermatozoa were examined by counting 100 spermatozoa using the classificationof Christiansen (1984. Sperm viability was assessed by eosinnegrosin staining. The result showed that coconut water substitution has significant effect on the motility and livespermatozoa . Combination between the levels of coconut water in the egg yolk citratediluent could be applied as an alternative diligent instead of egg yolk diligent for dogsemen up to 75%.

  8. Viable acrosome-intact human spermatozoa in the ejaculate as a marker of semen quality and fertility status

    DEFF Research Database (Denmark)

    Egeberg Palme, Dorte Louise; Rehfeld, Anders; Bang, Anne Kirstine

    2018-01-01

    STUDY QUESTION: Is it possible, in an unbiased and clinical relevant way, to determine the number of viable acrosome-intact human spermatozoa in ejaculates and to use this as a measure of fertility chances? SUMMARY ANSWER: Image cytometry enables easy and unbiased quantification of viable acrosome......-intact spermatozoa and it correlates with semen quality and fertility status. WHAT IS KNOWN ALREADY: The presence of the acrosome and its ability to respond to physiological inducers (e.g. progesterone) in the female reproductive tract at the appropriate time and place is required for fertilization. However......, the available assays are labor intensive and therefore not used clinically. STUDY DESIGN, SIZE, DURATION: Washed semen samples and capacitated swim-up fractions from volunteers were used to develop the assay. Subsequently washed ejaculates from patients in fertility treatment (n = 156), proven fertile men (n...

  9. Spectra of molecular changes induced in DNA of Drosophila spermatozoa by 1-ethyl-1-nitrosourea and X-rays

    International Nuclear Information System (INIS)

    Batzer, M.A.; Fossett, N.G.; Lee, W.R.; Louisiana State Univ., Baton Rouge; Tedeschi, B.; Tucker, A.; Kilroy, G.; Arbour, P.

    1988-01-01

    Mutations induced in Drosophila spermatozoa at the alcohol dehydrogenase Adh locus by 1-ethyl-1-nitrosourea (ENU) were compared to X-ray-induced mutations using genetic tests for complementation, southern blotting, western blotting and northern blotting. 8 of 10 ENU-induced mutations complemented all known adjacent loci and were presumed to be intragenic. In contrast, 8 of 30 X-ray-induced mutations were intragenic. The interpretation of these results is that in spermatozoa X-rays induce primarily deletions that either produce deficiencies of the Adh locus or nonsense mutations within the locus, whereas ENU induces primarily missense mutations. This forward mutation assay based on loss of enzymatic activity efficiently recovered a broad spectrum of mutations ranging from missense to intragenic deletions and multi-locus deficiencies. Only 3 of these 40 mutations produced a polypeptide detectable as an electrophoretic variant. 37 refs.; 6 figs.; 2 tabs

  10. Epididymal sperm from Spix's yellow-toothed cavies sperm successfully cryopreserved in Tris extender with 6% glycerol and 20% egg yolk.

    Science.gov (United States)

    Silva, Andréia M; Praxedes, Erica C G; Campos, Lívia B; Bezerra, Luana G P; Moreira, Samara S J; Maia, Keilla M; Souza, Ana L P; Silva, Alexandre R

    2018-04-01

    As a non-threatened hystricognath rodent species, Spix's yellow-toothed cavies can be used as a model for the development of assisted reproductive techniques for the conservation of closely related species. The objective was to establish a functional protocol for cryopreservation of epididymal sperm from these cavies. Twelve sexually mature males, ∼2 y old and weighing ∼300 g, were euthanized. Sperm were recovered by retrograde flushing of the vas deferens and cauda epididymis with Tris extender. Thereafter, sperm were extended in Tris plus 20% egg yolk, with 3%, 6% or 9% glycerol or dimethyl sulfoxide (DMSO), placed in 0.25 mL straws and cryopreserved in liquid nitrogen. Sperm concentration, motility (using computer-assisted sperm analysis; CASA), plasma membrane integrity, osmotic response, morphology and sperm binding-ability were determined in fresh and frozen-thawed sperm. For most sperm endpoints, glycerol was a more desirable cryoprotectant than DMSO. Data (mean ± SEM) were similar with use of 3%, 6%, and 9% glycerol (P > 0.05) in osmotic response (40.66 ± 6.3%, 42.5 ± 7.1%, and 39.5 ± 5.0% respectably), and membrane integrity (55.17 ± 5.5%, 68.4 ± 4.1%, and 59.1 ± 4.9% respectably). Among concentrations assessed, the use of 6% glycerol resulted in the greatest (P < 0.05) post-thaw values for total motility (60.9 ± 4.4%), rapid subpopulation motility (27.7 ± 3.1%) and sperm-binding capability (227.0 ± 20.2). In conclusion, epididymal sperm from the Spix's yellow-toothed cavies (G. spixii) are optimally cryopreserved in Tris extender with 6% glycerol and 20% egg yolk. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. Comparative proteome analysis of cryopreserved flagella and head plasma membrane proteins from sea bream spermatozoa: effect of antifreeze proteins.

    Science.gov (United States)

    Zilli, Loredana; Beirão, José; Schiavone, Roberta; Herraez, Maria Paz; Gnoni, Antonio; Vilella, Sebastiano

    2014-01-01

    Cryopreservation induces injuries to fish spermatozoa that in turn affect sperm quality in terms of fertilization ability, motility, DNA and protein integrity and larval survival. To reduce the loss of sperm quality due to freezing-thawing, it is necessary to improve these procedures. In the present study we investigated the ability of two antifreeze proteins (AFPI and AFPIII) to reduce the loss of quality of sea bream spermatozoa due to cryopreservation. To do so, we compared viability, motility, straight-line velocity and curvilinear velocity of fresh and (AFPs)-cryopreserved spermatozoa. AFPIII addition to cryopreservation medium improved viability, motility and straight-line velocity with respect to DMSO or DMSO plus AFPI. To clarify the molecular mechanism(s) underlying these findings, the protein profile of two different cryopreserved sperm domains, flagella and head plasma membranes, was analysed. The protein profiles differed between fresh and frozen-thawed semen and results of the image analysis demonstrated that, after cryopreservation, out of 270 proteins 12 were decreased and 7 were increased in isolated flagella, and out of 150 proteins 6 showed a significant decrease and 4 showed a significant increase in head membranes. Mass spectrometry analysis identified 6 proteins (4 from isolated flagella and 2 present both in flagella and head plasma membranes) within the protein spots affected by the freezing-thawing procedure. 3 out of 4 proteins from isolated flagella were involved in the sperm bioenergetic system. Our results indicate that the ability of AFPIII to protect sea bream sperm quality can be, at least in part, ascribed to reducing changes in the sperm protein profile occurring during the freezing-thawing procedure. Our results clearly demonstrated that AFPIII addition to cryopreservation medium improved the protection against freezing respect to DMSO or DMSO plus AFPI. In addition we propose specific proteins of spermatozoa as markers related to

  12. Mitochondrial functions in spermatozoa of minipig boars carrying transgene with the N-terminal part of human mutated huntingtin

    Czech Academy of Sciences Publication Activity Database

    Spáčilová, J.; Rodinová, M.; Kratochvílová, H.; Ondrušková, N.; Mačáková, Monika; Bohuslavová, Božena; Ellederová, Zdeňka; Zeman, J.; Motlík, Jan; Hansíková, H.

    2015-01-01

    Roč. 78, Suppl 2 (2015), s. 24-24 ISSN 1210-7859. [Conference on Animal Models for neurodegenerative Diseases /3./. 08.11.2015-10.11.2015, Liblice] R&D Projects: GA MŠk ED2.1.00/03.0124; GA MŠk(CZ) 7F14308 Institutional support: RVO:67985904 Keywords : Huntington ´s disease * large animal model * spermatozoa Subject RIV: FH - Neurology

  13. Effect modification by apoptosis-related gene polymorphisms on the associations of phthalate exposure with spermatozoa apoptosis and semen quality.

    Science.gov (United States)

    Yang, Pan; Gong, Ya-Jie; Wang, Yi-Xin; Liang, Xin-Xiu; Liu, Qing; Liu, Chong; Chen, Ying-Jun; Sun, Li; Lu, Wen-Qing; Zeng, Qiang

    2017-12-01

    Human studies indicate that phthalate exposure is associated with adverse male reproductive health, and this association may be modified by genetic polymorphisms. We investigated whether apoptosis-related gene polymorphisms modified the associations of phthalate exposure with spermatozoa apoptosis and semen quality. In this Chinese population who sought for semen examination in an infertility clinic, we measured 8 phthalate metabolites in two urine samples to assess the individual's exposure levels. Apoptosis-related gene (Fas, FasL, and caspase3) polymorphisms were performed by real-time PCR. Spermatozoa apoptosis and semen quality parameters were evaluated by Annexin V/PI assay and computer-aided semen analysis, respectively. We found that Fas rs2234767, FasL rs763110, and caspase3 rs12108497 gene polymorphisms significantly modified the associations between urinary phthalate metabolites and spermatozoa apoptosis. For example, urinary monobutyl phthalate (MBP) associated with an increased percentage of Annexin V + /PI - spermatozoa of 25.11% (95% CI: 4.08%, 50.53%) were only observed among men with CT/TT genotype of FasL rs763110. In addition, we found that caspase3 rs12108497 gene polymorphisms significantly modified the associations of urinary mono (2-ethyl-5-hydroxyhexyl) phthalate (MEHHP) with decreased sperm concentration and sperm count (both p-values for interactions = 0.02). Our results provided the first evidence that apoptosis-related gene polymorphisms might contribute to the effects of phthalate exposure on male reproductive health. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. No cytotoxic effects from application of pentoxifylline to spermatozoa on subsequent pre-implantation embryo development in mice

    Directory of Open Access Journals (Sweden)

    Mohammad Ali Khalili

    2017-06-01

    Full Text Available The aim was to assess the effect of spermatozoa exposed to PTX on the rates of fertilization and embryo development and apoptotic cells within blastocysts in an animal model. Mice Oocytes were inseminated with spermatozoa exposed to 3.6 mmol PTX for 30 min, or with neat spermatozoa. Then fertilization and embryo development rate, blastocyst formation and quality, as well as total cell number of blastocyst, and DNA fragmentation index (DFI in blastocysts were surveyed in both groups. Fertilization and embryo development rate were similar between the groups. The rates of blastocyst formation did not differ significantly between control and PTX groups (52.4% vs. 51.8%. The average of total cell count in blastocysts and DFI in control and PTX groups were also insignificant (31.08 ± 1.5 vs. 34.14 ± 1.5 and 9.76 ± 5.0 vs. 11.77 ± 5.4. Application of PTX for enhancing sperm motility does not cause a cytotoxic effect on subsequent embryo development and embryo genome integrity.

  15. Effects of multiple collections on spermatozoa quality of Persian sturgeon, Acipenser persicus: motility, density and seminal plasma composition.

    Science.gov (United States)

    Aramli, M S; Kalbassi, M R; Gharibi, M R

    2015-02-01

    In this study, we investigated the effects of multiple collections of sperm on the endangered Persian sturgeon, Acipenser persicus, in terms of a number of sperm functional parameters (percentage of motile spermatozoa, total time period of motility and sperm concentration) as well as on the ionic composition, protein concentration and osmolality of seminal plasma. Semen samples were collected from 12 induced male fish in three experimental groups that had been injected intramuscularly with LHRH-A2, at dosages of 5 μg/kg body weight, at a number of time regimes: at 12 h, 17 h and 24 h after spawning induction (1); at 24, 29 and 34 h after spawning induction (2); and at 36, 41 and 46 h after spawning induction (3). The percentage of motile spermatozoa and the period of sperm motility decreased significantly (p collections. The concentration of spermatozoa decreased after the third collection, but this decline was not significant. No significant effect of multiple collections on protein concentration and ionic content (with exception of the Cl(-) ion) of seminal plasma was observed. In all experimental groups, a moderate impact of sequential collection on the osmolality (p collections on spermatological characteristics in the Persian sturgeon. Our results confirm that sequential stripping after the third collections has a negative effect on a number of functional parameters associated with sperm. Journal of Animal Physiology and Animal Nutrition © 2014 Blackwell Verlag GmbH.

  16. Cryopreservation of Boer goat spermatozoa: Comparison of two freezing extenders based on post-thaw sperm quality and fertility rates

    Directory of Open Access Journals (Sweden)

    Fitra Aji Pamungkas

    2014-06-01

    Full Text Available Boer goat have recently been popularly used for cross breeding with local goats. However, it is currently considered a breed at very limited number with relatively high prices . In this context, the cryopreservation of spermatozoa is important because it could be conserved for a very long period of time. Egg yolk extenders are most commonly used for cryopreservation of goat sperm. The aim of this study was to compare the ability of two extenders to maintain sperm viability after cryopreservation. Semen from three male Boer goat aged about 2-3 years old was collected using artificial vagina and frozen with Tris and Triladyl extender. The results showed that percentage of motility, viability and membrane integrity of spermatozoa with Tris and Triladyl extenders at every stage of cryopreservation showed not significantly difference (P>0.05, except the percentage of sperm motility post thawing of Triladyl was higher than Tris extender (52.00±4.47% vs 47.50±2.74%, P<0.05. Cryopreserved semen in Tris extender provided the same fertility rates after cervical insemination compared to Triladyl (62.50% vs 60.00%. In conclusion, the Tris extender has the same capabilities to Triladyl in cryopreservation of Boer goat spermatozoa as to maintain sperm quality and fertility rates.

  17. Variations in creatine kinase activity and reactive oxygen species levels are involved in capacitation of bovine spermatozoa.

    Science.gov (United States)

    Córdoba, M; Pintos, L; Beconi, M T

    2008-12-01

    The generation of reactive oxygen species (ROS) is associated with some factors such as oxidative substrate sources, mitochondrial function and NAD(P)H oxidase activity. In bovine spermatozoa, heparin capacitation produces a respiratory burst sensitive to diphenyleneiodonium (DPI). Creatine kinase (CK) is related to extramitochondrial ATP disponibility. Our purpose was to determine the variation in ROS level and its relation with NAD(P)H oxidase sensitive to DPI and CK participation, as factors involved in redox state and energy generation in capacitation. The chlortetracycline technique was used to evaluate capacitation. CK activity and ROS level were measured by spectrophotometry and spectrofluorometry respectively. The capacitation percentage was increased by heparin or quercetin treatment (P level as control (238.62 +/- 23.47 arbitrary units per 10(8) spermatozoa) (P > 0.05). CK activity decreased by 50% with heparin or quercetin (P level variations were observed in heparin- or quercetin-treated samples (P bovine spermatozoa, capacitation requires equilibrium between oxidative damage susceptibility and ROS levels. CK activity is associated with redox state variation and energy sources. In conclusion, capacitation induction depends on NADPH oxidase and the shuttle creatine-creatine phosphate, both sensitive to DPI.

  18. Nascimento de bezerros normais após inseminação artificial utilizando espermatozóides criopreservados obtidos de epidídimos refrigerados de bovinos após a morte Birth of normal calves after artificial insemination using cryopreserved spermatozoa obtained from refrigerated epididymides of death bovine

    Directory of Open Access Journals (Sweden)

    Priscila de Melo Costa

    2011-05-01

    epidydimides for long periods and cryopreserved. Bovine testicles were collected in abattoir, transported to the laboratory and stored at 5°C for 0, 24, 48h e 72 hours (n=10 for each storage time treatment group. The spermatozoa were retrieved from each epidydimides, evaluated and diluted in tris-egg yolk-glycerol 7% medium and cryopreserved in liquid nitrogen. The morphological and functional characteristics of the spermatozoa were analyzed in vitro, by microscopic evaluation and in vivo, using artificial insemination. Morphological alterations as sperm immaturity and motility reduction decreased after 72h of epididymides refrigeration and after thaw sperm were observed. The membrane and acrosome integrity were only affected in G48 and G72 groups after cryopreservation. However, the sperm capacity of fertilization post-cryopreservation was sufficient to promote two pregnancies and birth of healthy calves from G24 h and G72h groups. These results indicated that recovery and cryopreservation of chilled epididymal sperm until 72h from dead animals is a viable option to preserve male gametes to compose a germplasm bank.

  19. Cholesterol addition aids the cryopreservation of dromedary camel (Camelus dromedarius) spermatozoa.

    Science.gov (United States)

    Crichton, Elizabeth G; Pukazhenthi, Budhan S; Billah, M; Skidmore, Julian A

    2015-01-15

    The cryopreservation of dromedary camel (Camelus dromedarius) sperm has proved challenging with little success reported. The routine application of artificial insemination with frozen semen would assist the flow of valuable genetic material nationally and internationally. The current study sought to examine the effects of cholesterol (cholesterol-loaded cyclodextrin [CLC]) preloading on camel sperm cryosurvival. Ejaculates (n = 3 males; 3 ejaculates per male) were collected using an artificial vagina during the breeding season and extended in HEPES-buffered Tyrode's albumin lactate pyruvate (TALP) and allowed to liquefy in the presence of papain (0.1 mg/mL) before removal of the seminal plasma by centrifugation. Sperm pellets were resuspended (120 million/mL) in fresh TALP and incubated (15 minutes; 37 °C) with 0, 1.5, or 4.5 mg CLC/mL. Sperm suspensions were then centrifuged and reconstituted in INRA-96 containing 20% (v:v) egg yolk and 2.5% (v:v) methylformamide, loaded in 0.5-mL plastic straws, sealed, and cooled for 20 minutes at 4 °C. Straws were frozen over liquid nitrogen (4 cm above liquid; 15 minutes), plunged, and stored. Sperm motility, forward progressive status, and acrosomal integrity were recorded at 0 and 3 hours after thawing and compared with these same parameters before freezing. Aliquots also were stained with chlortetracycline hydrochloride to assess spontaneous sperm capacitation status before freezing and post-thaw. Pretreatment with CLC (1.5 and 4.5 mg/mL) enhanced cryosurvival. Post-thaw sperm motility was highest (P < 0.05) in 1.5 mg CLC/mL immediately after thawing (44%) and after 3 hours incubation at room temperature (34%). Highest post-thaw sperm progressive status was also achieved in the presence of 1.5 CLC. Greater proportions of spermatozoa retained acrosomal membrane integrity when cryopreserved in the presence of CLC, but there was no difference between 1.5 and 4.5 CLC. Although thawed spermatozoa underwent spontaneous

  20. The effects of cholesterol on the viability and fertility of bull spermatozoa

    Directory of Open Access Journals (Sweden)

    Polmer Situmorang

    2002-12-01

    Full Text Available This study was conducted to evaluate the effect of cholesterol on the viability and fertility of chilled and deep-frozen bull spermatozoa. Semen was collected by means of artificial vagina, diluted in Tris-Citrat diluent and cooled to 50C for 60 minutes. Following an equiliberation for 4 hours, semen was frozen at 5 cm above surface of liquid nitrogen for 10 minutes. The experiment was 2 x 3 factorial designed with two level of egg yolk (10 and 20% v/v and 3 level of cholesterol (0; 0.5 and 1.0 mg/ml. The viability of spermatozoa was evaluated after the temperature reduced to 50C, stored at 50C for 1, 3 and 7 days and after thawing. For fertility test, cows were artificially inseminated (AI using chilled and frozen semen on the onset or 6 hours of oestrus. Rectal palpation was conducted 3 months after AI to determine the pregnancy. The percentages motile of chilled semen was higher in 0.5 mg/ml than those of 0.0 or 1.0 mg/ml cholesterol but this difference was not significant. After thawing, the effects of cholesterol on the percentage motile was significant (P<0.05. The mean percentage motile was 47.5; 51,5 and 56.0 for 0.0; 0.5 and 1.0 mg/ml cholesterol respectively. The percentage of live sperm and intact apical ridge was higher in cholesterol however this effects was not significant. The effects level of egg yolk and its interaction with cholesterol on the viability was not significant. The percentage of pregnant was higher in 1.0 mg/ml and the mean percentage of pregnant was 45.8; 48.2 and 55.7 for 0.0; 0.5 and 1.0 mg/ml cholesterol respectively. Percentage of pregnant was higher for chilled semen than those of frozen semen (54.3 vs 45.5. In conclusion the addition of 1 mg/ml cholesterol increase the percentage of motile after thawing and pregnancy of cows inseminated with chilled and frozen semen.

  1. pKI values of prazosin and idazoxan for receptors stimulated by neuronally released transmitter in the epididymal portion of rat isolated vas deferens.

    Science.gov (United States)

    Mackay, D; Kengatharan, M

    1994-01-01

    1. A new method has been used to measure pKI values of prazosin and idazoxan against neuronally-released transmitter in the epididymal portion of the rat isolated vas deferens. The most reproducible results were obtained with a prolonged antagonist equilibration time (1 h). 2. Under these conditions the pKI of prazosin was practically unaffected by addition of alpha, beta-methylene-adenosine-5'-triphosphate (10 microM) to desensitize purinoceptors. Addition of desmethylimipramine (DMI) (0.3 microM) produced a small, but statistically non-significant, reduction. 3. The same method has been used to measure the pKI of prazosin against exogenous noradrenaline. In the latter case addition of DMI (0.3 microM) and corticosterone (30 microM) together produced a statistically significant reduction in the apparent pKI of prazosin. 4. The new method for estimating pKI values shows that DMI itself acts either pseudo-irreversibly or non-competitively and may be reducing the apparent pKI of prazosin. 5. The pKI values obtained for prazosin and idazoxan against neuronally-released transmitter are in good agreement with those obtained by other workers for the actions of these drugs on alpha-adrenoceptors.

  2. Detection of lipid peroxidation in frozen-thawed avian spermatozoa using C(11)-BODIPY(581/591).

    Science.gov (United States)

    Partyka, Agnieszka; Lukaszewicz, Ewa; Niżański, Wojciech; Twardoń, Jan

    2011-06-01

    The aim of this study was to perform flow cytometric analysis of C11-BODIPY581/591 oxidation in fowl and geese sperm as a marker for membrane lipid peroxidation (LPO) and to establish if the cryopreservation process would make sperm membranes more susceptible to oxidative stress. The experiment was carried out on 10 meat type line Flex roosters and 10 White Koluda® geese. The semen was collected two times a week, by dorso-abdominal massage method and pooled from 10 individuals of each species. Fowl semen samples were subjected to cryopreservation using the "pellet" method and Dimethylacetamide (DMA) as a cryoprotectant. Geese semen samples were cryopreserved in plastic straws in a programmable freezing unit with Dimethyloformamide (DMF) as the cryoprotectant. A fluorescent lipid probe C11-BODIPY581/591 provided with two double bonds that are oxidized during their contact with ROS, was used for the purpose of the assessment of the LPO in freshly diluted semen samples and frozen-thawed semen samples. This probe changes its color according to its state (non peroxidized: red; peroxidized: green). Flow cytometric analysis was used to monitor these changes. The White Koluda® geese fresh semen had a higher level of LPO than the Flex fresh semen (P > 0.01). The cryopreservation of fowl semen significantly (P > 0.01) increased the percentage of live and dead spermatozoa with lipid peroxidation. In frozen-thawed semen of White Koluda® geese the percentage of live spermatozoa with LPO significantly decreased (P > 0.05) whereas significantly (P > 0.01) higher level of dead cells with LPO was observed. There were significant differences between the two studied species. After thawing, the percentage of live and dead spermatozoa with lipid peroxidation was higher in fowl semen than in geese semen (P > 0.01). In conclusion, our data clearly indicate the existence of species specific differences in susceptibility of spermatozoa to the oxidation of PUFAs in the cell membranes

  3. Processing of semen by density gradient centrifugation selects spermatozoa with longer telomeres for assisted reproduction techniques.

    Science.gov (United States)

    Yang, Qingling; Zhang, Nan; Zhao, Feifei; Zhao, Wanli; Dai, Shanjun; Liu, Jinhao; Bukhari, Ihtisham; Xin, Hang; Niu, Wenbing; Sun, Yingpu

    2015-07-01

    The ends of eukaryotic chromosomes contain specialized chromatin structures called telomeres, the length of which plays a key role in early human embryonic development. Although the effect of sperm preparation techniques on major sperm characteristics, such as concentration, motility and morphology have been previously documented, the possible status of telomere length and its relation with sperm preparation techniques is not well-known for humans. The aim of this study was to investigate the role of density gradient centrifugation in the selection of spermatozoa with longer telomeres for use in assisted reproduction techniques in 105 samples before and after sperm processing. After density gradient centrifugation, the average telomere length of the sperm was significantly longer (6.51 ± 2.54 versus 5.16 ± 2.29, P average motile sperm rate was significantly higher (77.9 ± 11.8 versus 44.6 ± 11.2, P average DNA fragmentation rate was significantly lower (11.1 ± 5.9 versus 25.9 ± 12.9, P sperm count (rs = 0.58; P sperm with longer telomeres. Copyright © 2015 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  4. Soya-lecithin in extender improves the freezability and fertility of buffalo (Bubalus bubalis) bull spermatozoa.

    Science.gov (United States)

    Akhter, S; Ansari, M S; Andrabi, S M H; Rakha, B A; Ullah, N; Khalid, M

    2012-10-01

    Egg yolk is routinely used as a cryoprotectant in semen extenders. However, it may contain cryoprotective antagonists, and there are hygienic risks associated with its use. Proteins of plant origin, like soya-lecithin, lack these hazards. The aim of this study was to use soya-lecithin as a cryoprotectant in extender and to investigate its effects on in vitro quality and in vivo fertility of buffalo semen. Semen from three buffalo bulls was frozen in tris-citric extender containing 5.0%, 10% or 15% soya-lecithin or 20% egg yolk. Sperm motility, plasma membrane integrity and viability were assessed post-dilution, pre-freezing and post-thaw. In Post-dilution and pre-freezing, the values for motility, plasma membrane integrity and viability remained higher (p ≤ 0.05) in extenders containing 10% soya-lecithin and control compared with extender containing 5% and 15% soya-lecithin. However, motility, plasma membrane integrity and viability were higher (p lecithin compared with control and extenders containing 5% and 15% soya-lecithin. Semen from two buffalo bulls was frozen in tris-citric extender containing either 10% soya-lecithin or 20% egg yolk. Higher (p lecithin (56%) compared with 20% egg yolk (41.5%). The results suggest that 10% soya-lecithin in extender improves the freezability and fertility of buffalo bull spermatozoa and can be used as an alternate to egg yolk in cryopreservation of buffalo semen. © 2011 Blackwell Verlag GmbH.

  5. Evaluation of chromatin integrity of motile bovine spermatozoa capacitated in vitro.

    Science.gov (United States)

    Reckova, Z; Machatkova, M; Rybar, R; Horakova, J; Hulinska, P; Machal, L

    2008-08-01

    The efficiency of in vitro embryo production is highly variable amongst individual sires in cattle. To eliminate that this variability is not caused by sperm chromatin damage caused by separation or capacitacion, chromatin integrity was evaluated. Seventeen of AI bulls with good NRRs but variable embryo production efficiency were used. For each bull, motile spermatozoa were separated on a Percoll gradient, resuspended in IVF-TALP medium and capacitated with or incubated without heparin for 6 h. Samples before and after separation and after 3-h and 6-h capacitacion or incubation were evaluated by the Sperm Chromatin Structure Assay (SCSA) and the proportion of sperm with intact chromatin structure was calculated. Based on changes in the non-DFI-sperm proportion, the sires were categorized as DNA-unstable (DNA-us), DNA-stable (DNA-s) and DNA-most stable (DNA-ms) bulls (n=3, n=5 and n=9, respectively). In DNA-us bulls, separation produced a significant increase of the mean non-DFI-sperm proportion (p Capacitacion produced a significant decrease in the mean non-DFI-sperm proportion in H+ sperm (p capacitacion, the mean non-DFI-sperm proportion remained almost unchanged. In DNA-ms bulls, neither separation nor capacitacion had any effect on the mean non-DFI-sperm proportion. It can be concluded that, although separation and capacitacion may produce some changes in sperm chromatin integrity, these are not associated with different in vitro fertility of the bulls involved.

  6. Age-dependent changes in metabolic profile of turkey spermatozoa as assessed by NMR analysis

    Science.gov (United States)

    Di Iorio, Michele; Mannina, Luisa; Paventi, Gianluca; Rosato, Maria Pina; Cerolini, Silvia; Sobolev, Anatoly P.

    2018-01-01

    Metabolic profile of fresh turkey spermatozoa at three different reproductive period ages, namely 32, 44 and 56 weeks, was monitored by Nuclear Magnetic Resonance (NMR) spectroscopy and correlated to sperm quality parameters. The age-related decrease in sperm quality as indicated by reduction of sperm concentration, sperm mobility and osmotic tolerance was associated to variation in the level of specific water-soluble and liposoluble metabolites. In particular, the highest levels of isoleucine, phenylalanine, leucine, tyrosine and valine were found at 32 weeks of age, whereas aspartate, lactate, creatine, carnitine, acetylcarnitine levels increased during the ageing. Lipid composition also changed during the ageing: diunsaturated fatty acids level increased from 32 to 56 weeks of age, whereas a reduction of polyunsaturated fatty acids content was observed at 56 weeks. The untargeted approach attempts to give a wider picture of metabolic changes occurring in ageing suggesting that the reduction of sperm quality could be due to a progressive deficiency in mitochondrial energy producing systems, as also prompted by the negative correlation found between sperm mobility and the increase in certain mitochondrial metabolites. PMID:29534088

  7. Antioxidative effects of melatonin on kinetics, microscopic and oxidative parameters of cryopreserved bull spermatozoa.

    Science.gov (United States)

    Ashrafi, Iraj; Kohram, Hamid; Ardabili, Farhad Farrokhi

    2013-06-01

    Reactive oxygen species generated during the freeze-thawing process may reduce sperm quality. This study evaluates the effects of melatonin supplementation as an antioxidant in the semen extender on post-thaw parameters of bull spermatozoa. Melatonin was added to the citrate-egg yolk extender to yield six different final concentrations: 0, 0.1, 1, 2, 3 and 4mM. Ejaculates were collected from six proven Holstein bulls. Semen was diluted in the extender packaged in straws, which was frozen with liquid nitrogen. The semen extender supplemented with various doses of melatonin increased (peffective concentration of melatonin in microscopic evaluations of the bull sperm freezing extender was 2mM. The highest (pconcentration of melatonin in the semen extender and the highest activity of catalase (0.7±0.1) was obtained by 2mM melatonin. Four millimolar concentration of melatonin were reduced (pconcentration of melatonin in the semen extender improved the quality of post-thawed semen, which may associate with a reduction in lipid peroxidation as well as an increase in the total antioxidant capacity and antioxidant enzyme activity. Copyright © 2013 Elsevier B.V. All rights reserved.

  8. The formation and recovery of two-break chromosome rearrangements from irradiated spermatozoa of Drosophila melanogaster

    International Nuclear Information System (INIS)

    Leigh, B.

    1978-01-01

    Chromosome and chromatid-type rearrangements can be induced by exposure of spermatozoa of Drosophila to ionising radiation. A model, proposed to explain the formation and recovery of compound autosomes, has been extended to account for the induction of centric fragments capped by a duplication of paternal chromosome material. Three basic assumptions have been used; (1) that the sperm nucleus contains a haploid set of unreplicated chromosomes, (2) that the broken chromosome ends can be joined together before or after replication, and (3) that one of the first two cleavage nuclei may be lost and an adult organism derived from the other. The present paper reports a theoretical application of this combination of aasumptions to the general case of the formation and recovery of two-break rearrangements. This has led to an elucidation of the relation between repeats, compounds, fragments, and deficiencies on the one hand and inversions and translocations on the other hand. Dicentric chromosomes and segmental aneuploidy can be simply explained. A selective screen is formed by the segregation of chromatid rearrangements and the aneuploidy tolerance levels of the early cleavage nuclei. Thus there is an alternative way of explaining observations which might indicate preferential breakage or joining

  9. Effect of lactose, skim milk and Tris diluents on frozen buffalo spermatozoa

    Directory of Open Access Journals (Sweden)

    Ali Rastegarnia

    2007-08-01

    Full Text Available The composition of the extender in which semen is diluted before freezing plays a major role in successful cryopreservation of spermatozoa. This study was carried out to identify the suitable buffer for cryopreservation of buffalo semen. Sixteen split pooled ejaculates from two buffalo bulls possessing more than 70% visual sperm motility, were diluted at 370c either in lactose, skim milk or Tris extenders. The diluted semen was cooled to 40c within 2 hours, equilibrated at 40c for 4-6 hours following the addition of glycerol, filled in 0.5 ml French straws and frozen in a programmable cell freezer before plunging into liquid nitrogen. Semen was thawed at 370c for 30 seconds after 48 hours of storage inside liquid nitrogen. Post thaw visual sperm motility, plasma membrane integrity and acrosome morphology of each semen sample were assessed by warm plate microscopy at 370c, hypo-osmotic swelling test (HOST and giemsa staining, respectively. Analysis of variance revelated that percentage of post thaw visual sperm motility (Mean± standard deviation tended to be higher in Tris (50±3.6 than skim milk (44.5±2.5 and lower in lactose (24.4±10.5 extenders (P

  10. Detection of oncogenic human papillomavirus genotypes on spermatozoa from male partners of infertile couples.

    Science.gov (United States)

    Schillaci, Rosaria; Capra, Giuseppina; Bellavia, Carmela; Ruvolo, Giovanni; Scazzone, Concetta; Venezia, Renato; Perino, Antonio

    2013-11-01

    To evaluate the prevalence of human papillomavirus (HPV) sperm infection and its correlation with sperm parameters in patients who attended a fertility clinic. Cross-sectional clinical study. University-affiliated reproductive medicine clinic. A total of 308 male partners of couples undergoing in vitro fertilization techniques. Specimens of semen were collected from all patients. Sperm parameters were evaluated according to the World Health Organization manual. The presence of HPV DNA was researched by the combined use of two HPV assays and a highly sensitive nested polymerase chain reaction assay followed by HPV genotyping. To examine whether HPV was associated with the sperm, in situ hybridization (ISH) analysis was performed. Results of HPV investigation were compared with sperm parameters and ISH analysis. Twenty-four out of 308 semen samples (7.8%) were HPV DNA positive, but HPV infection did not seem to affect semen quality. Moreover, ISH revealed a clear HPV localization at the equatorial region of sperm head in infected samples. Oncogenic HPV genotypes were detected on spermatozoa from asymptomatic subjects, but a role of the infection in male infertility was not demonstrated. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  11. Impact of Storage and Purification on Mitochondrial Membrane Potential of Boar Spermatozoa

    Directory of Open Access Journals (Sweden)

    Aristotelis G. Lymberopoulos

    2013-05-01

    Full Text Available This study aimed to evaluate the effect of semen purification and storage on sperm mitochondrial membrane potential (ΔΨm. Gel-free whole ejaculates were collected from five proven fertile Large White boars aged two to three years. Aliquots of fresh semen were split, diluted in one step with commercial extenders and incubated at 37oC for 5-10 minutes. Semen was cooled to 18oC and packaged in 15-ml sterile propylene tubes. After 4-10 hours post-semen collection, stored semen was purified by colloidal centrifugation. After 48 hours post-semen collection, stored semen was incubated at 37oC and evaluated after 45 minutes for motility, velocity and sperm ΔΨm. Samples were stained with 2.99 μM JC-1 and 2.32 μM EH-1 and assessed by Fluorescence microscopy. After centrifugation a significant improvement of motility (P<0.035, and velocity (P<0.012 was noticed. The percentage of spermatozoa with intact plasma membrane and high/low mitochondrial membrane potential was statistical higher after centrifugation and storage at 18°C for 48 hours. In conclusion, colloidal purification of boar semen can improve sperm quality and  mitochondrial membrane potential.

  12. Boar spermatozoa cryopreservation in low glycerol/trehalose enriched freezing media improves cellular integrity.

    Science.gov (United States)

    Gutiérrez-Pérez, Oscar; Juárez-Mosqueda, María de Lourdes; Carvajal, Salvador Uribe; Ortega, María Elena Trujillo

    2009-06-01

    The use of glycerol for boar semen cryopreservation results in low fertility, possibly due to toxicity. This has led to recommend the use of solutions with less than 4% glycerol. Trehalose is a disaccharide known to stabilize proteins and biologic membranes during processes such as cryopreservation. Thus, it was decided to evaluate the cryoprotective effect of glycerol/trehalose mixtures. Effects on motility (M), viability (Vb) and acrosomal integrity (nA) were evaluated. Sperm samples were frozen in three different extenders: G4 contained 4% glycerol; T1 contained 1% glycerol plus 250 mM trehalose and T0.5 was constituted by 0.5% glycerol plus 250 mM trehalose. All extenders yielded similar post-freezing/thawing motility rates. Viability was diminished in T0.5 as compared to the others. In regard to acrosome integrity, it was twice as high (Pextender. Thus, T1 twice as many spermatozoa were alive, motile and intact, than in either T0.5 or G4, i.e. during freeze/thawing the use of T1 resulted in twice as many fertile cells as when using the other extenders. During our study, we noted that there were wide individual variations both in sperm viability and in motility.

  13. Motility analysis of circularly swimming bull spermatozoa by quasi-elastic light scattering and cinematography.

    Science.gov (United States)

    Craig, T; Hallett, F R; Nickel, B

    1982-04-01

    The Rayleigh-Gans-Debye approximation is used to predict the electric field autocorrelation functions of light scattered from circularly swimming bull spermatozoa. Using parameters determined from cinematography and modeling the cells as coated ellipsoids of semiaxes a = 0.5 micrometers, b = 2.3 micrometers, and c = 9.0 micrometers, we were able to obtain model spectra that mimic the data exactly. A coat is found to be a necessary attribute of the particle. It is also clear that these model functions at 15 degrees may be represented by the relatively simple function used before by Hallett et al. (1978) to fit data from circularly swimming cells, thus giving some physical meaning to these functional shapes. Because of this agreement the half-widths of experimental functions can now be interpreted in terms of an oscillatory frequency for the movement of the circularly swimming cell. The cinematographic results show a trend to chaotic behavior as the temperature of the sample is increased, with concomitant decrease in overall efficiency. This is manifested by a decrease in oscillatory frequency and translational speed.

  14. Hypospermatogenesis and spermatozoa maturation arrest in rats induced by mobile phone radiation.

    Science.gov (United States)

    Meo, Sultan Ayoub; Arif, Muhammad; Rashied, Shahzad; Khan, Muhammad Mujahid; Vohra, Muhammad Saeed; Usmani, Adnan Mahmood; Imran, Muhammad Babar; Al-Drees, Abdul Majeed

    2011-05-01

    To determine the morphological changes induced by mobile phone radiation in the testis of Wistar albino rats. Cohort study. Department of Physiology, College of Medicine, King Saud University, Riyadh, Saudi Arabia, from April 2007 to June 2008. Forty male Wistar albino rats were divided in three groups. First group of eight served as the control. The second group [group B, n=16] was exposed to mobile phone radiation for 30 minutes/day and the third group [group C, n=16] was exposed to mobile phone radiation for 60 minutes/day for a total period of 3 months. Morphological changes in the testes induced by mobile phone radiations were observed under a light microscope. Exposure to mobile phone radiation for 60 minutes/day caused 18.75% hypospermatogenesis and 18.75% maturation arrest in the testis of albino rats compared to matched controls. However, no abnormal findings were observed in albino rats that were exposed to mobile phone radiation for 30 minutes/day for a total period of 3 months. Long-term exposure to mobile phone radiation can cause hypospermatogenesis and maturation arrest in the spermatozoa in the testis of Wistar albino rats.

  15. Effect of reduced glutathione supplementation in semen extender on tyrosine phosphorylation and apoptosis like changes in frozen thawed Hariana bull spermatozoa.

    Science.gov (United States)

    Shah, Nadeem; Singh, Vijay; Yadav, Hanuman Prasad; Verma, Meena; Chauhan, Dharmendra Singh; Saxena, Atul; Yadav, Sarvajeet; Swain, Dilip Kumar

    2017-07-01

    To provide new insights into the mechanisms through which reduced glutathione (GSH) is able to protect spermatozoa, we tested the hypothesis that cryocapacitation and apoptosis like changes can contribute to the negative effect of freezing and thawing on bull spermatozoa, and that GSH prevent this damage. Having known protective effects of GSH in terms of a potent antioxidant, we evaluated capacitation, tyrosine phosphorylation and apoptosis like changes in bull spermatozoa after freezing and thawing in egg yolk tris glycerol extender containing (0.5m M-GSH-T1 & 1mM GSH-T2) and without GSH serving as the control (C). Forty ejaculates were collected from four Hariana bulls and were pooled due to non significant variations among the bull ejaculates for the evaluation of sperm attributes. Capacitation like changes, tyrosine phosphorylation, localization of tyrosine phosphorylated proteins, apoptosis like changes in terms of mitochondrial transmembrane potential and DNA fragmentation after final dilution, 4h of equilibration at 4°C and 24h after freezing and thawing were evaluated. GSH supplementation at 0.5mM showed significant reduction in B- and AR- pattern spermatozoa during all stages of semen freezing and thawing. Immunoblot revealed six proteins which were tyrosine phosphorylated and protein of 30 and 75kDa (p30, p75) were the major tyrosine phosphorylted proteins. On further analysis, the p30 showed differential variation in intensity in all the three groups after freezing and thawing. Positive immune reactivity for tyrosine phosphorylated proteins was found in neck, middle piece and post-acrosomal regions of spermatozoa. Addition of 0.5mM GSH decreased percentage of spermatozoa showing fragmented DNA and increased the percentage of spermatozoa having high transmembrane mitochondrial potential (P<0.05). This study demonstrates that GSH favours survival of bull spermatozoa by interfering with apoptotic and cryocapacitation pathways, and thereby protects the

  16. Evidence for compromised metabolic function and limited glucose uptake in spermatozoa from the teratospermic domestic cat (Felis catus) and cheetah (Acinonyx jubatus).

    Science.gov (United States)

    Terrell, Kimberly A; Wildt, David E; Anthony, Nicola M; Bavister, Barry D; Leibo, Stanley P; Penfold, Linda M; Marker, Laurie L; Crosier, Adrienne E

    2010-11-01

    Cheetahs and certain other felids consistently ejaculate high proportions (≥ 60%) of malformed spermatozoa, a condition known as teratospermia, which is prevalent in humans. Even seemingly normal spermatozoa from domestic cat teratospermic ejaculates have reduced fertilizing capacity. To understand the role of sperm metabolism in this phenomenon, we conducted a comparative study in the normospermic domestic cat versus the teratospermic cat and cheetah with the general hypothesis that sperm metabolic function is impaired in males producing predominantly pleiomorphic spermatozoa. Washed ejaculates were incubated in chemically defined medium containing glucose and pyruvate. Uptake of glucose and pyruvate and production of lactate were assessed using enzyme-linked fluorescence assays. Spermatozoa from domestic cats and cheetahs exhibited similar metabolic profiles, with minimal glucose metabolism and approximately equimolar rates of pyruvate uptake and lactate production. Compared to normospermic counterparts, pyruvate and lactate metabolism were reduced in teratospermic cat and cheetah ejaculates, even when controlling for sperm motility. Rates of pyruvate and lactate (but not glucose) metabolism were correlated positively with sperm motility, acrosomal integrity, and normal morphology. Collectively, our findings reveal that pyruvate uptake and lactate production are reliable, quantitative indicators of sperm quality in these two felid species and that metabolic function is impaired in teratospermic ejaculates. Furthermore, patterns of substrate utilization are conserved between these species, including the unexpected lack of exogenous glucose metabolism. Because glycolysis is required to support sperm motility and capacitation in certain other mammals (including dogs), the activity of this pathway in felid spermatozoa is a target for future investigation.

  17. Fetal Bovine Serum dalam Pengencer Tris Mempertahankan Kehidupan dan Keutuhan Membran Plasma Spermatozoa Semen Beku Domba Garut (FETAL BOVINE SERUM IN TRIS EXTENDER MAINTAINING SPERMATOZOA VIABILITY AND PLASMA MEMBRANE INTEGRITY OF GARUT RAM FROZEN SEMEN

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    Muhammad Rizal

    2013-12-01

    Full Text Available The objective of this study was to examine the effectiveness of fetal bovine serum (FBS against thequality of garut ram frozen semen. Semen was collected from one mature garut ram using artificial vagina.Fresh semen were evaluated then divided into four tubes at equal volume and each tube were diluted withTris extender containing 20% egg yolk (TEY-20, as control; TEY-20 + 8% FBS (FBS-8; TEY-20 + 10% FBS(FBS-10; and TEY-20 + 12% FBS (FBS-12, respectively. Semen at the concentration of 100x106 motilespermatozoa was loaded in 0.25 ml mini straw. Semen was equilibrated at 50C for three hours, then freezeand stored in liquid nitrogen container. The quality of the spermatozoa including percentages of motileand live spermatozoa, intact plasma membrane (IPM were evaluated following diluting, equilibratingand thawing process. A Complete Randomized design using four treatments and five replicates were usedin the study. The results showed that there was no significant difference (p<0.05 in percentage of motilespermatozoa following thawing between the control (44.0% and FBS-8 (46.0%, FBS-10 (48.0%, andFBS-12 (47.0%, respectively. The percentage of live spermatozoa and IPM were significantly higher (p<0.05in the FBS-8 (69.0% and 58.2%; FBS-10 (72.4% and 61.2%; FBS-12 (72.2% and 64.4% compared to thecontrol (64.8% and 52.8%, respectively. In conclusion, the addition of FBS into Tris extender was able tomaintain the viability and integrity of plasma membrane of garut ram frozen semen.

  18. Integrative systems analysis of diet-induced obesity identified a critical transition in the transcriptomes of the murine liver and epididymal white adipose tissue.

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    Kim, J; Kwon, E-Y; Park, S; Kim, J-R; Choi, S-W; Choi, M-S; Kim, S-J

    2016-02-01

    It is well known that high-fat diet (HFD) can cause immune system-related pathological alterations after a significant body weight gain. The mechanisms of the delayed pathological alterations during the development of diet-induced obesity (DIO) are not fully understood. To elucidate the mechanisms underlying DIO development, we analyzed time-course microarray data obtained from a previous study. First, differentially expressed genes (DEGs) were identified at each time point by comparing the hepatic transcriptome of mice fed HFD with that of mice fed normal diet. Next, we clustered the union of DEGs and identified annotations related to each cluster. Finally, we constructed an 'integrated obesity-associated gene regulatory network (GRN) in murine liver'. We analyzed the epididymal white adipose tissue (eWAT) transcriptome usig the same procedure. Based on time-course microarray data, we found that the genes associated with immune responses were upregulated with an oscillating expression pattern between weeks 2 and 8, relatively downregulated between weeks 12 and 16, and eventually upregulated after week 20 in the liver of the mice fed HFD. The genes associated with immune responses were also upregulated at late stage, in the eWAT of the mice fed HFD. These results suggested that a critical transition occurred in the immune system-related transcriptomes of the liver and eWAT around week 16 of the DIO development, and this may be associated with the delayed pathological alterations. The GRN analysis suggested that Maff may be a key transcription factor for the immune system-related critical transition thatoccurred at week 16. We found that transcription factors associated with immune responses were centrally located in the integrated obesity-associated GRN in the liver. In this study, systems analysis identified regulatory network modules underlying the delayed immune system-related pathological changes during the development of DIO and could suggest possible

  19. Oleoylethanolamide enhances β-adrenergic-mediated thermogenesis and white-to-brown adipocyte phenotype in epididymal white adipose tissue in rat

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    Juan Suárez

    2014-01-01

    Full Text Available β-adrenergic receptor activation promotes brown adipose tissue (BAT β-oxidation and thermogenesis by burning fatty acids during uncoupling respiration. Oleoylethanolamide (OEA can inhibit feeding and stimulate lipolysis by activating peroxisome proliferator-activating receptor-α (PPARα in white adipose tissue (WAT. Here we explore whether PPARα activation potentiates the effect of β3-adrenergic stimulation on energy balance mediated by the respective agonists OEA and CL316243. The effect of this pharmacological association on feeding, thermogenesis, β-oxidation, and lipid and cholesterol metabolism in epididymal (eWAT was monitored. CL316243 (1 mg/kg and OEA (5 mg/kg co-administration over 6 days enhanced the reduction of both food intake and body weight gain, increased the energy expenditure and reduced the respiratory quotient (VCO2/VO2. This negative energy balance agreed with decreased fat mass and increased BAT weight and temperature, as well as with lowered plasma levels of triglycerides, cholesterol, nonessential fatty acids (NEFAs, and the adipokines leptin and TNF-α. Regarding eWAT, CL316243 and OEA treatment elevated levels of the thermogenic factors PPARα and UCP1, reduced p38-MAPK phosphorylation, and promoted brown-like features in the white adipocytes: the mitochondrial (Cox4i1, Cox4i2 and BAT (Fgf21, Prdm16 genes were overexpressed in eWAT. The enhancement of the fatty-acid β-oxidation factors Cpt1b and Acox1 in eWAT was accompanied by an upregulation of de novo lipogenesis and reduced expression of the unsaturated-fatty-acid-synthesis enzyme gene, Scd1. We propose that the combination of β-adrenergic and PPARα receptor agonists promotes therapeutic adipocyte remodelling in eWAT, and therefore has a potential clinical utility in the treatment of obesity.

  20. Cryopreservation of boar semen. III: Ultrastructure of boar spermatozoa frozen ultra-rapidly at various stages of conventional freezing and thawing.

    Science.gov (United States)

    Bwanga, C O; Ekwall, H; Rodriguez-Martinez, H

    1991-01-01

    Ejaculated boar spermatozoa subjected to a conventional freezing and thawing process, were ultra-rapidly fixed, freeze-substituted and examined by electron microscopy to monitor the presence of real or potential intracellular ice and the degree of cell protection attained with the different extenders used during the process. Numerous ice crystal marks representing the degree of hydration of the cells were located in the perinuclear space of those spermatozoa not in proper contact with the extender containing glycerol (i.e. prior to freezing). The spermatozoa which were in proper contact with the extenders presented a high degree of preservation of the acrosomes, plasma membranes as well as the nuclear envelopes. No ice marks were detected in acrosomes before thawing, indicating that the conventional assayed cryopreservation method provided a good protection against cryoinjury. The presence of acrosomal changes (internal vesiculization, hydration and swelling) in thawed samples however, raises serious questions about the thawing procedure employed.

  1. Exposure of spermatozoa to dibutyl phthalate induces abnormal embryonic development in a marine invertebrate Galeolaria caespitosa (Polychaeta: Serpulidae).

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    Lu, Yonggang; Lin, Minjie; Aitken, Robert John

    2017-10-01

    In this study, we have investigated the impact of dibutyl phthalate (DBP) on early embryogenesis in a sessile marine invertebrate, Galeolaria caespitosa. DBP was found to induce sperm dysfunction as well as impaired and defective embryogenesis characterised by a particular pattern of abnormality. Thus, after the first cleavage, one blastomere in these abnormal embryos was able to carry out further mitoses, while the other arrested. Analysis of microtubules, chromosomes and actin filaments demonstrated that the mitotic spindles in the abnormal embryos were irregularly bent, shortened and unable to anchor to the cortex, resulting in the defective segregation of chromosomes. Within the non-dividing blastomeres, karyokinesis was found to continue at a slow pace as indicated by the presence of multiple sets of abnormal mitotic spindles. However, cytokinesis had been disrupted in these arrested cells due to a failure to assemble the contractile actin ring, as a result of which one pole of the embryos remained as one large, undivided cell. DBP was found to suppress the activity of superoxide dismutase in spermatozoa and, in association with this change, DBP-treated cells experienced oxidative stress as indicated by the presence of lipid aldehydes, such as 4-hydroxynonenal (4-HNE) in the sperm acrosome and neck. Adduction of lipid aldehydes at the level of the acrosome would be expected to impede the acrosome reaction and account for the significant decrease in fertilisation rates. 4-HNE generated as a consequence of lipid peroxidation in the sperm neck resulted in alkylation of the sperm centrioles. Such paternally damaged centrioles were inherited by the embryos and disrupted cytoskeletal protein organisation during early cleavage, generating the observed abnormalities in embryonic development. This research emphasises the vulnerability of spermatozoa to oxidative damage and highlights novel potential mechanisms for reproductive toxicity involving the alkylation of

  2. Cryopreservation of Iberian pig spermatozoa. Comparison of different freezing extenders based on post-thaw sperm quality.

    Science.gov (United States)

    De Mercado, Eduardo; Rodríguez, Ana; Gómez, Emilio; Sanz, Elena

    2010-03-01

    The aim of this study was to evaluate the cryoprotective effect of different freezing extenders against cryopreservation injuries on Iberian boar sperm. The sperm-rich fraction was collected and pooled from six sexually mature Iberian boars, and was frozen in different extenders containing glucose, lactose or fructose as sugar source and including Orvus ES Paste only in the freezing extender-2 (Glucose; Lactose and Fructose) or in both freezing extenders (Glucose2; Lactose2 and Fructose2). During the cryopreservation process, the supernatant was removed after the centrifugation step, then was extended with freezing extender-1 for the equilibration period and with freezing extender-2 immediately before freezing. Post-thaw sperm characteristics, such as plasma membrane integrity (SYBR-14/PI), mitochondrial function (Rhodamine 123) and acrosome integrity (NAR), were monitored. Overall sperm motility and the individual kinematic parameters of motile spermatozoa (assessed by the computer-aided sperm analysis system Sperm Class Analyzer [SCA]) were recorded in the different experimental treatments. Measurements were taken at 30 and 150 min post-thaw. The state of the acrosome after thawing did not show significant differences between the freezing extenders studied. Freezing-thawing caused a significant decrease (Pextenders. Furthermore, spermatozoa frozen with Orvus ES Paste in both freezing extenders exhibited lower (Pextender. The spermatozoa frozen with the Lactose extender and with Orvus ES Paste only in the second freezing extender showed a better evolution of the motility and kinematic characteristics (Pextenders studied in the present experiment affected the quality of frozen-thawed semen in Iberian boar.

  3. Ultrastructure of spermatozoa of spider crabs, family Mithracidae (Crustacea, Decapoda, Brachyura): Integrative analyses based on morphological and molecular data.

    Science.gov (United States)

    Assugeni, Camila de O; Magalhães, Tatiana; Bolaños, Juan A; Tudge, Christopher C; Mantelatto, Fernando L; Zara, Fernando J

    2017-12-01

    Recent studies based on morphological and molecular data provide a new perspective concerning taxonomic aspects of the brachyuran family Mithracidae. These studies proposed a series of nominal changes and indicated that the family is actually represented by a different number and representatives of genera than previously thought. Here, we provide a comparative description of the ultrastructure of spermatozoa and spermatophores of some species of Mithracidae in a phylogenetic context. The ultrastructure of the spermatozoa and spermatophore was observed by scanning and transmission electron microscopy. The most informative morphological characters analysed were thickness of the operculum, shape of the perforatorial chamber and shape and thickness of the inner acrosomal zone. As a framework, we used a topology based on a phylogenetic analysis using mitochondrial data obtained here and from previous studies. Our results indicate that closely related species share a series of morphological characteristics of the spermatozoa. A thick operculum, for example, is a feature observed in species of the genera Amphithrax, Teleophrys, and Omalacantha in contrast to the slender operculum observed in Mithraculus and Mithrax. Amphithrax and Teleophrys have a rhomboid perforatorial chamber, while Mithraculus, Mithrax, and Omalacantha show a wider, deltoid morphology. Furthermore, our results are in agreement with recently proposed taxonomic changes including the separation of the genera Mithrax (previously Damithrax), Amphithrax (previously Mithrax) and Mithraculus, and the synonymy of Mithrax caribbaeus with Mithrax hispidus. Overall, the spermiotaxonomy of these species of Mithracidae represent a novel set of data that corroborates the most recent taxonomic revision of the family and can be used in future taxonomic and phylogenetic studies within this family. © 2017 Wiley Periodicals, Inc.

  4. Vaginal mucus from ewes treated with progestogen sponges affects quality of ram spermatozoa.

    Science.gov (United States)

    Manes, Jorgelina; Ríos, Glenda; Fiorentino, María Andrea; Ungerfeld, Rodolfo

    2016-03-15

    The use of intravaginal sponges (IS) to synchronize estrous onset in ewes provokes vaginitis, an increase in the vaginal bacterial load, and growth of bacterial species that are not present during spontaneous estrous behavior. The objective of the study was to compare the functional sperm parameters after incubating it with mucus collected from the vagina of ewes during spontaneous estrus or estrous synchronized with IS. Pooled spermatozoa were co-incubated with: (1) vaginal mucus collected from ewes in spontaneous estrus; (2) vaginal mucus collected from ewes in estrus pretreated with progestogen-impregnated IS; (3) synthetic mucus; and (4) medium without mucus as a control group. Sperm samples were evaluated after incubating it for 30 and 90 minutes. The number of colony-forming units (CFUs/mL), pH, and osmolality were greater in the mucus collected from ewes treated with IS than from those untreated (P = 0.046; P ewes treated with IS than in the other three treatments both, 30 and 90 minutes after incubation (P = 0.0009 and P ewes treated with IS had a lower percentage of sperm with intact plasma membrane than all the other treatments (P ewes treated with IS than in the other three treatments (P ewes during their spontaneous estrus (P = 0.0005). The lowest percentages of sperm with acrosome damage were observed in sperm incubated with mucus collected from sheep in spontaneous estrus for 30 and 90 minutes (P ewes treated with IS than in the other three groups (P = 0.0005). The functionality and the viability of ram sperm is negatively affected by the cervical mucus of ewes pretreated with progestagen-impregnated IS used in estrous synchronization treatments. This may partially explain the decrease in conception rate obtained with treatments with IS. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. The quantification of spermatozoa by real-time quantitative PCR, spectrophotometry, and spermatophore cap size.

    Science.gov (United States)

    Doyle, Jacqueline M; McCormick, Cory R; DeWoody, J Andrew

    2011-01-01

    Many animals, such as crustaceans, insects, and salamanders, package their sperm into spermatophores, and the number of spermatozoa contained in a spermatophore is relevant to studies of sexual selection and sperm competition. We used two molecular methods, real-time quantitative polymerase chain reaction (RT-qPCR) and spectrophotometry, to estimate sperm numbers from spermatophores. First, we designed gene-specific primers that produced a single amplicon in four species of ambystomatid salamanders. A standard curve generated from cloned amplicons revealed a strong positive relationship between template DNA quantity and cycle threshold, suggesting that RT-qPCR could be used to quantify sperm in a given sample. We then extracted DNA from multiple Ambystoma maculatum spermatophores, performed RT-qPCR on each sample, and estimated template copy numbers (i.e. sperm number) using the standard curve. Second, we used spectrophotometry to determine the number of sperm per spermatophore by measuring DNA concentration relative to the genome size. We documented a significant positive relationship between the estimates of sperm number based on RT-qPCR and those based on spectrophotometry. When these molecular estimates were compared to spermatophore cap size, which in principle could predict the number of sperm contained in the spermatophore, we also found a significant positive relationship between sperm number and spermatophore cap size. This linear model allows estimates of sperm number strictly from cap size, an approach which could greatly simplify the estimation of sperm number in future studies. These methods may help explain variation in fertilization success where sperm competition is mediated by sperm quantity. © 2010 Blackwell Publishing Ltd.

  6. Soy lecithin interferes with mitochondrial function in frozen-thawed ram spermatozoa.

    Science.gov (United States)

    Del Valle, I; Gómez-Durán, A; Holt, W V; Muiño-Blanco, T; Cebrián-Pérez, J A

    2012-01-01

    Egg yolk and milk are the 2 major membrane cryoprotectants commonly used in freezing media for the long-term preservation of semen (alone or in combination with others). However, in recent years, there have been increasing arguments against the use of egg yolk or milk because of the risk of introducing diseases through the use of cryopreserved semen. In this study, we analyzed the protective effect of lecithin as an alternative to egg yolk for the cryopreservation of ram semen, using a range of functional markers for sperm viability, motility, apoptosis, and mitochondrial functionality analyses (mitochondrial inner membrane surface [MIMS], mitochondrial inner membrane potential [MIMP], and cell membrane potential) as methods of assessment in samples diluted in 3 different media: Tris-citrate-glucose as control and 2 media supplemented with soy lecithin or egg yolk. The results showed that lecithin was able to effectively protect certain sperm quality characteristics against freezing-induced damage. However, lecithin induced loss of mitochondrial membrane potential or mitochondrial loss that was not reflected by modifications in sperm motility in fresh semen. MIMS and MIMP values decreased in thawed lecithin-treated samples, concomitant with a lower (P lecithin may have affected the inner mitochondrial membrane in frozenthawed spermatozoa and confirmed that sublethal damages that seriously affect sperm functionality, not detected by classic sperm quality analyses, can be evidenced by changes in the inner mitochondrial membrane surface. These findings strengthen the relationship between mitochondrial membrane potential and motility and show that the mitochondrial alterations induced by the cryopreservation process could be specific targets for the improvement of semen cryopreservation protocols.

  7. Cooperative functions of manganese and thiol redox system against oxidative stress in human spermatozoa

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    Amrit Kaur Bansal

    2009-01-01

    Full Text Available Aims: In this study, the effects of 0.1 mM Mn 2+ on thiol components (total thiols [TSH], glutathione reduced [GSH], glutathione oxidized [GSSG] and redox ratio [GSH/ GSSG] have been determined in human spermatozoa. Settings and Design: The subjects of the study were healthy males having more than 75% motility and 80 x 10 6 sperms/mL. Materials and Methods: Fresh semen was suspended in phosphate-buffered saline (PBS (pH 7.2 and this suspension was divided into eight equal fractions. All fractions, control (containing PBS and experimental (treated/untreated with [ferrous ascorbate, FeAA - 200 FeSO 4 μM, 1000 μM ascorbic acid, nicotine (0.5 mM and FeAA + nicotine], supplemented/unsupplemented with Mn 2+ [0.1 mM], were incubated for 2 h at 378C. These fractions were assessed for determining the thiol components. Statistical Analysis: The data were statistically analyzed by Students " t" test. Results and Conclusions: Ferrous ascorbate, nicotine and ferrous ascorbate + nicotine induced oxidative stress and decreased GSH and redox ratio (GSH/GSSG ratio but increased the TSH and GSSG levels. Mn 2+ supplementation improved TSH, GSH and redox ratio (GSH/GSSG but decreased the GSSG level under normal and oxidative stress conditions. Thiol groups serve as defense mechanisms of sperm cells to fight against oxidative stress induced by stress inducers such as ferrous ascorbate, nicotine and their combination (ferrous ascorbate + nicotine. In addition, Mn 2+ supplementation maintains the thiol level by reducing oxidative stress.

  8. Evaluation of Serial Thawing-Refreezing on Human Spermatozoa Resistance Using Cryovials and Straws

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    Fatemeh Ghasemian

    2012-01-01

    Full Text Available Background: We designed this study to detect the cryoinjury rate on human sperm after serialfreezing and thawing, taking into consideration the effects of using cryovials and straws.Materials and Methods: In this experimental study, semen specimens obtained from 15 subjectswere divided into normozoospermic and oligozoospermic groups. Each of the normozoospermicand oligozoo spermic semen specimens were additionally divided into two groups: i. washed andii. unwashed. Specimens were repeatedly freeze-thawed by using cryovials and straws with thefast liquid nitrogen vapor method, until no motile sperm remained. Sperm motility, recovery, andmorphology rate were then determined after thawing, and compared between the groups whiletaking into consideration the effects of using cryovials and straws.Results: Motile spermatozoa were observed in all normozoospermic samples up to thaw 6 with bothcryovials and straws while in oligozoospermic specimens up to thaw 4 (straw and thaw 3 (cryovialin the freeze-thawing cycle. Normozoospermic sample analysis showed no significant difference inmorphology rate. There was a significant increase in motility and recovery percentages for washedsamples, which was observed with straws in compared to the unwashed groups. Oligozoospermicsample analysis indicated a significant increase in motility, recovery (p<0.01, and morphology(p<0.001 rates in washed specimens compared to unwashed specimens using straws. Theimportance of washing sperm was obvious for oligozoospermic specimens.Conclusion: Normozoospermic sperm resisted freezing longer than oligozoospermic sperm. Use ofstraws and cryovials made significant differences in motility, recovery, and morphology of sperm ineach thaw. This difference was slightly higher for oligozoospermic specimens. Results indicated thatthe percentage of motility was higher for washed normozoospermic specimens in each thaw whenstraws were used, whereas the percentage of motility, recovery, and

  9. Quality and fertilizing capacity of boar spermatozoa during liquid storage in extender supplemented with different antibiotics.

    Science.gov (United States)

    Bryła, Magdalena; Trzcińska, Monika

    2015-12-01

    The aim of the study was to determine the effect of antibiotics on quality parameters and fertilizing capacity of boar sperm during liquid preservation. In the first experiment, semen was diluted in an extender containing 200 μg/mL of gentamicin as a control and diluted in a modified extenders: Ext I (contained 200 μg/mL florfenicol), Ext II (contained 200 μg/mL polymyxin B), Ext III (contained 100 μg/mL gentamicin and 100 μg/mL florfenicol) and Ext IV (contained 100 μg/mL gentamicin and 100 μg/mL polymyxin B). The semen was stored for ten days. Sperm quality was evaluated based on the motility (CASA; TM: total motility; PM: progressive motility), membrane integrity (YO-PRO-1/PI assay), mitochondrial activity (JC-1) and DNA integrity (TUNEL). The highest PM% (62.5 ± 9.6) was observed in Ext III at Day 6 of storage. The highest sperm viability and mitochondrial transmembrane potential was noticed at the end of the storage period in Ext III. Long-term storage did not induce DNA fragmentation in the extenders analyzed. In the second experiment, semen diluted in the control extender and in the extender providing the highest quality spermatozoa on Day 10 (Ext III) was used for artificial insemination (AI) of synchronized gilts. Our studies showed that the highest reproductive performance of inseminated gilts (pregnant gilts: 97.0%, litter size: 11.4 ± 1.2) occurred with Ext III semen dilution. The combination of 100 μg/mL gentamicin and 100 μg/mL florfenicol in the extender maintained sperm motility, membrane integrity and mitochondrial activity and enhanced the higher reproduction success. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. PEMBERIAN EKSTRAK BUAH JUWET (Syzygium cumini L. TERHADAP JUMLAH DAN MORFOLOGI SPERMATOZOA TIKUS PUTIH (Rattus sp. JANTAN YANG TERPAPAR ASAP ROKOK

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    Fitria Dwijayanti

    2017-03-01

    Full Text Available The research to determine the effect of fruit extracts juwet on the number and morphology of spermatozoa were superbly exposed to smoke has been conducted. This study uses rats aged 3 month, 200-210 gram, total 24 rats consisting of 4 treatment, is control, smoke of cigarette, juwet fruit extracts, and smoke of cigarette to juwet fruit extracts. Each treatment consisting 6 replications, each consisting of 1 rat. Before, acclimatization of rat treated 7 days. Way of giving juwet fruit extracts the gavage method 2 ml once daily for 48 days, while the CMC-Na 0,5% is given control. The exposure to cigarette smoke is given from an aerator pump , to given once daily for 48 days. Data were analyzed with ANOVA, if they were 5% significantly different would be followed by a DMRT. The result showed that the number of abnormal sperm morphology were significantly diferent (p<0,05. The averange number of abnormal spermatozoa K1 group was higher than K3 group. The number of spermatozoa was not significant different between treatments. The average number of spermatozoa K1 lower than K3. This is because the smoke of cigarette increased formation of ROS and the resulting stress oxidative and cause cell damage tissues and organs, especially to the reproductive system. Juwet fruits extract have been able to improve the count and morphologyof rats spermatozoa expose to smoke. This case because of antioxidant from juwet fruit can be protect cell from the free radical attack. Keywords: Rattus sp, smoke of cigarette, java plum  fruit, spermatozoa, , ROS, antioxidant.

  11. The Drosera Extract as an Alternative In Vitro Supplement to Animal Semen: Effects on Bovine Spermatozoa Activity and Oxidative Balance

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    Eva Tvrdá

    2015-05-01

    Full Text Available In vitro storage and processing of animal semen is considered to be a risk factor to spermatozoa activity, possibly leading to reduced fertility and litter sizes following artificial insemination (AI. A variety of substances isolated from natural resources have the potential to exhibit protective or antioxidant properties on the spermatozoon, thus they may extend the lifespan of stored semen. Drosera (Drosera rotundifolia L. has been shown to possess antimicrobial, anti-inflammatory and antioxidant properties, making the plant extract a potential candidate for preserving liquid animal semen during in vitro storage. This study compared the ability of different concentrations of Drosera extract on the motility, viability and superoxide production of bovine spermatozoa during different time periods (0, 2, 6, 12 and 24h of in vitro culture. Spermatozoa motility was assessed using the SpermVisionTM CASA (Computer aided sperm analysis system. Cell viability was examined using the metabolic activity MTT assay and the nitroblue-tetrazolium (NBT test was applied to quantify the intracellular superoxide formation. The CASA analysis revealed that Drosera extract supplementation was able to prevent a rapid decline of spermatozoa motility, especially in the case of concentrations ranging between 1 and 5 mg/mL (P<0.001 with respect to Times 6h, 12h and 24h. At the same time, concentrations ranging between 1 and 10 mg/mL of the extract led to a significant preservation of the cell viability throughout short-term (P<0.05 in case of Time 6h as well as long-term periods of the experiment (P<0.01 with respect to Time 12h, and P<0.001 in case of Time 24h. 10 and 5 mg/mL of the extract exhibited antioxidant characteristics, translated into a significant reduction of the intracellular superoxide production, particularly notable at Times 12h (P<0.01 and 24h (P<0.001. The results indicate that the Drosera extract is capable of delaying the damage inflicted to the

  12. Comparative studies of dose-response curves for recessive lethal mutations induced by ethylnitrosourea in spermatogonia and in spermatozoa of Drosophila melanogaster

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    Yoshikawa, I.; Ayaki, T.; Ohshima, K.

    1984-01-01

    Induction of recessive lethal mutation by N-ethyl-N-nitrosourea (ENU) was studied for the second chromosome of spermatogonia and spermatozoa in Drosophila melanogaster. ENU (0.03, 0.3, and 1.0 mM) was given to flies by dissolving it in feeding sucrose solution. When plotted against absorbed doses of ENU, the observed frequencies to recessive lethals showed a linear relationship for induction in spermatozoa but a sigmoidal relationship for induction in spermatogonia. These results suggest that in spermatogonia ENU-induced mutational damage is more repairable in a lower dose range of ENU. Mosaic lethal mutations were induced by ENU but not in spermatogonia.

  13. Negative effect of combined cysteine and glutathione in soy lecithin-based extender on post-thawed ram spermatozoa.

    Science.gov (United States)

    Zhandi, Mahdi; Sharafi, Mohsen

    2015-09-01

    This study was conducted to evaluate the effect of combined cysteine and glutathione in soy lecithin-based semen extender on post-thawed ram sperm quality. A total of 28 ejaculates were collected twice a week (from four rams) during breeding season. In each replicate, semen samples (n = 4, one ejaculate for each ram) were pooled and divided into three equal parts, and each part was diluted with one of following extender: (1) soy lecithin-based extender containing no cysteine and no glutathione (C0-G0), (2) soy lecithin-based extender containing cysteine (5 mM) and glutathione (5 mM) (C5-G5), and (3) soy lecithin-based extender containing cysteine (10 mM) and glutathione (10 mM) (C10-G10). After freeze-thawing process, motility and velocity parameters, plasma membrane integrity and functionality, mitochondrial activity, and apoptosis features of spermatozoa were evaluated. The obtained results showed that total and progressive motility, plasma membrane integrity and functionality, and live post-thawed spermatozoa was lower in C10-G10 extender compared to C0-G0 and C5-G5 extenders (P 0.05). In conclusion, it seems that high concentration of combined cysteine and glutathione in soy lecithin-based semen extender has a detrimental effect of post-thawed ram sperm quality.

  14. Flow cytometric evaluation of antibiotic effects on viability and mitochondrial function of refrigerated spermatozoa of Nile tilapia

    Science.gov (United States)

    Segovia, M.; Jenkins, J.A.; Paniagua-Chavez, C.; Tiersch, T.R.

    2000-01-01

    Improved techniques for storage and evaluation of fish sperm would enhance breeding programs around the world. The goal of this study was to test the effect of antibiotics on refrigerated sperm from Nile tilapia (Oreochromis niloticus) by use of flow cytometry with 2 dual-staining protocols for objective assessment of sperm quality. Concentrations of 1 x 109 sperm/mL were suspended in Ringer's buffer at 318 mOsmol/kg (pH 8.0). The fluorescent stains Sybr 14 (10 ??M), propidium iodide (2.4 mM), and rhodamine 123 (0.13 ??M) were used to assess cell viability and mitochondrial function. Three concentrations of ampicillin, gentamicin, and an antibiotic/antimycotic solution were added to fresh spermatozoa. Motility estimates and flow cytometry measurements were made daily during 7 d of refrigerated storage (4 ??C). The highest concentrations of gentamicin and antibiotic/antimycotic and all 3 concentrations of ampicillin significantly reduced sperm viability. The highest of each of the 3 antibiotic concentrations significantly reduced mitochondrial function. This study demonstrates that objective sperm quality assessments can be made using flow cytometry and that addition of antibiotics at appropriate concentrations can lengthen refrigerated storage time for tilapia spermatozoa. With minor modifications, these protocols can be adapted for use with sperm from other species and with other tissue types.

  15. Freezability of water buffalo bull (Bubalus bubalis) spermatozoa is improved with the addition of curcumin (diferuoyl methane) in semen extender.

    Science.gov (United States)

    Shah, S A H; Andrabi, S M H; Qureshi, I Z

    2017-10-01

    Effects of curcumin as antioxidant in extender were evaluated on freezability of buffalo spermatozoa. Semen from each of the five bulls (n = 3 replicates, six ejaculates/bull, a total of 30 ejaculates) was diluted in Tris-citric acid extender containing curcumin (0.5, 1.0, 1.5 or 2.0 mM) or control. At pre-freezing and post-thawing, total antioxidant contents (μM/L) and lipid peroxidation levels (μM/ml) were higher (p curcumin and control. At post-thawing, progressive motility (PM, %) and rapid velocity (RV, %) were higher (p curcumin and control (except in case of RV, 1.5 was similar with 1.0 mM). Kinematics (average path velocity, μm/s; straight-line velocity, μm/s; curved-line velocity, μm/s; straightness, %; linearity, %), in vitro longevity (%, PM and RV) and DNA integrity (%) at post-thawing were higher (p curcumin and control. We concluded that freezability of water buffalo spermatozoa is improved with the addition of 1.5 mM curcumin in extender. © 2016 Blackwell Verlag GmbH.

  16. Localisation of calcium-binding proteins in ram spermatozoa using the immunofluorescence technique

    International Nuclear Information System (INIS)

    Sabrina Sukardi; Watson, P.F.

    2000-01-01

    Localization of two calcium-binding proteins (proteins A and B) believed to be involve in membrane fusion on whole spermatozoa were carried out in two stages; before and after the acrosome reaction, the reaction being a prerequisite to fertilization. Determination of the acrosome reaction and sperm viability is carried out using fluorescent dyes i.e., FITC-conjugated Pisum sativum agglutinin (PSA) and propidium iodide (PI) respectively. Polyclonal antibodies were raised in rabbits. Ejaculated semen was diluted in buffer and loaded into tubes. Acrosome reaction was induced with calcium ionophore A23187 at 390 degree C. PI was added to the sub-samples at time 0 and 45 minutes. Excess PI, ionophore and seminal plasma was filtered out with a syringe. Smears were made on slides and air-dried. The cells were pemeabilised with ethanol and rinsed in PBS. Batch 1 slides were incubated with FITC-PSA in the dark while batch H slides were incubated in 1% sheep serum. Batch II slides were then rinsed in PBS twice and incubated in both antiserum and pre-immune serum (negative control). These slides were then incubated in FITC-conjugated secondary antibody (anti-rabbit IgG) and kept in the dark. After final washing and mounted, both batches of slides were viewed immediately using fluorescence microscope. Results obtained before acrosome reaction showed localization of both antibodies to the whole sperm head, along the midpiece and tail. The acrosomes were also intact and cells were viable. After the acrosome reaction, localization of both antibodies were observed at the post-acrosomal region, midpiece, tail and the equatorial segment with no binding to the acrosome. Cells were mainly acrosome-reacted and dying. No binding was observed with pre-immune serum. Results indicate that the antigens were present in the acrosome and the change in binding suggests that the antigens have been redistributed after commencement of the acrosome reaction. The findings suggest that the proteins

  17. Effects of ions on the motility of fresh and demembranate spermatozoa of common carp (Cyprinus carpio)and paddlefish (Polyodon spathula)

    Czech Academy of Sciences Publication Activity Database

    Linhart, Otomar; Cosson, J.; Mims, S. D.; Rodina, Marek; Gela, David; Shelton, W. L.

    2004-01-01

    Roč. 28, - (2004), s. 203-205 ISSN 0920-1742 R&D Projects: GA ČR GA524/03/0178 Grant - others:CBG Program(US) KYX-01-11469 Institutional research plan: CEZ:AV0Z5045916 Keywords : fish * spermatozoa * motility Subject RIV: ED - Physiology Impact factor: 0.399, year: 2004

  18. Dose-response relationship for the induction of structural chromosome aberrations in human spermatozoa after in vitro exposure ti tritium. beta. -rays

    Energy Technology Data Exchange (ETDEWEB)

    Kamiguchi, Yujiroh; Tateno, Hiroyuki; Mikamo, Kazuya (Asahikawa Medical College (Japan). Department of Biological Sciences)

    1990-02-01

    THe effects of tritium (HTO) {beta}-rays on human sperm chromosomes were studied using our interspecific in vitro fertilization system between human spermatozoa and zona-free hamster oocytes. Semen samples were treated with media containing 1.53-24.3 mCi/ml HTO for about 80 min. 1290 spermatozoa from the controls and 1842 spermatozoa from the irradiated groups were karyotyped. The incidence of spermatozoa with structural chromosome aberrations increased linearly with increasing dosage. Breakage-type aberrations occurred far more frequently than exchange-type. Chromosome-type aberrations appeared far more frequently than chromatid-ype. All of these types of aberrations showed linear dose-dependent increases. The RBE valus of HTO {beta}-rays relative to X-rays were calculated for the above-mentioned 5 indices, respectively. Their RBE values franged from 1.89 to 3.00 when the absorbed dose was estimated to be the minimum, whereas the values ranged between 1.04 and 1.65 when the absorbed dose was estimated to be the maximum. (author). 15 refs.; 3 figs.; 4 tabs.

  19. Injection of Carica papaya L. Seed Extract of Cibinong Variety to Macaca fascicularis L. and its Effect to Quality of Spermatozoa and Level of Testosterone Hormone.

    Directory of Open Access Journals (Sweden)

    Tuti Nuraini

    2012-06-01

    Full Text Available Lack of contraceptive choices which meet the requirements is one of the contributing factors to less participation of man in contraceptive use. This research aimed to study the effectiveness of natural material for alternative male contraception, by injecting papaya seed extract with Cibinong variety (Carica papaya L. to long tail monkey (Macaca fascicularis L. The research was conducted at Primates Study Center, Institute of Agriculture, Bogor. Total samples of this research were 8 monkeys, with three intervention groupsand one control group. Papaya seed extract was injected via intramuscular in 21 days, with dose for each group were 40 mg/monkey, 80 mg/monkey, and 120 mg/monkey. Data analysis of spermatozoa quality (motility, viability, morfology was done by using Cochran test before and after intervention stages, and during recovery stage. Meanwhile, data aalysis of spermatozoa concentration and testosterone hormone level was done by using Friedman test. Result of this reseachdemonstrated reduction of motility, viability, and morfology ofspermatozoa after inejction of papaya seed extract and increase to normal level at recovery stage (p ≤ 0.05. These results was supported with cement aglutination. The most effective dose was at 40 mg/monkey/day, with reduction of spermatozoa motility from 87.5 % to 40%, and reduction of testosterone level from 2.35 ng/mL to 1.83 ng/mL. Even though spermatozoa motility and testosterone hormone levelreduced, but its conditions were still in good condition category.

  20. Cross-talk between free and bound spermatozoa to modulate initial sperm:egg ratios at the site of fertilization in the mammalian oviduct.

    Science.gov (United States)

    Hunter, R H F; Gadea, J

    2014-08-01

    This essay proposes that highly localized communication between free and bound spermatozoa in the caudal portion of the oviduct acts to regulate the numbers detaching from the epithelium and progressing to the site of fertilization close to the time of ovulation. Low initial sperm:egg ratios are essential for monospermic fertilization. Liberation of surface macromolecules and metabolic prompting from activated spermatozoa, together with altered patterns of sperm movement and dynamic differences in intracellular Ca(2+) ion status between neighboring sperm cells, would influence the progressive release of spermatozoa from the reservoir in the oviduct isthmus. Different intensities of preovulatory epithelial binding, reflecting a range of states in the sperm surface membranes and associated proteins, would provide a further explanation for a chronologically staggered periovulatory detachment of spermatozoa. Intimate sperm-sperm interactions within the confines of the oviduct isthmus offer a sensitive means of fine-tuning the vanguard of competent male gametes reaching the isthmo-ampullary junction. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Dimensions of human ejaculated spermatozoa in Papanicolaou-stained seminal and swim-up smears obtained from the Integrated Semen Analysis System (ISAS(®)).

    Science.gov (United States)

    Bellastella, Giuseppe; Cooper, Trevor G; Battaglia, Marina; Ströse, Anda; Torres, Inma; Hellenkemper, Barbara; Soler, Carles; Sinisi, Antonio A

    2010-11-01

    Objective measurements are required for computer-aided sperm morphometric analysis (CASMA) machines to distinguish normal from abnormal sperm heads. The morphometric characteristics of spermatozoa in 72 samples of semen and of spermatozoa from 72 other semen samples after swim-up were quantified by the semi-automated Integrated Sperm Analysis System (ISAS) computer-aided system, which measured the sperm head parameters length (L), width (W), area (A), perimeter (P), acrosomal area (Ac), and the derived values L/W and P/A. For each man a homogeneous population of distributions characterized seminal spermatozoa (7 942 cells: median values L 4.4 μm, W 2.8 μm, A 9.8 μm(2), P 12.5 μm, Ac 47.5%, L/W 1.57, P/A 1.27), and there was no significant difference in within- and among-individual variation. Different men could have spermatozoa of significantly different dimensions. Head dimensions for swim-up spermatozoa from different men (4 812 cells) were similar to those in semen, differing only by 2%-5%. The values of L, W and L/W fell within the limits given by the World Health Organization (WHO). Although these samples were not biologically matched, linear mixed-effects statistical analyses permitted valid comparison of the groups. A subpopulation of 404 spermatozoa considered to fit the stringent criteria of WHO 'normal' seminal spermatozoa from both semen and swim-up were characterized by median values (and 95% confidence intervals) of L, 4.3 μm (3.8-4.9), W, 2.9 μm (2.6-3.3), A, 10.2 μm(2) (8.5-12.2), P, 12.4 μm (11.3-13.9), Ac, 49% (36-60), L/W, 1.49 (1.32-1.67) and P/A, 1.22 (1.11-1.35). These median values fall within the 95th centile confidence limits given by WHO, but the confidence intervals for L and W were larger. Although these differences in head dimensions among men and after swim-up could be detected by CASMA, the small differences make it unlikely that technicians would be able to distinguish them. The values could be used as default sperm head

  2. Normal sperm morphology and changes of semen characteristics and abnormal morphological spermatozoa among peri-mating seasons in captive japanese black bears (Ursus thibetanus japonicus).

    Science.gov (United States)

    Okano, Tsukasa; Murase, Tetsuma; Nakamura, Sachiko; Komatsu, Takeshi; Tsubota, Toshio; Asano, Makoto

    2009-04-01

    The objectives of this study were to obtain morphological data for normal spermatozoa and to investigate seasonal changes (the early, mid- and post-mating seasons) in abnormal morphology of spermatozoa and the characteristics of semen in Japanese black bears. Semen was collected by electroejaculation from 34 captive male Japanese black bears a total of 74 times. Length of head, width of head, length of midpiece and total length of the spermatozoa were 6.3 +/- 0.4, 4.5 +/- 0.3, 10.4 +/- 0.7 and 69.6 +/- 3.1 mum (mean +/- SD; 20 semen, 200 spermatozoa), respectively. In the semen collected during the mid-mating season, ejaculate volume, ejaculate pH, sperm concentration, total sperm count, motility, viability and intact acrosomes were 0.46 +/- 0.36 ml, 7.3 +/- 0.4, 659 +/- 644 x 10(6)/ml, 214 +/- 208 x 10(6), 82.9 +/- 9.6%, 89.3 +/- 9.5% and 97.0 +/- 3.2% (mean +/- SD; n=21, in ejaculate pH n=8), respectively. Sperm motility and viability in the early (n=7) and mid-mating (n=21) seasons were significantly higher than in the post-mating (n=8) season. The rates of detached heads in the early and mid-mating season were significantly lower than in the post-mating season. The main abnormal morphologies observed (mean +/- SD%; n=23) were simply bent tail (19.9 +/- 22.6), distal droplets (13.5 +/- 11.7), proximal droplets (9.6 +/- 7.8), teratoid spermatozoa (6.7 +/- 10.7), knobbed acrosome (4.9 +/- 8.6), acrosome damage (3.7 +/- 2.8) and bent midpiece (3.7 +/- 5.1). The data will be useful for artificial breeding and further research on male reproductive physiology in this species.

  3. Superoxide dismutase (SOD) in boar spermatozoa: purification, biochemical properties and changes in activity during semen storage (16°C) in different extenders.

    Science.gov (United States)

    Orzołek, Aleksandra; Wysocki, Paweł; Strzeżek, Jerzy; Kordan, Władysław

    2013-03-01

    The antioxidant system in semen is composed of enzymes, low-molecular weight antioxidants and seminal plasma proteins. Loss of enzymatic activity of superoxide dismutase (SOD) during semen preservation may cause insufficient antioxidant defense of boar spermatozoa. The aim of this study was to isolate and characterize SOD molecular forms from spermatozoa and to describe changes in SOD activity in boar sperm during preservation at 16°C. Sperm extracts were prepared from fresh or diluted semen and used for SOD purification or activity measurement. Ion-exchange chromatography and gel filtration was used to purify SOD molecular forms. BTS, Dilu Cell, M III and Vitasem were used as diluents for 5-day storage of semen at +16°C. The molecular form of SOD released from spermatozoa after cold shock and homogenization had a molecular weight of approximately 67kDa. The activity of the SOD form was the highest at pH 10 within the temperature range between 20 and 45°C. The enzymatic activity of form released after cold shock was inhibited by H2O2 and diethyldithiocarbamate (DDC; by 65 and 40%, respectively). The SOD form released by homogenization was inhibited by H2O2 and DDC (40%). The molecular form released after urea treatment was a 30kDa protein with maximum activity at 20°C and pH 10. Enzymatic activity of this form was inhibited by H2O2 by 35%, DDC by 80% and 2-mercaptoethanol by 15%. The antigenic determinants of SOD isolated from boar seminal plasma and spermatozoa were similar to each other. Susceptibility of spermatozoa to cold shock increased during storage, but the differences between extenders were statistically non-significant. Copyright © 2013 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  4. A technical assessment of the porcine ejaculated spermatozoa for a sperm-specific RNA-seq analysis.

    Science.gov (United States)

    Gòdia, Marta; Mayer, Fabiana Quoos; Nafissi, Julieta; Castelló, Anna; Rodríguez-Gil, Joan Enric; Sánchez, Armand; Clop, Alex

    2018-04-26

    The study of the boar sperm transcriptome by RNA-seq can provide relevant information on sperm quality and fertility and might contribute to animal breeding strategies. However, the analysis of the spermatozoa RNA is challenging as these cells harbor very low amounts of highly fragmented RNA, and the ejaculates also contain other cell types with larger amounts of non-fragmented RNA. Here, we describe a strategy for a successful boar sperm purification, RNA extraction and RNA-seq library preparation. Using these approaches our objectives were: (i) to evaluate the sperm recovery rate (SRR) after boar spermatozoa purification by density centrifugation using the non-porcine-specific commercial reagent BoviPure TM ; (ii) to assess the correlation between SRR and sperm quality characteristics; (iii) to evaluate the relationship between sperm cell RNA load and sperm quality traits and (iv) to compare different library preparation kits for both total RNA-seq (SMARTer Universal Low Input RNA and TruSeq RNA Library Prep kit) and small RNA-seq (NEBNext Small RNA and TailorMix miRNA Sample Prep v2) for high-throughput sequencing. Our results show that pig SRR (~22%) is lower than in other mammalian species and that it is not significantly dependent of the sperm quality parameters analyzed in our study. Moreover, no relationship between the RNA yield per sperm cell and sperm phenotypes was found. We compared a RNA-seq library preparation kit optimized for low amounts of fragmented RNA with a standard kit designed for high amount and quality of input RNA and found that for sperm, a protocol designed to work on low-quality RNA is essential. We also compared two small RNA-seq kits and did not find substantial differences in their performance. We propose the methodological workflow described for the RNA-seq screening of the boar spermatozoa transcriptome. FPKM: fragments per kilobase of transcript per million mapped reads; KRT1: keratin 1; miRNA: micro-RNA; miscRNA: miscellaneous

  5. Patients with acephalic spermatozoa syndrome linked to SUN5 mutations have a favorable pregnancy outcome from ICSI.

    Science.gov (United States)

    Fang, Jianzheng; Zhang, Jingjing; Zhu, Fuxi; Yang, Xiaoyu; Cui, Yugui; Liu, Jiayin

    2018-01-10

    Are Sad1 and UNC84 domain containing 5 (SUN5) mutations associated with the outcomes of ICSI in patients with acephalic spermatozoa syndrome (ASS)? Despite highly abnormal sperm morphology, ASS patients with SUN5 mutations have a favorable pregnancy outcome following ICSI. ASS is a rare cause of infertility characterized by the production of a majority of headless spermatozoa, along with a small proportion of intact spermatozoa with an abnormal head-tail junction. Previous studies have demonstrated that SUN5 mutations may cause ASS. Several studies showed that ICSI could help patients with ASS father children. This retrospective cohort study included 11 infertile ASS males with SUN5 mutations. Five of them underwent five ICSI cycles. Their ICSI results were compared to men with ASS without SUN5 mutations (n = 3) and to men with multiple morphological abnormalities of the sperm flagella (MMAF) (n = 9). All ICSI treatments were completed between Jan 2011 and May 2017. Sanger DNA sequencing was used to detect mutations in SUN5. Clinical and biological data were collected from patients at the fertility center. Sanger sequencing validated 11 patients with SUN5 mutations. Three novel mutations in SUN5 (c.829C>T [p.Q277*]; c.1067G>A [p.R356H]; c.211+1 insGT [p.S71Cfs11*]) were identified in three patients. The rates of fertilization, good-quality embryos and pregnancy for five patients with SUN5 mutations following ICSI were 81.5%, 81.8% and 100%, respectively. The rates of fertilization and good-quality embryos in patients with MMAF were significantly lower compared with ASS patients (65.6 versus 82.4%, P = 0.039 and 53.6 versus 85.2%, P = 0.031, respectively). There were no differences in ICSI results between ASS patients with and without SUN5 mutations. Only a small number patients with SUN5 mutations was available because of its rare incidence. Patients with ASS can be effectively treated with ICSI. SUN5 mutations may be one of the genetic causes of ASS. This study

  6. High-fat diet reprograms the epigenome of rat spermatozoa and transgenerationally affects metabolism of the offspring

    DEFF Research Database (Denmark)

    de Castro Barbosa, Thais; Ingerslev, Lars R; Alm, Petter S

    2016-01-01

    OBJECTIVES: Chronic and high consumption of fat constitutes an environmental stress that leads to metabolic diseases. We hypothesized that high-fat diet (HFD) transgenerationally remodels the epigenome of spermatozoa and metabolism of the offspring. METHODS: F0-male rats fed either HFD or chow diet......1 male offspring showed common DNA methylation and small non-coding RNA expression signatures. Altered expression of sperm miRNA let-7c was passed down to metabolic tissues of the offspring, inducing a transcriptomic shift of the let-7c predicted targets. CONCLUSION: Our results provide insight...... into mechanisms by which HFD transgenerationally reprograms the epigenome of sperm cells, thereby affecting metabolic tissues of offspring throughout two generations....

  7. Radioiodination of surface proteins of bull spermatozoa and their characterization by sodium dodecyl sulphate-polyacrylamide gel electrophoresis

    International Nuclear Information System (INIS)

    Vierula, M.

    1980-01-01

    Surface proteins of ejaculated bull spermatozoa were radioiodinated using Ma 125 I, solubilized and characterized by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The electron microscopic autoradiographs showed that the labelling was equally distributed to all parts of the spermatozoon and restricted to the sperm surface. The electrophoresis of solubilized radioactivity revealed 6 radioactive fractions with approximate molecular weights of 67 000-69 000, 47 000-50 000, 34 000-37 000, 25 000-28 000 and 14 000-16 000. The 6th fraction probably represented labelled lipids. The electrophoresis of radioiodinated seminal plasma proteins revealed only 2 radioactive protein peaks which coincided with the sperm surface protein fractions IV and V. (author)

  8. Bilateral nekrose af caput femoris efter brug af arbejdssele

    DEFF Research Database (Denmark)

    Husted, Henrik; Nielsen, Reno E Tilgreen; Linnet, Jan A Rørgaard

    2002-01-01

    The case of a 42-year old man, who developed bilateral necrosis of the femoral head from traumatic causes is described. He hung in stirrups, working 4-6 hours a day decorating for three weeks, during which he complained of numbness, fatigue and radiating pain in both legs. X-rays and a bone scan...

  9. Proksimal bilateral humerusfraktur med posterior luksation af caput

    DEFF Research Database (Denmark)

    Kristensen, W; Osther, P

    1990-01-01

    . The mechanism of the injury is described and the treatment discussed. Regardless of the method of primary reposition employed, no difference in the subsequent joint function appears to be observed. Nevertheless, recognition of the condition and early reduction are of importance for the result....

  10. Utilization of fluorescent probe association for simultaneous assessment of plasmatic, acrosomal, and mitochondrial membranes of rooster spermatozoa

    Directory of Open Access Journals (Sweden)

    ECC Celeghini

    2007-09-01

    Full Text Available This experiment was designed with the objective of developing a simple, practical, and high repeatability technique for the simultaneous evaluation of the integrity of the plasmatic and acrosomal membranes, as well as funcional mitochondria of domestic fowl spermatozoa using an association of fluorescent probes. Four ejaculates (motility > 80% and abnormal morphology < 10% from each of six Ross male broiler breeder (n=24 were diluted in TALP sperm medium (25x10(6 spermatozoa/mL and split into two aliquots, and one of these aliquots was flash frozen in liquid nitrogen and thawed to damage all cellular membranes. Three treatments were prepared from these aliquots, with the following ratios of Fresh semen:Flash frozen semen: 100:0 (T100, 50:50 (T50, and 0:100 (T0. A 150-µL aliquot of diluted semen was placed in a microcentrifuge tube with the addition of 2-µL PI, 2-µL MITO, and 50-µL FITC-PSA, and incubated at 38.5º C/8 min in the dark. An 8-µL sample was placed on a slide, coverslipped, and examined by epifluorescence microscopy. Each sample had 200 cells counted and classified based on the fluorescence emitted by each probe. By regression analysis, plasma membrane integrity, as detected by PI, was determined as: v=4.17+0.82X (R²=0.95. Acrosome integrity, as detected by FITC-PSA, generated the equation: v=4.19+0.84X (R²=0.96. Functional mitochondria was estimated by the equation v=3.20+0.83X (R²=0.96. This is an efficient technique to simultaneously evaluate plasmatic, acrosomal, and mitochondrial membranes in fowl sperm. It is suggested that its application in flow cytometry systems allows this methodology to be applied in large scale.

  11. Impact of estrogenic compounds on DNA integrity in human spermatozoa: Evidence for cross-linking and redox cycling activities

    International Nuclear Information System (INIS)

    Bennetts, L.E.; De Iuliis, G.N.; Nixon, B.; Kime, M.; Zelski, K.; McVicar, C.M.; Lewis, S.E.; Aitken, R.J.

    2008-01-01

    A great deal of circumstantial evidence has linked DNA damage in human spermatozoa with adverse reproductive outcomes including reduced fertility and high rates of miscarriage. Although oxidative stress is thought to make a significant contribution to DNA damage in the male germ line, the factors responsible for creating this stress have not been elucidated. One group of compounds that are thought to be active in this context are the estrogens, either generated as a result of the endogenous metabolism of androgens within the male reproductive tract or gaining access to the latter as a consequence of environmental exposure. In this study, a wide variety of estrogenic compounds were assessed for their direct effects on human spermatozoa in vitro. DNA integrity was assessed using the Comet and TUNEL assays, lesion frequencies were quantified by QPCR using targets within the mitochondrial and nuclear (β-globin) genomes, DNA adducts were characterized by mass spectrometry and redox activity was monitored using dihydroethidium (DHE) as the probe. Of the estrogenic and estrogen analogue compounds evaluated, catechol estrogens, quercetin, diethylstilbestrol and pyrocatechol stimulated intense redox activity while genistein was only active at the highest doses tested. Other estrogens and estrogen analogues, such as 17β-estradiol, nonylphenol, bisphenol A and 2,3-dihydroxynaphthalene were inactive. Estrogen-induced redox activity was associated with a dramatic loss of motility and, in the case of 2-hydroxyestradiol, the induction of significant DNA fragmentation. Mass spectrometry also indicated that catechol estrogens were capable of forming dimers that can cross-link the densely packed DNA strands in sperm chromatin, impairing nuclear decondensation. These results highlight the potential importance of estrogenic compounds in creating oxidative stress and DNA damage in the male germ line and suggest that further exploration of these compounds in the aetiology of male

  12. Effects of Aging and Oxidative Stress on Spermatozoa of Superoxide-Dismutase 1- and Catalase-Null Mice.

    Science.gov (United States)

    Selvaratnam, Johanna S; Robaire, Bernard

    2016-09-01

    Advanced paternal age is linked to complications in pregnancy and genetic diseases in offspring. Aging results in excess reactive oxygen species (ROS) and DNA damage in spermatozoa; this damage can be transmitted to progeny with detrimental consequences. Although there is a loss of antioxidants with aging, the impact on aging male germ cells of the complete absence of either catalase (CAT) or superoxide dismutase 1 (SOD1) has not been investigated. We used CAT-null (Cat(-/-)) and SOD1-null (Sod(-/-)) mice to determine whether loss of these antioxidants increases germ cell susceptibility to redox dysfunction with aging. Aging reduced fertility and the numbers of Sertoli and germ cells in all mice. Aged Sod(-/-) mice displayed an increased loss of fertility compared to aged wild-type mice. Treatment with the pro-oxidant SIN-10 increased ROS in spermatocytes of aged wild-type and Sod(-/-) mice, while aged Cat(-/-) mice were able to neutralize this ROS. The antioxidant peroxiredoxin 1 (PRDX1) increased with age in wild-type and Cat(-/-) mice but was consistently low in young and aged Sod(-/-) mice. DNA damage and repair markers (γ-H2AX and 53BP1) were reduced with aging and lower in young Sod(-/-) and Cat(-/-) mice. Colocalization of γ-H2AX and 53BP1 suggested active repair in young wild-type mice but reduced in young Cat(-/-) and in Sod(-/-) mice and with age. Oxidative DNA damage (8-oxodG) increased in young Sod(-/-) mice and with age in all mice. These studies show that aged Sod(-/-) mice display severe redox dysfunction, while wild-type and Cat(-/-) mice have compensatory mechanisms to partially alleviate oxidative stress and reduce age-related DNA damage in spermatozoa. Thus, SOD1 but not CAT is critical to the maintenance of germ cell quality with aging. © 2016 by the Society for the Study of Reproduction, Inc.

  13. Impact of estrogenic compounds on DNA integrity in human spermatozoa: Evidence for cross-linking and redox cycling activities

    Energy Technology Data Exchange (ETDEWEB)

    Bennetts, L.E.; De Iuliis, G.N.; Nixon, B.; Kime, M.; Zelski, K. [ARC Centre of Excellence in Biotechnology and Development and Discipline of Biological Sciences, University of Newcastle, NSW (Australia); McVicar, C.M.; Lewis, S.E. [Obstetrics and Gynaecology, Queen' s University, Belfast (United Kingdom); Aitken, R.J. [ARC Centre of Excellence in Biotechnology and Development and Discipline of Biological Sciences, University of Newcastle, NSW (Australia)], E-mail: jaitken@mail.newcastle.edu.au

    2008-05-10

    A great deal of circumstantial evidence has linked DNA damage in human spermatozoa with adverse reproductive outcomes including reduced fertility and high rates of miscarriage. Although oxidative stress is thought to make a significant contribution to DNA damage in the male germ line, the factors responsible for creating this stress have not been elucidated. One group of compounds that are thought to be active in this context are the estrogens, either generated as a result of the endogenous metabolism of androgens within the male reproductive tract or gaining access to the latter as a consequence of environmental exposure. In this study, a wide variety of estrogenic compounds were assessed for their direct effects on human spermatozoa in vitro. DNA integrity was assessed using the Comet and TUNEL assays, lesion frequencies were quantified by QPCR using targets within the mitochondrial and nuclear ({beta}-globin) genomes, DNA adducts were characterized by mass spectrometry and redox activity was monitored using dihydroethidium (DHE) as the probe. Of the estrogenic and estrogen analogue compounds evaluated, catechol estrogens, quercetin, diethylstilbestrol and pyrocatechol stimulated intense redox activity while genistein was only active at the highest doses tested. Other estrogens and estrogen analogues, such as 17{beta}-estradiol, nonylphenol, bisphenol A and 2,3-dihydroxynaphthalene were inactive. Estrogen-induced redox activity was associated with a dramatic loss of motility and, in the case of 2-hydroxyestradiol, the induction of significant DNA fragmentation. Mass spectrometry also indicated that catechol estrogens were capable of forming dimers that can cross-link the densely packed DNA strands in sperm chromatin, impairing nuclear decondensation. These results highlight the potential importance of estrogenic compounds in creating oxidative stress and DNA damage in the male germ line and suggest that further exploration of these compounds in the aetiology of

  14. Effect of Priangan ram seminal plasma on viability of Peranakan Etawah buck spermatozoa preserved at 3–5oC

    Directory of Open Access Journals (Sweden)

    Muhammad Rizal

    2008-03-01

    Full Text Available In processing of buck semen, seminal plasma is a problem because it contains a phospholipase A enzime produced by the Cowper gland. If this enzime interacts with egg yolk, it causes semen coagulation, and consequently death of spermatozoa. The purpose of this research was to examine the effect of Priangan ram seminal plasma on viability of Peranakan Etawah (PE buck spermatozoa preserved at 3–5oC. Semen was collected using artificial vagina once a week. Fresh semen was divided into three tubes then centrifuged at 3,000 RPM for 30 min. Supernatant of the first tube was mixed again with Pasteur pipette (treatment A or control. Supernatant of the second tube was removed (treatment B or without seminal plasma. Supernatant of the third tube was removed and changed with Priangan ram seminal plasma in the same volume (treatment C. Semen was diluted with Tris extender containing 20% egg yolk and stored in refrigerator at 3–5oC. Quality of diluted-semen including percentages of motile spermatozoa (MS, live spermatozoa (LS, and intact plasma membrane (IPM was evaluated every day during storage at 3–5oC for three days. Results of this study showed that mean volume, colour, consistency, pH, mass activity, spermatozoa concentration, MS, LS, spermatozoa abnormal, and IPM of PE buck fresh semen, respectively was 0.68 ml, cream, thick, 7, ++/+++, 4,148.57 million cell/ml, 70%, 83.89%, 7.12% and 84%. At day-4 of storage, percentages of MS, LS, and IPM for treatment C (40, 52.2 and 51.6% was significantly (P<0.05 higher than that of: treatment B (31, 44.8 and 45.2% and treatment A (11, 15.6 and 14.8%. In conclusion, seminal plasma of Priangan ram could maintain the quality of PE buck semen preserved at 3–5oC for three days, and it prevent semen from coagulation.

  15. The use of comet assay to assess DNA integrity of boar spermatozoa following liquid preservation at 5 degrees C and 16 degrees C.

    Directory of Open Access Journals (Sweden)

    J Strzezek

    2004-03-01

    Full Text Available The comet assay, under neutral conditions, allows the assessment of DNA integrity influenced by sperm ageing, which is manifested in DNA double-strand breaks. Here, we attempted to use a modified neutral comet assay test (single-cell gel electrophoresis, to our knowledge for the first time, to assess DNA integrity of boar spermatozoa during liquid storage for 96 h at 5 degrees C and 16 degrees C. In this comet assay protocol we used 2% beta-mercaptoethanol prior to the lysis procedure, to aid in removing nuclear proteins. Ejaculates from 3 boars (designated A, C and G were diluted with a standard semen extender, Kortowo-3 (K-3, which was supplemented with lipoprotein fractions extracted from hen egg yolk (LPFh or ostrich egg yolk (LPFo. Irrespective of the extender type, the percentage of comet-detected spermatozoa with damaged DNA increased gradually during prolonged storage at 5 degrees C and 16 degrees C. Spermatozoa stored in K-3 extender exhibited elevated levels of DNA damage at both storage temperatures. Significant differences in DNA damage among the boars were more pronounced during storage in LPF-based extenders at 5 degrees C: spermatozoa of boars A and G were less susceptible to DNA damage. The percent of tail DNA in comets was lower in LPF-based extenders, and there were individual variations among the boars. We observed that changes in DNA integrity were dependent on the extender type and storage temperature. A higher level of DNA instability was observed in K-3 extended semen compared with K-3/LPFh or K-3/LPFo extended semen during storage at 5 degrees C. No significant difference in the level of DNA damage between K-3/LPFh and K-3/LPFo was observed. It seems that a long-term storage can affect genomic integrity of boar spermatozoa. The modified neutral comet assay can be used to detect low levels of DNA damage in boar spermatozoa during liquid preservation. Therefore, screening for sperm DNA damage may be used as an additional

  16. The use of comet assay to assess DNA integrity of boar spermatozoa following liquid preservation at 5 degrees C and 16 degrees C.

    Science.gov (United States)

    Fraser, L; Strzezek, J

    2004-01-01

    The comet assay, under neutral conditions, allows the assessment of DNA integrity influenced by sperm ageing, which is manifested in DNA double-strand breaks. Here, we attempted to use a modified neutral comet assay test (single-cell gel electrophoresis), to our knowledge for the first time, to assess DNA integrity of boar spermatozoa during liquid storage for 96 h at 5 degrees C and 16 degrees C. In this comet assay protocol we used 2% beta-mercaptoethanol prior to the lysis procedure, to aid in removing nuclear proteins. Ejaculates from 3 boars (designated A, C and G) were diluted with a standard semen extender, Kortowo-3 (K-3), which was supplemented with lipoprotein fractions extracted from hen egg yolk (LPFh) or ostrich egg yolk (LPFo). Irrespective of the extender type, the percentage of comet-detected spermatozoa with damaged DNA increased gradually during prolonged storage at 5 degrees C and 16 degrees C. Spermatozoa stored in K-3 extender exhibited elevated levels of DNA damage at both storage temperatures. Significant differences in DNA damage among the boars were more pronounced during storage in LPF-based extenders at 5 degrees C: spermatozoa of boars A and G were less susceptible to DNA damage. The percent of tail DNA in comets was lower in LPF-based extenders, and there were individual variations among the boars. We observed that changes in DNA integrity were dependent on the extender type and storage temperature. A higher level of DNA instability was observed in K-3 extended semen compared with K-3/LPFh or K-3/LPFo extended semen during storage at 5 degrees C. No significant difference in the level of DNA damage between K-3/LPFh and K-3/LPFo was observed. It seems that a long-term storage can affect genomic integrity of boar spermatozoa. The modified neutral comet assay can be used to detect low levels of DNA damage in boar spermatozoa during liquid preservation. Therefore, screening for sperm DNA damage may be used as an additional test of sperm

  17. The relationship of physical and chemical conditions of CEP diluent with egg yolk addition to bull spermatozoa quality before and after storage at temperaturof 4-5°C

    Science.gov (United States)

    Ducha, N.; Hariani, D.; Budijastuti, W.

    2018-01-01

    Storage of semen requires diluent to dilute semen and maintain sperm quality. One of the diluent for bull semen was CEP. The purpose of this study was to assess the association of bull spermatozoa quality with the physical and chemical conditions of CEP diluents with the addition of egg yolk before and after the storage process. The study used Limousin bull with 5 replications. The quality of spermatozoa included motility and viability. Physical and chemical conditions included the pH and osmolarity of the diluent. The motility of spermatozoa was observed under a light microscope with 200 X magnification at 37°C by two people. The viability of spermatozoa was observed under a light microscope with 400 X magnification with nigrosine eosin staining. Data were analyzed with ANOVA and continued Duncan’s test. Dilution pH was measured using pH indicator paper ranging from 6-8. The osmolarity of the diluent was measured by electrical osmolarity. The results showed that the addition of egg yolk in the CEP diluent decreased the pH and increased osmolartitas, but the quality of spermatozoa can be kept up to 8 days of storage. The conclusion in this study was the addition of egg yolk in the CEP diluent provided physical and chemical conditions that can maintain the quality of spermatozoa during storage at a temperature of 4-5 ° C.

  18. Chronic peroxisome proliferator-activated receptor gamma (PPARgamma) activation of epididymally derived white adipocyte cultures reveals a population of thermogenically competent, UCP1-containing adipocytes molecularly distinct from classic brown adipocytes

    DEFF Research Database (Denmark)

    Petrovic, Natasa; Walden, Tomas B; Shabalina, Irina G

    2009-01-01

    The recent insight that brown adipocytes and muscle cells share a common origin and in this respect are distinct from white adipocytes has spurred questions concerning the origin and molecular characteristics of the UCP1-expressing cells observed in classic white adipose tissue depots under certain...... physiological or pharmacological conditions. Examining precursors from the purest white adipose tissue depot (epididymal), we report here that chronic treatment with the peroxisome proliferator-activated receptor gamma agonist rosiglitazone promotes not only the expression of PGC-1alpha and mitochondriogenesis...... associated with classic brown adipocytes (Zic1, Lhx8, Meox2, and characteristically PRDM16) or for myocyte-associated genes (myogenin and myomirs (muscle-specific microRNAs)) and retain white fat characteristics such as Hoxc9 expression. Co-culture experiments verify that the UCP1-expressing cells...

  19. Therapeutic and fertility restoration effects of Ionidium suffruticosum on sub-fertile male albino Wistar rats: effects on testis and caudal spermatozoa.

    Science.gov (United States)

    Chenniappan, Kuppusamy; Murugan, Kadarkari

    2017-12-01

    Ionidium suffruticosum (L.) Ging (Violaceae) is an important medicinal plant widely used as a herbal traditional medicine in Ayurveda for the treatment of infertility. Currently, little pharmacological information is available on its male fertility properties following prolonged use. To investigate I. suffruticosum leaf extracts for male fertility parameters. The ethanol lyophilized fraction was administered orally on carbendazim-induced sub-fertility rats (250 mg/kg body weight for 28 days). The effects of fractions on rat's fertility parameters i.e., body and testes weight, sperm motility, sperm vitality, epididymal sperm counts, its morphology, enzyme and antioxidant stress and histopathology were studied and compared with clomiphene citrate. The sub-fertile male rats treated with I. suffruticosum leaf extract increased the body weight of 7 g, testis weight of 97 mg, increased cauda epididymal sperm counts of 34.2 × 10 6 sperm/mL, motility of sperm 46% and vitality 28% also increased and normal sperm morphology also improved up to 32%. The carbendazim-treated group showed loss in body weight of 33 g, testis weight of 851 mg, decreased epididymal sperm counts of 15 × 10 6 sperm/mL, with sluggish motility and a highly significant fall in the live sperms of about 57%. The leaf fraction of I. suffructicosum increased the testicular weight, spermatogenesis, sperm counts, lessened sperm agglutination, and increased testicular oxidative biomarkers, SOD, and CAT. This study therefore supports the usage of I. suffructicosum in traditional medicine for infertility.

  20. Expression of the vitamin D metabolizing enzyme CYP24A1 at the annulus of human spermatozoa may serve as a novel marker of semen quality

    DEFF Research Database (Denmark)

    Jensen, Martin Blomberg; Jørgensen, A; Nielsen, J E

    2012-01-01

    Vitamin D (VD) is important for male reproduction in mammals and the VD receptor (VDR) and VD-metabolizing enzymes are expressed in human spermatozoa. The VD-inactivating enzyme CYP24A1 titrates the cellular responsiveness to VD, is transcriptionally regulated by VD, and has a distinct expression...... at the sperm annulus. Here, we investigated if CYP24A1 expression serves as a marker for VD metabolism in spermatozoa, and whether CYP24A1 expression was associated with semen quality. We included 130 men (53 healthy young volunteers and 77 subfertile men) for semen analysis and immunocytochemical (ICC.......3%. Functional studies revealed that 1,25(OH)(2) D(3) increased [Ca(2+) ](i) and sperm motility in young healthy men, while 1,25(OH)(2) D(3) was unable to increase motility in subfertile patients. In conclusion, we suggest that CYP24A1 expression at the annulus may serve as a novel marker of semen quality...

  1. Birth of puppies of predetermined sex after artificial insemination with a low number of sex-sorted, frozen-thawed spermatozoa in field conditions.

    Science.gov (United States)

    Wei, Yun-Fang; Chen, Fang-Liang; Tang, Shu-Sheng; Mao, Ai-Guo; Li, Li-Guang; Cheng, Lu-Guang; Chen, Chao; Li, Fei-Xiang; Wang, Bin; Xu, Tao; Zhang, Yue-Jun; Li, Jing; Wan, Jiu-Sheng

    2017-08-01

    The aim of this study was to evaluate fertility and sex ratios after artificial insemination in dogs under field conditions. Semen was cryopreserved as unsorted (control) or was separated into X- and Y-chromosome-bearing sperm using a cell sorter. Sixty female dogs were inseminated with frozen-thawed spermatozoa of 100 × 10 6 unsorted (a dose in practice) and 4 × 10 6 sorted (X and Y group, respectively). A total of 20 dogs became pregnant and 126 puppies were born from the three groups. The percentage of parturition was similar for the X (5/20; 25.0%) and Y (4/20; 20.0%) group (P > 0.05), but lower than controls (11/20; 55.0%) (P dog spermatozoa at a farm level, making sperm-sexing technology potentially applicable for elite breeding units. © 2017 Japanese Society of Animal Science.

  2. Impact of cigarette smoking on histone (H2B) to protamine ratio in human spermatozoa and its relation to sperm parameters.

    Science.gov (United States)

    Hamad, M F; Shelko, N; Kartarius, S; Montenarh, M; Hammadeh, M E

    2014-09-01

    Smoking is strongly associated with abnormalities in histone-to-protamine transition and with alteration of protamine expression in human spermatozoa. A proper protamine to histone ratio is, however, essential for sperm chromatin maturity and DNA integrity. Alterations in these sperm nuclear proteins were observed in infertile men. The present prospective study is aimed at evaluating the possible relationship among smoking, semen quality and the histone-to-protamine transition ratio in mature spermatozoa. Histone H2B and protamine 1 (P1) and 2 (P2) were quantified using acid-urea polyacrylamide gel electrophoresis in the spermatozoa of 35 smokers and 19 non-smokers. Levels of lipid peroxidation marker malondialdehyde (MDA) were measured in seminal plasma by thiobarbituric acid assay. Cotinine concentrations were determined in seminal plasma using an enzyme-linked immunosorbent assay. Histone H2B levels in smokers (292.27 ± 58.24 ng/10(6)) were significantly higher (p = 0.001) than that of non-smokers (109.1 ± 43.70 ng/10(6)), besides, a significant difference (p > 0.0001) was found for the P1 and P2 ratio between smokers (1.71 ± 0.071) and non-smokers (1.05 ± 0.033). The H2B/(H2B+P1 + P2) ratio (0.29 ± 0.71) of smokers were significantly higher (p = sperm count, motility (p = 0.018), vitality (p = 0.009) and membrane integrity (p = 0.0001) than non-smokers. These results reveal that patients who smoke possess a higher proportion of spermatozoa with an alteration of the histone to protamine ratio than patients who do not smoke, and suggest that cigarette smoking may inversely affect male fertility. © 2014 American Society of Andrology and European Academy of Andrology.

  3. Effects of the spermatozoa: oocyte ratio, water volume and water temperature on artificial fertilization and sperm activation of cascudo-preto

    Directory of Open Access Journals (Sweden)

    Robie Allan Bombardelli

    2013-01-01

    Full Text Available The objective of this study was to evaluate the effects of water volume and water temperature on the sperm motility duration and the number of spermatozoa, and the water volume on the fertilization rates of oocytes of Rhinelepis aspera. Experiments were carried out to evaluate the effect of semen dilutions (1.74×10-5, 1.74×10-4, 1.74×10-3, 1.74×10-2, 1.74×10-1 and 1.00 mL of sperm.mL-1 of water and water temperature (5, 10, 15, 20, 25, 30, 35, 40, 45, and 50 ºC on spermatozoa motility duration. In addition, the effects of insemination dose (7×10³, 7×10(4, 7×10(5, 7×10(6 and 7×10(7 spermatozoa.oocyte-1 and water volume (1.0, 30.0, 60.0, 90.0 and 120.0 mL water.2.0 mL-1 oocytes on the artificial fertilization rates of oocytes were evaluated. The longest sperm motility duration were observed for the semen dilution of 1.74×10-5 mL semen.mL-1 water and in water at 5 ºC. The highest fertilization rates were obtained for insemination doses between 7.00×10³ and 1.23×10(7 spermatozoa. oocyte-1 and water volume of 28.11 mL water.2.0 mL-1 oocytes.

  4. The copper transporter (SLC31A1/CTR1) is expressed in bovine spermatozoa and oocytes: Copper in IVF medium improves sperm quality.

    Science.gov (United States)

    Anchordoquy, J P; Anchordoquy, J M; Pascua, A M; Nikoloff, N; Peral-García, P; Furnus, C C

    2017-07-15

    Adequate dietary intake of copper (Cu) is required for normal reproductive performance in cattle. The objective of this study was to investigate the pregnancy rates from cattle with deficient, marginal and adequate Cu plasma concentration at the beginning of artificial insemination protocol. Moreover, we determined Cu concentrations present in bovine oviductal fluid (OF), and the effects of Cu on fertilizing ability of bovine spermatozoa. Also, the presence of Cu transporter, SLC31A1 (also known as CTR1), in spermatozoa and in vitro matured oocyte were investigated. We found no differences in pregnancy rates among animals with adequate, marginal, and deficient Cu concentrations measured in plasma at the beginning of fixed-time artificial insemination (FTAI) protocol. Copper concentrations in OF were 38.3 ± 2.17 μg/dL (mean ± SEM) regardless of cupremia levels. The addition of 40 μg/dL Cu to IVF medium enhanced total and progressive motility, sperm viability, functional sperm membrane integrity (HOST), sperm-zona binding, and pronuclear formation. On the other hand, the presence of Cu in IVF medium did not modify acrosome integrity and cleavage rates after IVF, but impaired blastocyst rates. Cu transporter SLC31A1 was detected in bovine spermatozoa in the apical segment of acrosome, and in the oocyte matured in vitro. In conclusion, the results obtained in the present study determined that cupremia levels at the beginning of FTAI protocol did not influence the pregnancy rates at 60 d after insemination. The presence of CTR1 in bovine mature oocyte and spermatozoa, as well as the beneficial effect of Cu on sperm quality would suggest an important role of this mineral during the fertilization process. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Effect of commercial long-term extenders on metabolic activity and membrane integrity of boar spermatozoa stored at 17 degrees C.

    Science.gov (United States)

    Dziekońska, A; Fraser, L; Majewska, A; Lecewicz, M; Zasiadczyk, Ł; Kordan, W

    2013-01-01

    This study was aimed to analyze the metabolic activity and membrane integrity of boar spermatozoa following storage in long-term semen extenders. Boar semen was diluted with Androhep EnduraGuard (AeG), DILU-Cell (DC), SafeCell Plus (SCP) and Vitasem LD (VLD) extenders and stored for 10 days at 17 degrees C. Parameters of the analyzed sperm metabolic activity included total motility (TMOT), progressive motility (PMOT), high mitochondrial membrane potential (MMP) and ATP content, whereas those of the membrane integrity included plasma membrane integrity (PMI) and normal apical ridge (NAR) acrosome. Extender type was a significant (P semen storage. In all extenders the metabolic activity and membrane integrity of the stored spermatozoa decreased continuously over time. Among the four analyzed extenders, AeG and SCP showed the best performance in terms of TMOT and PMI on Days 5, 7 and 10 of storage. Marked differences in the proportions of spermatozoa with high MMP were observed between the extenders, particularly on Day 10 of storage. There were not any marked differences in sperm ATP content between the extenders, regardless of the storage time. Furthermore, the percentage of spermatozoa with NAR acrosomes decreased during prolonged storage, being markedly lower in DC-diluted semen compared with semen diluted with either AeG or SCP extender. The results of this study indicated that components of the long-term extenders have different effects on the sperm functionality and prolonged semen longevity by delaying the processes associated with sperm ageing during liquid storage.

  6. Effect of tributyltin on adenylate content and enzyme activities of teleost sperm: a biochemical approach to study the mechanisms of toxicant reduced spermatozoa motility.

    Science.gov (United States)

    Rurangwa, E; Biegniewska, A; Slominska, E; Skorkowski, E F; Ollevier, F

    2002-03-01

    The effects of tributyltin (TBT) on the energy metabolism and motility of fish spermatozoa were investigated in vitro in African catfish and common carp. A significant (PTBT for 24 h. Exposure of catfish spermatozoa to 2.7-27 microg/l TBT caused an instant decrease in ATP content. In the presence of 27 microg/l TBT approximately 55% of the initial ATP concentration in catfish semen was lost after 60 min incubation while AMP concentrations increased and the total adenine nucleotide (TAN) pool remained unchanged. The reduction in sperm ATP levels could not be attributed to cell death since viability decreased only slightly over the period of exposure. In carp by contrast, none of the adenylates concentrations studied (ATP, ADP and AMP) were affected by TBT exposure at any experimental condition. However, carp sperm motility was significantly reduced by exposure to 2.7 microg/l TBT. Among the enzymes investigated only lactate dehydrogenase (LDH) in catfish sperm was significantly (PTBT treatment with a reduction in activity of approximately 75%. Compared with carp sperm before TBT exposure, that of catfish had lower adenylate contents and overall lower enzymatic activities; this explains its slower sperm velocity and shorter duration of movement as measured by computer assisted sperm analysis (CASA). The present in vitro study shows that catfish spermatozoa are more sensitive to TBT exposure (and probably to other toxicants) than those of carp.

  7. l-Cysteine improves antioxidant enzyme activity, post-thaw quality and fertility of Nili-Ravi buffalo (Bubalus bubalis) bull spermatozoa.

    Science.gov (United States)

    Iqbal, S; Riaz, A; Andrabi, S M H; Shahzad, Q; Durrani, A Z; Ahmad, N

    2016-11-01

    The effects of l-cysteine in extender on antioxidant enzymes profile during cryopreservation, post-thaw quality parameters and in vivo fertility of Nili-Ravi buffalo bull spermatozoa were studied. Semen samples from 4 buffalo bulls were diluted in Tris-citric acid-based extender having different concentrations of l-cysteine (0.0, 0.5, 1.0, 2.0 and 3.0 mm) and frozen in 0.5-ml French straws. The antioxidative enzymes [catalase, super oxide dismutase and total glutathione (peroxidase and reductase)] were significantly higher (P l-cysteine as compared to other groups. Post-thaw total motility (%), progressive motility (%), rapid velocity (%), average path velocity (μm s -1 ), straight line velocity (μm s -1 ), curvilinear velocity (μm s -1 ), beat cross frequency (Hz), viable spermatozoa with intact plasmalemma (%), acrosome and DNA integrity (%) were higher with the addition of 2.0 mm l-cysteine as compared to other groups (P l-cysteine than in the control. In conclusion, the addition of 2.0 mm l-cysteine in extender improved the antioxidant enzymes profile, post-thaw quality and in vivo fertility of Nili-Ravi buffalo bull spermatozoa. © 2016 Blackwell Verlag GmbH.

  8. Analysis of sperm quality in recombinant inbred mouse strains: correlation of sperm head shape with sperm abnormalities and with the incidence of supplementary spermatozoa in the perivitelline space.

    Science.gov (United States)

    Krzanowska, H; Styrna, J; Wabik-Sliz, B

    1995-07-01

    Recombinant inbred strains were developed from reciprocal crosses between two inbred strains of mice (CBA and KE) differing in sperm head shape, proportion of normal sperm heads (CBA, 95%; KE, 78%) and fertilization efficiency (CBA, 100% of fertilized ova; KE, 72%), to determine whether the indices of sperm morphology and function were correlated. The following parameters were analysed in recombinant inbred and progenitor strains: index of sperm head shape (head width in the middle of its length/head length), percentage of abnormal sperm heads, percentage of spermatozoa with progressive movements, efficiency of penetration of hyaluronic acid polymer (Sperm Select) and percentage of fertilized ova after mating males from the tested strains with females from an outbred stock. For each investigated character, recombinant inbred strains, recombinant inbred EXCB and CBXE, could be divided into at least three categories: KE-like, CBA-like and intermediate, suggesting that in each case a minimum of two genes was involved. Recombinant strains derived from the reciprocal crosses of progenitor strains differed only with respect to the proportion of abnormal sperm heads, showing the involvement of the Y chromosome in determining this character. Penetration into Sperm Select was significantly correlated both with fertilization efficiency and sperm motility, while correlation with the proportion of normal spermatozoa did not reach the level of significance. However, there was a significant negative correlation of both sperm abnormalities and the incidence of supplementary spermatozoa in the perivitelline space with the index of sperm head shape.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Evolutionary morphology of the male reproductive system, spermatozoa and seminal fluid of spiders (Araneae, Arachnida)--current knowledge and future directions.

    Science.gov (United States)

    Michalik, Peter; Ramírez, Martín J

    2014-07-01

    The male reproductive system and spermatozoa of spiders are known for their high structural diversity. Spider spermatozoa are flagellate and males transfer them to females in a coiled and encapsulated state using their modified pedipalps. Here, we provide a detailed overview of the present state of knowledge of the primary male reproductive system, sperm morphology and the structural diversity of seminal fluids with a focus on functional and evolutionary implications. Secondly, we conceptualized characters for the male genital system, spermiogenesis and spermatozoa for the first time based on published and new data. In total, we scored 40 characters for 129 species from 56 families representing all main spider clades. We obtained synapomorphies for several taxa including Opisthothelae, Araneomorphae, Dysderoidea, Scytodoidea, Telemidae, Linyphioidea, Mimetidae, Synotaxidae and the Divided Cribellum Clade. Furthermore, we recovered synspermia as a synapomorphy for ecribellate Haplogynae and thus propose Synspermiata as new name for this clade. We hope that these data will not only contribute to future phylogenetic studies but will also stimulate much needed evolutionary studies of reproductive systems in spiders. Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. Evaluation of swine fertilisation medium (SFM) efficiency in preserving spermatozoa quality during long-term storage in comparison to four commercial swine extenders.

    Science.gov (United States)

    Fantinati, P; Zannoni, A; Bernardini, C; Forni, M; Tattini, A; Seren, E; Bacci, M L

    2009-02-01

    In pig production, artificial insemination is widely carried out and the use of fresh diluted semen is predominant. For this reason, there are increasing interests in developing new extenders and in establishing the optimal storage conditions for diluted spermatozoa. In the last few decades, we utilised a homemade diluent (swine fertilisation medium (SFM)) for spermatozoa manipulation and biotechnological application as the production of transgenic pigs utilising the sperm-mediated gene transfer technique. The purpose of the present study is therefore to analyse the ability of SFM, in comparison to four commercial extenders, in preserving the quality of diluted boar semen stored at 16.5°C till 15 days. We utilised some of the main predictive tests as objectively measured motility, acrosome and sperm membrane integrity, high mitochondrial membrane potential and pH. Based on our in vitro study, SFM could be declared as a good long-term extender, able to preserve spermatozoa quality as well as Androhep Enduraguard for up to 6 to 9 days and more.

  11. Alexandria Journal of Medicine - Vol 47, No 2 (2011)

    African Journals Online (AJOL)

    Histological study of the possible protective effect of pomegranate juice on bisphenol-A induced changes of the caput epididymal epithelium and sperms of adult albino rats · EMAIL FREE FULL TEXT EMAIL FREE FULL TEXT DOWNLOAD FULL TEXT DOWNLOAD FULL TEXT. IF El Ghazzawy, AE Meleis, EF Farghaly, A ...

  12. Browse Title Index

    African Journals Online (AJOL)

    Items 151 - 200 of 379 ... Ibrahim K. Ragab, Hala Z.E. Mohamed. Vol 47, No 2 (2011), Histological study of the possible protective effect of pomegranate juice on bisphenol-A induced changes of the caput epididymal epithelium and sperms of adult albino rats, Abstract PDF. IF El Ghazzawy, AE Meleis, EF Farghaly, A Solaiman.

  13. Combinations of glycerol percent, glycerol equilibration time, and thawing rate upon freezability of bull spermatozoa in plastic straws.

    Science.gov (United States)

    Wiggin, H B; Almquist, J O

    1975-03-01

    Twelve ejaculates were used in a central composite experiment to test 15 combinations of glycerol (7, 9, 11, 13, or 15%), glycerol equilibration times (1, 2, 4, 8, or 16 h) and thawing rates (water at 35 C for 15 s, 50 C for 13 s, 65 C for 11 s, 80 C for 9 s, or 95 C for 7 s). Semen was diluted in heated skim milk-glycerol, packaged in .3-ml. Continental U.S. straws and frozen in liquid nitrogen vapor. Based on post-thaw progressive sperm motility after storage at -196 C for 9 to 11 days, estimated optima from multiple regression were 10.7% for glycerol, 2.0 h for glycerol equilibration time, and 76 C for thawing bath temperature. Only the linear effect for each variable was significant. Much faster thawing rates and shorter glycerol equilibration times than those for freezing bull spermatozoa in glass ampules should be used for maximum post-thaw sperm motility in straws.

  14. Analysis of DNA damage in sea lamprey (Petromyzon marinus) spermatozoa by UV, hydrogen peroxide, and the toxicant bisazir

    International Nuclear Information System (INIS)

    Ciereszko, Andrzej; Wolfe, Tobie D.; Dabrowski, Konrad

    2005-01-01

    In this study we sought to demonstrate that Comet assay can be applied to sea lamprey sperm DNA fragmentation and used to describe the relationship between sperm DNA damage and sperm fertilizing ability. We show that the assay can be used reliably and accurately, and unlike in the case of mammals, there is no need for additional steps related to improvement of efficacy of lysis and DNA decondensation. This agrees with the presence of histone proteins in lamprey sperm. An increase in DNA fragmentation was noted during short-term storage of milt on ice (0-4 days). We demonstrated genotoxic effects of UV radiation and oxidative stress (exposure to hydrogen peroxide) and found that oxidative damage to sperm DNA was likely repaired after fertilization in the embryo. Repairing capacity of the oocyte toward sperm DNA lesions caused by UV was restricted. Toxic effect of p,p-bis-(1-aziridinyl)-N-methylphosphinothioic acid (p,p-bis(1-aziridinyl)-N-methylphosphinothioic amide), a sea lamprey chemosterilant, could not be linked to DNA fragmentation in the in vitro tests. Its genotoxicity in vivo may possibly be associated with other mechanisms of DNA degradation (oxidation or DNA-protein and DNA-DNA cross-linking). In conclusion, this study demonstrates that Comet assay can be successfully applied to monitor effects of environmental disturbances and imposed injuries in sea lamprey spermatozoa and possibly other species of ancient fish with acrosomal sperm

  15. Analysis of DNA damage in sea lamprey (Petromyzon marinus) spermatozoa by UV, hydrogen peroxide, and the toxicant bisazir

    Energy Technology Data Exchange (ETDEWEB)

    Ciereszko, Andrzej [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States); Semen Biology Group, Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, 10-747 Olsztyn (Poland); Wolfe, Tobie D. [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States); Dabrowski, Konrad [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States)]. E-mail: dabrowski.1@osu.edu

    2005-06-15

    In this study we sought to demonstrate that Comet assay can be applied to sea lamprey sperm DNA fragmentation and used to describe the relationship between sperm DNA damage and sperm fertilizing ability. We show that the assay can be used reliably and accurately, and unlike in the case of mammals, there is no need for additional steps related to improvement of efficacy of lysis and DNA decondensation. This agrees with the presence of histone proteins in lamprey sperm. An increase in DNA fragmentation was noted during short-term storage of milt on ice (0-4 days). We demonstrated genotoxic effects of UV radiation and oxidative stress (exposure to hydrogen peroxide) and found that oxidative damage to sperm DNA was likely repaired after fertilization in the embryo. Repairing capacity of the oocyte toward sperm DNA lesions caused by UV was restricted. Toxic effect of p,p-bis-(1-aziridinyl)-N-methylphosphinothioic acid (p,p-bis(1-aziridinyl)-N-methylphosphinothioic amide), a sea lamprey chemosterilant, could not be linked to DNA fragmentation in the in vitro tests. Its genotoxicity in vivo may possibly be associated with other mechanisms of DNA degradation (oxidation or DNA-protein and DNA-DNA cross-linking). In conclusion, this study demonstrates that Comet assay can be successfully applied to monitor effects of environmental disturbances and imposed injuries in sea lamprey spermatozoa and possibly other species of ancient fish with acrosomal sperm.

  16. Egg yolk and glycerol requirements for freezing boar spermatozoa treated with methyl β-cyclodextrin or cholesterol-loaded cyclodextrin.

    Science.gov (United States)

    Blanch, Eva; Tomás, Cristina; Hernández, Marta; Roca, Jordi; Martínez, Emilio A; Vázquez, Juan M; Mocé, Eva

    2014-04-24

    Egg yolk (EY) and glycerol are common constituents of extenders used for sperm cryopreservation. It has been demonstrated that using cholesterol-loaded cyclodextrins (CLC) improves sperm cryosurvival in several species. However, standard freezing extenders might not be the most appropriate for CLC-treated sperm. This study evaluated the EY and glycerol requirements for freezing CLC-treated boar spermatozoa. Semen samples from 34 ejaculates coming from 4 boars were used. Each ejaculate was split into three aliquots: one was used untreated (control), and the other two were treated with 1 mg of CLC or methyl-β-cyclodextrin/120 × 10(6) sperm for 15 min at 22 C prior to cryopreservation. Our results indicated that reducing the concentration of EY was detrimental for sperm viability after thawing (31.57 ± 2 vs. 19.89% ± 2 for 20 and 10% EY, respectively; P semen treated with CLC. On the other hand, it was observed that the traditional concentration of glycerol (3%) was not the appropriate for freezing CLC-treated sperm (61.10 ± 3 vs. 47.87% ± 3 viable sperm for control and CLC-treated sperm, respectively; P extenders for CLC-treated sperm. Nevertheless, additional studies will be needed to evaluate alternative cryoprotectants and to determine the effect of high glycerol concentrations on sperm functionality.

  17. Assessment of Chromatin Maturity in Human Spermatozoa: Useful Aniline Blue Assay for Routine Diagnosis of Male Infertility

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    Afifa Sellami

    2013-01-01

    Full Text Available During spermatogenesis, sperm chromatin undergoes structural changes and results in a high condensation. This nuclear compaction would be useful as a predictor of sperm fertilization capacity and pregnancy outcome. We purpose to evaluate firstly the relationship among chromatin maturity assessed by aniline blue staining (AB and the semen parameters in infertile men. Secondly, we analyzed whether the sperm gradient density centrifugation is effective to select mature spermatozoa. Fifty-one ejaculates were investigated by semen analysis and stained for chromatin condensation with AB to distinguish between unstained mature sperm and stained immature sperm. AB was applied also on 12 ejaculates which proceeded by density gradient centrifugation to compare the rates of immature sperm before and after selection. Neat semen were divided into two groups: G1 (: immature sperm <20% and G2 (: immature sperm ≥20%. No significant differences were detected in sperm concentration, motility, and normal morphology between G1 and G2. However, the rates of some morphology abnormalities were higher in G2: head abnormalities ( and microcephalic sperm (. We founded significant correlation between sperm immaturity and acrosome abnormalities (; . Sperm selection has significantly reduced the rates of immature sperm. A better understanding of chromatin structure and its impact on the sperm potential is needed to explore male infertility.

  18. Effect of dietary selenium and vitamin E on the ultrastructure and ATP concentration of boar spermatozoa, and the efficacy of added sodium selenite in extended semen on sperm motility.

    Science.gov (United States)

    Marin-Guzman, J; Mahan, D C; Whitmoyer, R

    2000-06-01

    Three experiments evaluated the effects of dietary Se and vitamin E on the ultrastructure of spermatozoa, ATP concentration of spermatozoa, and the effects of adding sodium selenite to semen extenders on subsequent sperm motility. The experiment was a 2 x 2 arrangement of treatments in a randomized complete block design. A total of 10 mature boars were fed from weaning to 18 mo of age diets fortified with two levels of supplemental Se (0 or .5 ppm) or vitamin E (0 or 220 IU/kg diet). The nonfortified diets contained .06 ppm Se and 4.4 IU vitamin E/kg. In Exp. 1, the spermatozoa from all boars were examined by electron microscopy. Vitamin E had no effect on structural abnormalities in the spermatozoa. When the low-Se diet was fed the acrosome or nuclei of the spermatozoa was unaffected, but the mitochondria in the tail midpiece were more oval with wider gaps between organelles. The plasma membrane connection to the tail midpiece was not tightly bound as when boars were fed Se. Immature spermatozoa with cytoplasmic droplets were more numerous when boars were fed the low-Se diet, but the occurrence of midpiece abnormalities occurred in boars fed diets with or without Se or vitamin E. Our results suggest that Se may enhance spermatozoa maturation in the epididymis and may reduce the number of sperm with cytoplasmic droplets. In Exp. 2, the concentration of ATP in the spermatozoa was evaluated in the semen of all treatment boars. When the low-Se diet was fed, ATP concentration was lower (P boar semen with a semen extender with sodium selenite added at 0, .3, .6, or .9 ppm Se. Three ejaculates from each boar were used to evaluate these effects on sperm motility to 48 h after dilution. Sperm motility declined (P extender, and this decline was exacerbated as the concentration of added Se increased (P boars resulted in abnormal spermatozoal mitochondria, a lower ATP concentration in the spermatozoa, and a loose apposition of the plasma membrane to the helical coil of the

  19. In vivo sodium, potassium, and sperm concentrations in the rat epididymis.

    Science.gov (United States)

    Turner, T T; Hartmann, P K; Howards, S S

    1977-02-01

    In vivo samples of epididymal fluids were obtained through the use of micropuncture techniques. Microsamples from four areas of the rat epididymis were analyzed for Na+ and K+ concentrations and for sperm density. Na+ values declined significantly from caput to corpus epididymidis (P less than 0.01), while K+ and sperm concentrations increased significantly (P less than 0.01). A large water loss from the epididymal lumen was calculated, as well as net losses of both cations. Water losses may be explained on the basis of an active Na+ pump; however, the effect of the absolute values of epididymal Na+ and K+ concentrations on sperm motility and fertility remains unresolved.

  20. Morphological alterations in cryopreserved spermatozoa of scallop Argopecten purpuratus Alteraciones morfológicas en espermatozoides criopreservados de concha de abanico Argopecten purpuratus

    Directory of Open Access Journals (Sweden)

    Carlos Espinoza

    2010-01-01

    Full Text Available The present work identifies and quantifies the morphological alterations of scallop Argopecten purpuratus spermatozoa caused by long-term cryopreservation. Percentages of motility, fertilization and injured spermatozoa were quantified by optic microscopy and scanned electron microscopy. These parameters were evaluated in sperm without treatment (CTR, spermatozoa incubated in cryoprotective solution but not freezed (ICS and freezed-thawed spermatozoa (FTS. Spermatozoa of ICS treatment remained motile longer than those of CTR, whereas those of FTS treatment were lowest. Morphology of the spermatozoa was affected in several ways by the freeze-thawing treatment; some had their head deformed or swollen, others had their cell membrane folded or broken; acrosome reaction; anomalous positions or absence of mitochondria as well as broken, stiff or loss of lineal structure of tail. CTR and ICS treatments had higher percentages of undamaged sperm (87.7% and 79.0% respectively, while FTS samples had 14.2% of undamaged sperm. The tail was the spermatic structure most commonly injured in FTS (77.0%, the percentage of sperm with head injury was 55.1% and with acrosome reaction was 28.7%, whereas middle piece was affected in 23.9% of sperm. Percentages of fertilization were 68.3%, 67.9% and 58.2% for CTR, ICS and FTS respectively, which were not significantly different. There was a higher correlation between injuries and motility than between injuries and fertilization success. Correlation between motility and fertilization was low (0.605 and 0.668 with motility at 5 and 30 min, respectively.El presente trabajo identifica y cuantifica las alteraciones morfológicas en espermatozoides de concha de abanico A. purpuratus causadas por la criopreservación en nitrógeno líquido. Porcentajes de motilidad, fecundación de ovocitos frescos y espermatozoides lesionados (en cabeza, acrosoma, pieza media y flagelo fueron determinados bajo microscopía óptica y electr

  1. Altered PIWI-LIKE 1 and PIWI-LIKE 2 mRNA expression in ejaculated spermatozoa of men with impaired sperm characteristics.

    Science.gov (United States)

    Giebler, Maria; Greither, Thomas; Müller, Lisa; Mösinger, Carina; Behre, Hermann M

    2018-01-01

    In about half the cases of involuntary childlessness, a male infertility factor is involved. The PIWI-LIKE genes, a subclade of the Argonaute protein family, are involved in RNA silencing and transposon control in the germline. Knockout of murine Piwi-like 1 and 2 homologs results in complete infertility in males. The aim of this study was to analyze whether the mRNA expression of human PIWI-LIKE 1-4 genes is altered in ejaculated spermatozoa of men with impaired sperm characteristics. Ninety male participants were included in the study, among which 47 were with normozoospermia, 36 with impaired semen characteristics according to the World Health Organization (WHO) manual, 5 th edition, and 7 with azoospermia serving as negative control for the PIWI-LIKE 1-4 mRNA expression in somatic cells in the ejaculate. PIWI-LIKE 1-4 mRNA expression in the ejaculated spermatozoa of the participants was measured by quantitative real-time PCR. In nonazoospermic men, PIWI-LIKE 1-4 mRNA was measurable in ejaculated spermatozoa in different proportions. PIWI-LIKE 1 (100.0%) and PIWI-LIKE 2 (49.4%) were more frequently expressed than PIWI-LIKE 3 (9.6%) and PIWI-LIKE 4 (15.7%). Furthermore, a decreased PIWI-LIKE 2 mRNA expression showed a significant correlation with a decreased sperm count (P = 0.022) and an increased PIWI-LIKE 1 mRNA expression with a decreased progressive motility (P = 0.048). PIWI-LIKE 1 and PIWI-LIKE 2 mRNA expression exhibited a significant association with impaired sperm characteristics and may be a useful candidate for the evaluation of the impact of PIWI-LIKE 1-4 mRNA expression on male infertility.

  2. Conservation science in a terrorist age: the impact of airport security screening on the viability and DNA integrity of frozen felid spermatozoa.

    Science.gov (United States)

    Gloor, Kayleen T; Winget, Doug; Swanson, William F

    2006-09-01

    In response to growing terrorism concerns, the Transportation Security Administration now requires that all checked baggage at U.S. airports be scanned through a cabinet x-ray system, which may increase risk of radiation damage to transported biologic samples and other sensitive genetic material. The objective of this study was to investigate the effect of these new airport security regulations on the viability and DNA integrity of frozen felid spermatozoa. Semen was collected from two domestic cats (Felis silvestris catus) and one fishing cat (Prionailurus viverrinus), cryopreserved in plastic freezing straws, and transferred into liquid nitrogen dry shippers for security screening. Treatment groups included frozen samples from each male scanned once or three times using a Transportation Security Administration-operated cabinet x-ray system, in addition to non-scanned samples (i.e., negative control) and samples previously scanned three times and exposed to five additional high-intensity x-ray bursts (i.e., positive control). Dosimeters placed in empty dry shippers were used to quantify radiation exposure. Following treatment, straws were thawed and spermatozoa analyzed for post-thaw motility (percentage motile and rate of progressive movement), acrosome status, and DNA integrity using single-cell gel electrophoresis (i.e., the comet assay). Dosimeter measurements determined that each airport screening procedure produced approximately 16 mrem of radiation exposure. Our results indicated that all levels of radiation exposure adversely affected (P 0.05) among treatment groups. Results also showed that the amount of double-stranded DNA damage was greater (P cat species scanned three times compared to samples scanned once or negative controls. Findings suggest that new airport security measures may cause radiation-induced damage to frozen spermatozoa and other valuable biologic samples transported on passenger aircraft and that alternative modes of sample

  3. Seminal Plasma Characteristics and Expression of ATP-binding Cassette Transporter A1 (ABCA1) in Canine Spermatozoa from Ejaculates with Good and Bad Freezability.

    Science.gov (United States)

    Schäfer-Somi, S; Palme, N

    2016-04-01

    The composition of seminal plasma and the localization of the ATP-binding cassette transporter A1 (ABCA1) in spermatozoa from good and bad freezers were compared to frozen-thawed spermatozoa from the same dog. Ejaculates were obtained from 31 stud dogs, and the sperm-rich fraction (SRF) was kept for analysis. One aliquot was used for the analysis of concentration, progressive motility (P; CASA), viability (V; CASA) and leucocyte count, and the analysis was performed by flow cytometry (FITC-PNA/PI), SCSA and HOST. In seminal plasma, concentration of albumin, cholesterol, calcium, inorganic phosphate, sodium, potassium, zinc and copper was measured. Semen smears were prepared and evaluated for the expression of ABCA1. The remainder of each ejaculate was frozen. After thawing, the quality assessment was repeated and further smears were prepared. According to post-thaw semen quality, dogs were assigned to good freezers (n = 20) or bad freezers (n = 11), the latter were defined as 40% morphologically abnormal sperm and/or < 50% viability. Bad freezers were older than good freezers (5.3 vs 3.4 years, p < 0.05). In bad freezers, the percentage of sperm with ABCA1 signal in the acrosome was lower (26.3% vs 35.7%, p < 0.01) and the percentage of sperm with complete loss of ABCA1 signal higher (46.7% vs 30%, p < 0.01); the percentage of dead spermatozoa was higher (36.1% vs 25.5%, p < 0.05), and the concentration of cholesterol and sodium in seminal plasma was lower than in good freezers (p < 0.05). We conclude that in thawed bad freezer sperm, an increase in acrosome damages coincided with an increased loss of cholesterol transporters and cell death, and a lower cholesterol concentration in seminal plasma. Follow-up studies revealed whether a relation exists between these findings. © 2016 Blackwell Verlag GmbH.

  4. Effects of the technique of cryopreservation and dilution/centrifugation after thawing on the motility and vitality of spermatozoa of oligoasthenozoospermic men.

    Science.gov (United States)

    Esteves, Sandro C; Spaine, Deborah M; Cedenho, Agnaldo P; Srougi, Miguel

    2003-01-01

    Comparing in human semen samples with low initial quality, the effects of 2 techniques of cryopreservation and dilution/centrifugation after thawing on the spermatic motility and vitality. Semen samples from 15 oligo and/or asthenozoospermic individuals assisted in the infertility sector of a tertiary hospital were obtained through masturbation. The samples were divided into 2 portions of equal volume, and diluted (1:1; v/v) with the cryoprotector containing glycerol (Test yolk buffer). One portion was frozen through the technique of liquid nitrogen vapor with static phases (group I - GI), while the other was frozen through a programmable biological freezer with linear speed (Planer, Kryo 10, series III) (group II - GII). The following parameters were assessed before freezing and after thawing: percentage of spermatozoa with progressive motility (Prog%) and percentage of live spermatozoa (Vit%). After defrosting, Prog% was assessed before and after removal of cryoprotector diluent, in different time intervals (zero, 3 h, and 24 h). The statistical analysis has been accomplished by using the non-parametric tests of Wilcoxon and Friedman. There was significant reduction of Prog% and Vit% from before freezing to after defrosting in both groups, I and II (p live spermatozoa with progressive motility. The effects of dilution and centrifugation to remove the cryoprotector had a negative impact only in samples frozen through the automated technique. In both techniques, progressive motility is kept constant during the first 3 hours after thawing and removal of the cryoprotector, but is drastically diminished by the end of an incubation period of 24 hours.

  5. Estrous cycle characterisation and artificial insemination using frozen-thawed spermatozoa in the bottlenose dolphin (Tursiops truncatus).

    Science.gov (United States)

    Robeck, T R; Steinman, K J; Yoshioka, M; Jensen, E; O'Brien, J K; Katsumata, E; Gili, C; McBain, J F; Sweeney, J; Monfort, S L

    2005-05-01

    The reproductive endocrinology of the bottlenose dolphin, Tursiops truncatus, was characterized to facilitate the development of artificial insemination using cryopreserved spermatozoa. Specific objectives were: (i) to determine the excretory dynamics of urinary luteinizing hormone (LH) and ovarian steroid metabolites during the estrous cycle; (ii) to evaluate the effect of an exogenously administered synthetic progesterone analog (altrenogest) on reproductive hormone excretion; (iii) to correlate follicular growth and ovulation (as determined by transabdominal ultrasound) to urinary LH and ovarian steroid metabolites; (iv) examine the in vivo fertilisation capacity of cryopreserved semen, and (v) to develop an intrauterine insemination technique. Based on urinary endocrine monitoring of natural estrous cycles (2 consecutive cycles) and nine post altrenogest cycles in ten females, estrous cycles were found to be 36 days long and comprised of an 8 day and 19 day follicular and luteal phase, respectively. Peak estrogen conjugates (EC; 5.4+/-3.8 ng/mg creatinine (Cr)) occurred 8 h prior to the LH surge (70.9+/-115.7 ng/mg Cr). The time of ovulation, as determined by ultrasonography, occurred 32.1+/-8.9 h and 24.3+/-7.0 h after the onset of the LH surge and LH peak, respectively. Mean preovulatory follicular diameter and circumference were 2.1+/-0.5 cm and 6.5+/-1.5 cm, respectively. Of the 27 estrous synchronisation attempts, 13 resulted in an ovulatory cycle, with ovulation occurring 21 days post-altrenogest treatment. Intrauterine (4 of 5) and intracornual (1 of 3) inseminations conducted across eight estrous cycles resulted in five pregnancies (63%), one pregnancy resulted from the use of liquid stored semen, whereas four were achieved using cryopreserved semen. These data provide new information on female bottlenose dolphin reproductive physiology, and demonstrate that the combination of endocrine monitoring and serial ultrasonography contributed to successful AI

  6. Osmotic tolerance of avian spermatozoa: Influence of time, temperature, cryoprotectant and membrane ion pump function on sperm viability

    Science.gov (United States)

    Blanco, J.M.; Long, J.A.; Gee, G.; Donoghue, A.M.; Wildt, D.E.

    2008-01-01

    Potential factors influencing sperm survival under hypertonic conditions were evaluated in the Sandhill crane (Grus canadensis) and turkey (Meleagridis gallopavo). Sperm osmotolerance (300-3000 mOsm/kg) was evaluated after: (1) equilibration times of 2, 10, 45 and 60 min at 4 ?C versus 21 ?C; (2) pre-equilibrating with dimethylacetamide (DMA) or dimethylsulfoxide (Me2SO) at either 4 ?C or 21 ?C; and (3) inhibition of the Na+/K+ and the Na+/H+ antiporter membrane ionic pumps. Sperm viability was assessed using the eosin-nigrosin live/dead stain. Species-specific differences occurred in response to hypertonic conditions with crane sperm remaining viable under extreme hypertonicity (3000 mOsm/kg), whereas turkey sperm viability was compromised with only slightly hypertonic (500 mOsm/kg) conditions. The timing of spermolysis under hypertonic conditions was also species-specific, with a shorter interval for turkey (2 min) than crane (10 min) sperm. Turkey sperm osmotolerance was slightly improved by lowering the incubation temperature from 21 to 4 ?C. Pre-equilibrating sperm with DMA reduced the incidence of hypertonic spermolysis only in the crane, at both room and refrigeration temperature. Inhibiting the Na+/K+ and the Na+/H+ antiporter membrane ion pumps did not impair resistance of crane and turkey spermatozoa to hypertonic stress; pump inhibition actually increased turkey sperm survival compared to control sperm. Results demonstrate marked species specificity in osmotolerance between crane and turkey sperm, as well as in the way temperature and time of exposure affect sperm survival under hypertonic conditions. Differences are independent of the role of osmotic pumps in these species.

  7. Aspects pertinent to the usefulness of a solid phase radio-immuno-sorbent assay for the detection of spermatozoa antibodies in sera of infertility patients

    International Nuclear Information System (INIS)

    Hinrichs-Reiche, I.

    1987-01-01

    The solid phase Radio-Immuno-Sorbent Assay (RISA) is a highly sensitive and valid test to detect 125-iodinetagged antibodies to spermatozoa that allows qualitative and quantitative evaluations of sperm-incapacitating immunglobulin Ig G in sera from patients believed to be infertile for immunological reasons. The study failed to reveal any correlations between the results of RISA and those of micro-sperm-agglutination or micro-sperm-immobilisation tests. There was a major body of evidence pointing to possible links between female isoimmunity and male autoimmunity. (TRV) [de

  8. Effect of hydrogen peroxide on the main kinetic parameters of ATP hydrolysis by ouabain sensitive Na+, K+-ATP-ase in spermatozoa of infertile men

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    Р. В. Фафула

    2017-12-01

    Full Text Available Background: It is known that Na+,K+-ATP-ase plays important role in physiology of spermatozoa including their motility. Na+,K+-ATP-ase is one of the targets for reactive oxygen species. Hyperproduction of reactive oxygen species can damage sperm cells and it is considered to be as one of the mechanisms of male infertility. Objectives: To evaluate the H2O2 effect on the main kinetic parameters of ATP hydrolysis by ouabain-sensitive Na+,K+-ATPase of spermatozoa of fertile (normozoospermia and infertility men (asthenozoospermia. Materials and methods: Na+, K+-ATP-ase activity was determined spectrophotometrically by production of Pi. Concentration dependencies ware linearized in Lineweaver-Burk plot. Results: Effective inhibitory effect of H2O2 on ouabain-sensitive Na+,K+-ATP-ase activity of sperm cells of fertile and infertile men was demonstrated. The effects of H2O2 on the main kinetic parameters of the ATP hydrolysis with the involvement of Na+, K+-ATP-ase was studied. In the whole range of studied concentrations of ATP the Na+, K+-ATP-ase activity of spermatozoa of fertile and infertile men was reduced in the presence of H2O2 in the incubation medium. However, the optimal activity of the Na+, K+-ATP-ase activity of sperm cells in both normozoospermic and asthenozoospermic men was observed in the presence of 5 mM ATP in the incubation medium. By linearization of concentration curves in Lineweaver-Burk plot the main kinetic parameters of Na+, K+-activated, Mg2+-dependent ATP hydrolysis in the sperm cells of fertile and infertile men were determined. Under the effect of H2O2, the affinity constant of enzyme to ATP in normozoospermic and asthenozoospermic men increases several times. The initial maximum rate of ATP hydrolysis was significantly reduced only in the spermatozoa of fertile men with normozoospermia. Conclusions: Under conditions of H2O2-induced oxidative stress the inhibition of ouabain-sensitive Na+,K+-ATP-ase activity in sperm cells

  9. Predictive value of serum Inhibin-B levels as an indicator of the presence of testicular spermatozoa in non-obstructive azoospermia

    Directory of Open Access Journals (Sweden)

    Marwan Alhalabi

    2016-12-01

    Full Text Available Introduction: Inhibin-B is glycoproteins of the transforming growth factor-β superfamily, is produced almost exclusively by the Sertoli cells and has been proposed as direct markers of their function and indirect markers of spermatogenesis. Our objective was to evaluate the predictive value of serum Inhibin-B levels as an indicator of the presence of testicular spermatozoa in non-obstructive azoospermia (NOA, compared with other markers such as FSH and testicular volume. Material and methods: A total of 228 patients with idiopathic non-obstructive azoospermia (NOA were recruited in this prospective study. Inhibin-B in serum has been evaluated before sperm retrieval by Enzyme-Linked Immuno-Sorbent Assay (Elisa, in addition to FSH and testosterone. Testicular volume was measured by sonography. Testicular Fine Needle Aspiration (FNA and Testicular Sperm Extraction (TESE were performed for sperm retrieval. The patients were classified according to the presence or absence of Testicular spermatozoa into two groups (I and II respectively. Results: Spermatozoa were retrieved in 87 patients (38.16%, classified as group I and were absent in 141 patients (61.84% classified as group II. Mean serum Inhibin-B ± SD was significantly higher in group I than in group II, 71.77 versus 27.49 respectively (P < 0.001. The odds ratio of Inhibin-B for success of sperm retrieval was 1.0409 (95% CI: 1.0286–1.0533. Serum Inhibin-B demonstrated that the best inhibin-B value threshold for discriminating between successful and failed sperm retrieval was 35 pg/ml, with a sensitivity of 75.86 (95% Confidence interval [CI]: 65.5–84.4 and specificity of 80.85 (CI: 73.4–87.0 for the prediction of the presence of testicular spermatozoa as determined by the receiver operating characteristic (ROC curve analysis and area under curve (AUC. Conclusions: Inhibin-B might be a predictive marker of persistent spermatogenesis in the men with non-obstructive azoospermia.

  10. Spermatozoa from the maned wolf (Chrysocyon brachyurus) display typical canid hyper-sensitivity to osmotic and freezing-induced injury, but respond favorably to dimethyl sulfoxide.

    Science.gov (United States)

    Johnson, Amy E M; Freeman, Elizabeth W; Wildt, David E; Songsasen, Nucharin

    2014-06-01

    We assessed the influences of medium osmolality, cryoprotectant and cooling and warming rate on maned wolf (Chrysocyon brachyurus) spermatozoa. Ejaculates were exposed to Ham's F10 medium (isotonic control) or to this medium plus NaCl (350-1000mOsm), sucrose (369 and 479mOsm), 1M glycerol (1086mOsm) or dimethyl sulfoxide (Me2SO, 1151mOsm) for 10 min. Each sample then was diluted back into Ham's medium and assessed for sperm motility and plasma membrane integrity. Although glycerol and Me2SO had no influence (P>0.05), NaCl and sucrose solutions affected sperm motility (P0.05) to the control. As osmolality of the NaCl solution increased, motility decreased to maned wolf spermatozoa are similar to domestic dog sperm in their sensitivity to osmotic-induced motility damage, the plasma membranes tolerate dehydration, and the cells respond favorably to Me2SO as a cryoprotectant. Published by Elsevier Inc.

  11. [A thermodynamic study on bovine spermatozoa by microcalorimetry after Percoll density-gradient centrifugation - experimental probe of its utility in andrology].

    Science.gov (United States)

    Fischer, C; Scherfer-Brähler, V; Müller-Schlösser, F; Schröder-Printzen, I; Weidner, W

    2007-05-01

    Microcalorimetric measurements can be used for recording exothermic or endothermic summation effects of a great variety of biological processes. The aim of the present study was to examine the usefullness of the microcalorimetry method to characterise the biological activity of spermatozoa. The heat flow of bovine fresh sperm as well as cryosperm samples were measured after Percoll density-gradient centrifugation in a 4-channel microcalorimeter. Various calibration times, volumes of samples and sperm concentrations were tested and analysed. Sperm concentration was recorded by a computer-assisted, computer-aided software system method (CASA). Using a calibration time of 15 minutes, the heat signal of the fresh and cryosperm samples showed a characteristic peak after 39.5 min and 38.1 min (mean), respectively, with a significant correlation to sample volume and sperm concentration (p < 0.05). For obtaining the best results, a sample volume of 1 ml and a sperm concentration of more than 50 x 10 (6)/mL was used. With microcalorimetric measurements the biological activity of spermatozoa could be recorded for reproducible results, thus opening the way to an automatised ejaculate analysis in the future. More investigations are necessary to correlate microcalorimetric parameters with semen function.

  12. On methods for the detection of reactive oxygen species generation by human spermatozoa: analysis of the cellular responses to catechol oestrogen, lipid aldehyde, menadione and arachidonic acid.

    Science.gov (United States)

    Aitken, R J; Smith, T B; Lord, T; Kuczera, L; Koppers, A J; Naumovski, N; Connaughton, H; Baker, M A; De Iuliis, G N

    2013-03-01

    Oxidative stress is known to have a major impact on human sperm function and, as a result, there is a need to develop sensitive methods for measuring reactive oxygen species (ROS) generation by these cells. A variety of techniques have been developed for this purpose including chemiluminescence (luminol and lucigenin), flow cytometry (MitoSOX Red, dihydroethidium, 4,5-diaminofluorescein diacetate and 2',7'-dichlorodihydrofluorescein diacetate) and spectrophotometry (nitroblue tetrazolium). The relative sensitivity of these assays and their comparative ability to detect ROS generated in different subcellular compartments of human spermatozoa, have not previously been investigated. To address this issue, we have compared the performance of these assays when ROS generation was triggered with a variety of reagents including 2-hydroxyestradiol, menadione, 4-hydroxynonenal and arachidonic acid. The results revealed that menadione predominantly induced release of ROS into the extracellular space where these metabolites could be readily detected by luminol-peroxidase and, to a lesser extent, 2',7'-dichlorodihydrofluorescein. However, such sensitivity to extracellular ROS meant that these assays were particularly vulnerable to interference by leucocytes. The remaining reagents predominantly elicited ROS generation by the sperm mitochondria and could be optimally detected by MitoSOX Red and DHE. Examination of spontaneous ROS generation by defective human spermatozoa revealed that MitoSOX Red was the most effective indicator of oxidative stress, thereby emphasizing the general importance of mitochondrial dysregulation in the aetiology of defective sperm function. © 2013 American Society of Andrology and European Academy of Andrology.

  13. Tin accumulation in spermatozoa of the rats exposed to tributyltin chloride by synchrotron radiation X-ray fluorescence (SR-XRF) analysis with microprobe

    International Nuclear Information System (INIS)

    Homma-Takeda, S.; Nishimura, Y.; Terada, Y.; Ueno, S.; Watanabe, Y.; Yukawa, M.

    2005-01-01

    Organotin compounds are widely used in industry and its environmental contamination by these compounds has recently become a concern. It is known that they act as endocrine disruptors but details of the dynamics of Sn in reproductive organs are still unknown. In the present study, we attempted to determine Sn distribution in the testis of rats exposed to tributyltin chloride (TBTC) by inductively coupled argon plasma-mass spectrometry (ICP-MS) for microdissectioned seminiferous tubules and cell-selective metal determination of synchrotron radiation X-ray florescence (SR-XRF) analysis. TBTC was orally administered to rats at a dose of 45 μmol/kg per day for 3 days. One day later, Sn was detected in the microdissectioned seminiferous tubules at a level approximately equivalent to that in the testis. Significant stage-specificity of Sn accumulation was not observed in the experimental model. Sn was also detected in spermatozoa at the stage VIII seminiferous tubule, which are the final step of spermatogenesis in the testis. These data indicate that Sn accumulates in germ cells as well as in spermatozoa in a short period of TBTC exposure

  14. Tin accumulation in spermatozoa of the rats exposed to tributyltin chloride by synchrotron radiation X-ray fluorescence (SR-XRF) analysis with microprobe

    Energy Technology Data Exchange (ETDEWEB)

    Homma-Takeda, S. [Environmental Radiation Protection Research Group, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku Chiba 263-8555 (Japan)]. E-mail: shino_ht@nirs.go.jp; Nishimura, Y. [Environmental Radiation Protection Research Group, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku Chiba 263-8555 (Japan); Terada, Y. [Japan Synchrotron Radiation Research Institute, Mikazuki, Hyogo 679-5198 (Japan); Ueno, S. [School of Veterinary Medicine and Animal Sciences, Kitasato University, Towada, Aomori 034-8628 (Japan); Watanabe, Y. [Environmental Radiation Protection Research Group, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku Chiba 263-8555 (Japan); Yukawa, M. [Environmental Radiation Protection Research Group, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku Chiba 263-8555 (Japan)

    2005-04-01

    Organotin compounds are widely used in industry and its environmental contamination by these compounds has recently become a concern. It is known that they act as endocrine disruptors but details of the dynamics of Sn in reproductive organs are still unknown. In the present study, we attempted to determine Sn distribution in the testis of rats exposed to tributyltin chloride (TBTC) by inductively coupled argon plasma-mass spectrometry (ICP-MS) for microdissectioned seminiferous tubules and cell-selective metal determination of synchrotron radiation X-ray florescence (SR-XRF) analysis. TBTC was orally administered to rats at a dose of 45 {mu}mol/kg per day for 3 days. One day later, Sn was detected in the microdissectioned seminiferous tubules at a level approximately equivalent to that in the testis. Significant stage-specificity of Sn accumulation was not observed in the experimental model. Sn was also detected in spermatozoa at the stage VIII seminiferous tubule, which are the final step of spermatogenesis in the testis. These data indicate that Sn accumulates in germ cells as well as in spermatozoa in a short period of TBTC exposure.

  15. Networks Models of Actin Dynamics during Spermatozoa Postejaculatory Life: A Comparison among Human-Made and Text Mining-Based Models

    Directory of Open Access Journals (Sweden)

    Nicola Bernabò

    2016-01-01

    Full Text Available Here we realized a networks-based model representing the process of actin remodelling that occurs during the acquisition of fertilizing ability of human spermatozoa (HumanMade_ActinSpermNetwork, HM_ASN. Then, we compared it with the networks provided by two different text mining tools: Agilent Literature Search (ALS and PESCADOR. As a reference, we used the data from the online repository Kyoto Encyclopaedia of Genes and Genomes (KEGG, referred to the actin dynamics in a more general biological context. We found that HM_ALS and the networks from KEGG data shared the same scale-free topology following the Barabasi-Albert model, thus suggesting that the information is spread within the network quickly and efficiently. On the contrary, the networks obtained by ALS and PESCADOR have a scale-free hierarchical architecture, which implies a different pattern of information transmission. Also, the hubs identified within the networks are different: HM_ALS and KEGG networks contain as hubs several molecules known to be involved in actin signalling; ALS was unable to find other hubs than “actin,” whereas PESCADOR gave some nonspecific result. This seems to suggest that the human-made information retrieval in the case of a specific event, such as actin dynamics in human spermatozoa, could be a reliable strategy.

  16. Quantity and quality of guinea pig (cavia porcellus) spermatozoa after administration of methanol extract of bitter melon (momordica charantia) seed and depot medroxy progesterone acetate (DMPA)

    Science.gov (United States)

    Ilyas, Syafruddin; Hutahaean, Salomo; Nursal

    2018-03-01

    The discovery of male contraceptive drugs continues to be pursued, due to the few participation of men associated with the lack of contraceptive options for men. The combination of bitter melon seed methanol extract and DMPA are the options that currently apply to men. Therefore, the use of guinea pigs as experimental animals conducted research using experimental methods with complete randomized design (CRD). There are 4 control groups and 4 treatment groups. The first group, control group of dimethyl sulphoxide (DMSO) for 0 week (K0), The second one, bitter melon seed extract of 50 mg/100g Body Weight/day for 0 week (P0), the third one, control group of dimethyl sulfoxide (DMSO) for 4 weeks (K1), the fourth one, bitter melon seed extract of 50 mg/100g BW/day for 4 weeks + Depot medroxy Progesterone Acetate (P1), the fifth one, control group of dimethyl sulfoxide (DMSO) for 8 weeks (K2), the sixth one, bitter melon seed extract of 50 mg/100g BW/day for 8 weeks + DMPA (P2), the seventh one, control group of dimethyl sulfoxide (DMSO) for 12 weeks (K3), the eighth one, bitter melon seed extract of 50 mg/100g BW/day for 12 weeks + DMPA (P3). Methanol extract of bitter melon seed to decrease the quantity and quality of guinea pig spermatozoa decreased significantly, i.e. viability and normal morphology of spermatozoa (p<0.05).

  17. Synergism between caffeine and γ-radiation in the induction of dominant lethal mutations in oocytes and spermatozoa of Musca domestica

    International Nuclear Information System (INIS)

    Targa, H.J.

    1983-01-01

    Caffeine was studied with regard to its synergism with γ-radiation in the induction of dominant lethal mutations in S14 oocytes and mature spermatozoa of M. domestica. In S14 oocytes an increase in the frequency of such a type of mutation was observed only when the exposure to γ-radiation followed a pretreatment with a diet containing 0.2% of caffeine. Negative results were obtained with (a) post-treatment with the same kind of diet, (b) pretreatment with diets containing 0.1 and 0.02% of caffeine and (c) exposure to the radiation 6 h after interruption of the feeding treatment with the diet containing 0.2% of caffeine. Such influence of the conditions under which the treatment is performed and the synergistic effects is probably related to the food intake pattern and the rapid metabolism of the caffeine. When the 0.2% caffeine pretreatment was combined with an exposure of the oocytes to variable doses of γ-radiation, the increments in the mutations observed seemed to be negatively correlated to the radiation doses used. Also, under such conditions, the dose/survival relationship fits well an exponential curve expressed by in y=-0.866chi. With mature spermatozoa, synergism by caffeine was found only when the females, after having been mated with the irradiated males, were fed for 24 h on a diet supplemented with 0.2% of caffeine. (orig.)

  18. Effect of cyclodextrin-loaded cholesterol conjugates on plasma membrane viability of Piau swine breed frozen/thawed spermatozoa.

    Science.gov (United States)

    Pinho, R O; Lima, D M A; Shiomi, H H; Siqueira, J B; Silveira, C O; Faria, V R; Lopes, P S; Guimarães, S E F; Guimarães, J D

    2016-08-01

    The objective of this study was to investigate the effect of cyclodextrin-loaded cholesterol conjugates addition to freezing extenders on plasma membrane viability of frozen-thawed spermatozoa of the Piau swine breed. Twenty semen samples were used from five males. The freezing extender was based on lactose-egg yolk extender, added to 2% glycerol, 3% dimethylacetamide. The addition of cyclodextrin-loaded cholesterol conjugates was performed after centrifugation, when semen was diluted with the cooling extender. Four groups were subjected to the following treatment: without addition (group 1); 1.5 mg of cyclodextrin-loaded cholesterol/120 × 10(6) sperm (group 2); 1.5 mg of cyclodextrin-loaded cholestanol/120 × 10(6) sperm (group 3); 1.5 mg of cyclodextrin-loaded desmosterol/120 × 10(6) sperm (group 4). To check post-thawing sperm quality sperm motility and sperm morphology evaluation were used. Additionally, to check sperm viability the hypoosmotic swelling test, supravital staining, and fluorescent assay were used. The mean values recorded for total sperm motility of semen immediately after thawing were 54.5 ± 5.8, 55.5 ± 5.3, 53.7 ± 6.7, and 52.5 ± 6.6% respectively for groups one to four, without difference between themselves (p > 0.05). Regarding fluorescent assay the results were 28.3 ± 13.2, 26.9 ± 12.2, 22.2 ± 11.4, and 32.0 ± 15.3% respectively for groups one to four, also without difference between groups (p > 0,05). Similarly, complementary tests for evaluating the integrity and functionality of the plasma membrane showed no difference between treatments (p > 0.05). In conclusion, use of cyclodextrin-loaded cholesterol conjugates added to the plasma membrane of sperm did not demonstrate any additive effect on increasing and/or maintaining sperm motility. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Effect of freezing and thawing rates on the post-thaw viability of boar spermatozoa frozen in FlatPacks and Maxi-straws.

    Science.gov (United States)

    Eriksson, B M; Rodriguez-Martinez, H

    2000-11-01

    The effects of different freezing and thawing rates on the post-thaw motility and membrane integrity of boar spermatozoa, processed as split samples in Maxi-straws or flat PET-plastic packages (FlatPack) were studied. A programmable freezing device was used to obtain freezing rates of either 20, 50 or 80 degrees C/min. Thawing of the samples was performed in a bath of circulating water; for 40s at 50 degrees C or 27s at 70 degrees C for Maxi-straws and 23s at 35 degrees C, 13s at 50 degrees C or 8s at 70 degrees C for the FlatPacks. Sperm motility was assessed both visually and with a computer assisted semen analysis (CASA) apparatus, while plasma membrane integrity was assessed using the fluorescent probes Calcein AM and ethidium homodimer-1. Temperature changes during freezing and thawing were monitored in both forms of packaging. Values for motile spermatozoa, sperm velocity and lateral head displacement variables were significantly (pstraws, with superior results at higher thawing rates. Freezing at 50 degrees C/min yielded better motility than 20 or 80 degrees C/min, although the effect was rather small. Neither freezing rate nor thawing rate had any effect on membrane integrity (p>0.05). A significant boar effect was seen for several parameters. The most striking difference in temperature courses between containers was a 4-5-fold lowering of the thawing rate, between -20 and 0 degrees C, in the center of the Maxi-straw, compared with the FlatPack. This is apparently due to the insulating effect of the thawed water in the periphery of the Maxi-straw. The improvement in sperm motility seen when using the FlatPack appears to be related to the rapid thawing throughout the sample, which decreases the risk of cell damage due to recrystallization during thawing. Since sperm motility patterns have been reported to be correlated with fertility both in vitro and in vivo it is speculated that the use of the FlatPack might improve the results when using frozen

  20. Effects of the technique of cryopreservation and dilution/centrifugation after thawing on the motility and vitality of spermatozoa of oligoasthenozoospermic men

    Directory of Open Access Journals (Sweden)

    Esteves Sandro C.

    2003-01-01

    Full Text Available OBJECTIVE: Comparing in human semen samples with low initial quality, the effects of 2 techniques of cryopreservation and dilution/centrifugation after thawing on the spermatic motility and vitality. MATERIALS AND METHODS: Semen samples from 15 oligo and/or asthenozoospermic individuals assisted in the infertility sector of a tertiary hospital were obtained through masturbation. The samples were divided into 2 portions of equal volume, and diluted (1:1; v/v with the cryoprotector containing glycerol (Test yolk buffer. One portion was frozen through the technique of liquid nitrogen vapor with static phases (group I - GI, while the other was frozen through a programmable biological freezer with linear speed (Planer, Kryo 10, series III (group II - GII. The following parameters were assessed before freezing and after thawing: percentage of spermatozoa with progressive motility (Prog% and percentage of live spermatozoa (Vit%. After defrosting, Prog% was assessed before and after removal of cryoprotector diluent, in different time intervals (zero, 3 h, and 24 h. The statistical analysis has been accomplished by using the non-parametric tests of Wilcoxon and Friedman. RESULTS: There was significant reduction of Prog% and Vit% from before freezing to after defrosting in both groups, I and II (p < 0.001. Values of Prog% and Vit% were not statistically different between groups, after thawing. It has been observed a significant reduction in Prog% among portions frozen with the automated technique after dilution and centrifugation for removal of cryoprotector (p = 0.006. After cryoprotector removal, Prog% has been kept unaltered, in both groups, during the first 3 hours of incubation, although being superior in group I (p = 0,04. There was a significant decrease in Prog% after 24 hours of incubation, in both groups (p < 0,01. CONCLUSION: For human semen samples with low initial quality, freezing through vapor technique or through the automated technique

  1. VITRIFICATION OF MAMMALIAN SPERMATOZOA

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    Sánchez R

    2015-12-01

    Full Text Available La criopreservación de espermatozoides y su utilización en inseminación artificial ha causado gran impacto sobre la reproducción de especies comerciales y en humanos. Sin embargo, a pesar del gran avance en el tema, constantemente se modifican los procedimientos de congelamiento, a fin de obtener mejores resultados de viabilidad y movilidad espermática, para lograr de esta forma mayores porcentajes de fecundaciones exitosas. No obstante, el proceso de criopreservación induce perdida de la función espermática que se refleja en alteraciones de la membrana plasmática, daño a nivel mitocondrial, deterioro en la integridad del ADN, aumento en la producción de las especies reactivas de oxígeno (ROS y alteraciones del citoesqueleto, lo que finalmente tiene efectos negativos en la viabilidad y motilidad espermática, elementos esenciales para la fecundación. En la búsqueda de métodos que induzcan el menor daño celular, es que ha desarrollado en los últimos años los métodos de vitrificación espermática. Estos han permitido no solo superar en cuanto a la mantención de la función espermática a los métodos convencionales de congelación, sino también que lo hace más rápido, seguro y de menor costo. Esto último al eliminar los equipos necesarios para la congelación lenta y también en algunos casos la no utilización de nitrógeno líquido para el almacenamiento, bastando un congelador de - 80°C. Su aplicación en mamíferos, ha sido reportada exitosamente en humanos con nacimientos de niños sanos, tanto en técnicas de fecundación in vitro como inseminación intrauterina y recientemente con el nacimiento de los primeros felinos (gatos con técnicas de inseminación intrauterina.

  2. Royal jelly supplemented soybean lecithin-based extenders improve post-thaw quality and incubation resilience of goat spermatozoa.

    Science.gov (United States)

    Alcay, Selim; Toker, M Berk; Onder, N Tekin; Gokce, Elif

    2017-02-01

    The aim of the present study was to evaluate different concentrations of royal jelly (RJ) supplemented extenders for post-thaw quality and incubation resilience of goat spermatozoa. Semen samples were collected from five goats. Pooled semen were diluted with soybean lecithin-based extender without RJ (control) or supplemented with different concentrations (0.25, 0.5 and 0.75%) of RJ (RJ0.25, RJ0.5, RJ0.75 respectively), at a final concentration of 150 × 10 6 spermatozoon/mL. Semen samples were assessed for sperm motility, plasma membrane integrity using hypoosmotic swelling test (HOST) damaged acrosome using FITC-Pisum sativum agglutinin (PSA-FITC) and DNA integrity using terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL). The addition of RJ (0.5%, 0.75%) led to higher percentages of subjective motilities (55.33 ± 2.29%, 57.67 ± 2.58%) compared to control and RJ0.25 groups (49.00 ± 2,80%, 51.67 ± 3.09%) (P  0.05). In the end of incubation, motility and HOST rates of RJ0.5 (14.00 ± 3.87%, 31.20 ± 3.70%) and RJ0.75 (15.00 ± 3.27%, 29.20 ± 2.59%) groups were higher than control (8.00 ± 2.54%, 18.20 ± 3.11%) and RJ0.25 (9.00 ± 2.07%, 20.60 ± 2.88%) groups (P < 0.05). Also defected acrosome and DNA fragmation rates of RJ0.5 (32.20 ± 1.30%, 5.4 ± 0.55%) and RJ0.75 (29.20 ± 1.30%, 5.80 ± 0.45%) groups were significantly lower than control (38.80 ± 0.84%, 7.40 ± 1.34%) and RJ0.25 (39.80 ± 2.05%, 7.00 ± 1.58) groups. This study shows that RJ supplemented extenders have beneficial effect on goat sperm parameters at 0 h and 6 h of incubation. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. DESCRIPTION OF ULTRASTRUCTURAL DAMAGES IN FROZEN-THAWED CANINE SPERMATOZOA DESCRIÇÃO DE DANOS ULTRAESTRUTURAIS EM ESPERMATOZOIDES CANINOS CONGELADOS-DESCONGELADOS

    Directory of Open Access Journals (Sweden)

    Viviane Helena Chirinéa

    2009-07-01

    Full Text Available In spite of the advances in transmission electronic microscopy (TEM, there are few studies presenting a systematic description of the canine spermatozoa and they are only focused on sperm cell heads. The goal of the present study was to describe ultrastructural appearance of canine fresh and frozen-thawed spermatozoa, focusing on damages induced by freezing-thawing in different sperm regions. Ten ejaculates from five proven stud dogs (two ejaculates/dog were collected, evaluated, extended in Tris-egg yolk-glycerol, frozen and stored in liquid nitrogen, and thawed six weeks later.  Samples were evaluated for progressive motility, morphology, and for ultrastructural analysis by TEM. Concerning to TEM, the most striking differences between fresh and frozen-thawed samples were observed over the mid-piece since the fresh spermatozoa showed a well preserved mid-piece. However, the frozen-thawed spermatozoa mid-piece showed signs of damage such as mitochondrial vacuolization. In conclusion, freezing-thawing processes using a Tris-egg yolk extender induce ultrastructural damages in head and mid-piece of canine sperm, affecting the mitochondrial ultrastructure besides.

    KEY WORDS: Cryopreservation, dog, semen, ultrastructure.
    Apesar dos avanços na microscopia eletrônica de transmissão (MET, existem poucos estudos apresentando uma descrição sistemática do espermatozoide canino, os quais estão focados apenas na cabeça espermática. Objetivou-se, com o presente estudo, descrever a aparência ultraestrutural do espermatozoide canino fresco e congelado-descongelado, enfocando os danos induzidos pela congelação e descongelação em diferentes regiões espermáticas. Dez ejaculados obtidos de cinco cães (dois ejaculados por cão foram coletados, avaliados e diluídos em Tris-gema-glicerol, congelados e armazenados em nitrogênio líquido, e descongelados seis semanas após. Avaliaram-se as amostras quanto à motilidade progressiva

  4. Análise computacional da compactação da cromatina de espermatozoides de galo Computational analysis of chromatin condensation of rooster spermatozoa

    Directory of Open Access Journals (Sweden)

    A.C.N. Rodrigues

    2009-12-01

    Full Text Available Testaram-se variantes metodológicas utilizando azul de toluidina (AT, até se estabelecer um protocolo confiável para a avaliação computacional da compactação da cromatina em espermatozoides de galo. Para tal, foram utilizados sêmen de 10 galos com 35 semanas de idade e sêmen de 10 galos com 60 semanas de idade. O melhor método foi o de hidrólise com ácido clorídrico 1N por 10 minutos, coloração em cubeta com AT 0,025%, pH 4,0, por 20 minutos, desidratação em álcool, diafanização em xilol e montagem com bálsamo do Canadá. Todas as amostras de sêmen foram submetidas a este protocolo e posteriormente avaliadas por análise de imagem computacional, em que foram feitas mensurações da área, comprimento, largura, perímetro, homogeneidade da compactação da cromatina dentro de cada cabeça e intensidade de compactação da cromatina. Os espermatozoides de galos velhos apresentaram mais alterações na cromatina que os de galos jovens. Os galos jovens apresentaram cabeça dos espermatozoides maior que os galos mais velhos. A análise computacional da compactação da cromatina mostrou-se um método menos subjetivo e mais preciso que a avaliação visual das cabeças dos espermatozoides.The methodological variants using toluidina blue (AT to establish a trustworthy protocol for the computational analysis of chromatin condensation of rooster spermatozoa were studied. Twenty semen samples were used: ten from 35-week-old roosters and ten from 60-week-old roosters. Different methods of denaturation and staining were tested. The best method was hydrolysis with 1N HCl for 10 minutes, staining in bucket with 0.025% AT, pH 4.0, for 20 minutes, dehydration in alcohol, clearing in xylol, and mounted with Canada balsam. All the semen samples were submitted to this protocol and later evaluated by computational image analysis. Area, length, width, perimeter, and chromatin compaction homogeneity of head spermatozoa were measured. The sperm

  5. TESTICULAR AND EPIDIDYMAL HISTOPATHOLOGICAL EVALUATION IN MONGREL ADULT DOGS FROM SÃO JOÃO DA BOA VISTA (SP REGION AVALIAÇÃO HISTOPATOLÓGICA TESTICULAR E EPIDIDIMÁRIA EM CÃES ADULTOS SEM RAÇA DEFINIDA (SRD DA REGIÃO DE SÃO JOÃO DA BOA VISTA, SP

    Directory of Open Access Journals (Sweden)

    Renée Laufer Amorim

    2007-12-01

    Full Text Available

    Little has been explored about testicular and epidi-dymal changes in dogs in different Brazilian regions and their influence on canine fertility, mainly in mongrel dogs. This study was performed to verify the prevalence of testicular and epididymal histopathological changes in mongrel adult dogs from São João da Boa Vista (SP region, as well as the frequency according to animal size and occurrence of simultaneous testicular and epididymal lesions. The use of mongrel dogs was based on the fact that it is the most common breed in the Brazilian canine popula-tion. Testicles and epipidymis were obtained from 60 adult dogs, between 3 and 6 years old, in castration campaign and sorted to in small breed (28 animals, medium breed (28 animals and large breed (14 animals. Histopathological evaluation of the 120 testicles showed 107 cases of degeneration, 89 of tubular atrophy, 16 of hipoplasia, 6 of orchitis, 3 leydig cell tumor, 2 normal tissue, one sertolli cell tumor and one case of edema. In the epididymal evaluation, 55 were considered normal, 36 inflammatory, 22 papillary hyperplasia, 14 degenerations, 12 cribriform hyperplasia, 3 cases of fibrosis, one edema, one adeno-miosis and one spermatic granuloma. Testicular and epididymal lesions were more frequents in medium and large breeds. Cribriform epididymal hyperplasia was observed and papillary epididymal hyperplasia for papillary epithe-lial projections with normal morphology in the lumen was proposed.

    Key-words: Dog, epididym, testicle. histopathological alterations.

    A ocorrência de alterações testiculares e epididi-márias em cães nas diferentes regiões do país, assim como a influência delas sobre a fertilidade da espécie são ainda pouco exploradas, especialmente de cães sem raça definida (SRD. Neste estudo buscou-se determinar a prevalência das alterações histopatológicas testiculares e epididimárias nos

  6. Penambahan Vitamin C Pada Pengencer Spermatozoa Babi Landrace Yang Disimpan Pada Suhu 15ºC (ADDITION OF VITAMIN C IN DILUENT OF LANDRACE SPERM STORED IN TEMPERATURE OF 15ºC

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    Wayan Bebas

    2015-08-01

    Full Text Available The aims of this study were to determine the viability and motility of Landrace pig’sspermatozoa in the Beltsville Thawing Solution (BTS diluent with addition vitamin C which werestored at a temperature of 15ºC. During the storage process, metabolic activity of spermatozoaproduce free radicals that can reduce the viability and motility of spermatozoa thus it needs theaddition of antioxidants. Vitamin C is an antioxidant that can be used to scavengethe free radicals.This study used vitamin C added into the BTS diluent with scavenge dose is 0.1 mg/ml, 0.2 mg/mland a dose of 0.3 mg/ml. The results showed that a dose of 0.1 mg/ml is the best dose to maintainthe viability and motility of Landrace pigs spermatozoa were storage at a temperature of 15ºC.

  7. Effect of season on scrotal circumference, semen characteristics, seminal plasma composition and spermatozoa motility during liquid storage in INRA180 rams.

    Science.gov (United States)

    Benmoula, Anass; Badi, Abdelmoughit; El Fadili, Moussa; El Khalil, Kaoutar; Allai, Larbi; El Hilali, Abderaouf; El Amiri, Bouchra

    2017-05-01

    The present study was undertaken to assess the effect of seasons on scrotal circumference, semen characteristics, seminal plasma composition, and sperm motility during liquid storage of INRA180 rams. The semen was collected from five mature INRA180 rams (2-3 years of age) during one year (from April 2014 to March 2015). Scrotal circumferences, semen characteristics, some biochemical parameters of seminal plasma were evaluated. Immediately after collection and evaluation, the semen was pooled and extended in skim milk (SM) at 15°C to reach 0.8×109 spermatozoa/ml. Thereafter, samples were evaluated at different storage times (0, 8, and 24h). The results showed that scrotal circumference, semen quality and the concentration of total protein in seminal plasma were relatively constant during the year (P>0.05). However, total lipid and cholesterol concentrations increased significantly (P0.05). To conclude, the INRA180 rams have the ability to produce semen with high quality all over the year. The only parameters showing seasonal variations are cholesterol, total lipid, and progressive motility. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. IMPROVEMENT OF QUALITY OF LIFE AND MORPHOFUNCTIONAL CHARACTERISTICS OF SPERMATOZOA IN MEN WITH CHRONIC NONBACTERIAL PROSTATITIS AND IN A PROGRAM OF PRECONCEPTION PREPARATION TO FATHERHOOD

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    O. B. Zhukov

    2017-01-01

    Full Text Available In the article, the results of diagnostics and treatment of 30 men aged 30–45 (mean age 36.2 ± 5.3 years with chronic nonbacterial prostatitis (CNP and male factor infertility in marriage are presented. Among the patients, 15 patients had CNP and secretory infertility (1st group; mean age 35.2 ± 4.3 years, 15 men with infertility caused by CNP and varicocele in the postoperative period (2nd group; mean age 33.6 ± 3.3 years. Duration of the male factor infertility varied from 1 to 4.5 years, and on average it was 2.8 ± 1.6 years. After detailed questioning and examination, including a spermiogram, determination of antisperm antibodies, DNA fragmentation test, and electron microscopy of the ejaculate, patients in the 1st group received Prostatilen AC as rectal suppositories for 10 days. Patients in the second group received Prostatilen AC in the form of rectal suppositories for 20 days. We showed an improvement of quality of life and spermatogenesis characteristics with decreased MAR test (more pronounced in the 2nd group, a decrease in the DNA fragmentation index, and an improvement in ultrastructural morphofunctional characteristics of spermatozoa. We justify the use of Prostatilen AC to increase effectiveness of assisted reproductive technologies in men in programs of preconception preparation to fatherhood. 

  9. THE EFFECT OF SEMEN WASHING AND SOYBEAN LECITHIN LEVEL ON MOTILITY AND VIABILITY OF RAM SPERMATOZOA STORED AT 5⁰C

    Directory of Open Access Journals (Sweden)

    S. Salmin

    2014-10-01

    Full Text Available The objectives of this reserarch were to study the effect of semen washing and soybean lecithinlevel on the motility and viability of ram spermatozoa stored at 5⁰C. Semen was collected from threemature rams by artificial vagina. Semen was evaluated by macroscopic and microscopic. Semen wasdivided into ten tubes, the semen of first five tubes was unwashed (W0 and diluted with extender 0%,1%, 2%, 3% and 4% soybean lecithin (L0, L1, L2, L3 and L4. Semen of another five tubes werewashed (W1 then each diluted with similar extender levels. The diluted semen samples were stored at5⁰C and the sperm motility and viability were evaluated each day. The interaction of the semen washingand soybean lecithin levels were no significant differences (P>0.05 but the single factor of soybeanlecithin level was significant differeNces (P<0.05 and the single factor of semen washing were nosignificant differences (P>0.05 on progressive motility and viability of ram sperm.The best extenderwas 3% soybean lecithin (L3 with the percentage of ram sperm progressive motility was 63.18 ± 3.61%and viability was 71.76±2.32%.

  10. Effect of storage in short--and long-term commercial semen extenders on the motility, plasma membrane and chromatin integrity of boar spermatozoa.

    Science.gov (United States)

    De Ambrogi, Marco; Ballester, Juan; Saravia, Fernando; Caballero, Ignacio; Johannisson, Anders; Wallgren, Margareta; Andersson, Magnus; Rodriguez-Martinez, Heriberto

    2006-10-01

    For artificial insemination (AI) in pigs, preservation of liquid boar semen at 16-20 degrees C is still common practice as sperm cryopreservation remains suboptimal in this species. To meet the different needs of the swine industry, several extenders have been developed to preserve semen in liquid form for short--and long-term storage. In the present study, three different commercial extenders devised for short-term (BTS+) or long-term preservation (MR-A and X-Cell), were used to test whether storage of semen from four mature, fertile boars at 17 degrees C for 96 h would affect sperm characteristics relevant for fertility, such as motility, membrane integrity and chromatin stability. Computer-assisted sperm analysis, and stainings with the acylated membrane dye SYBR-14/propidium iodide, and acridine orange in connection with flow cytometry were used to evaluate these variables. Percentages of total motile spermatozoa decreased slightly, but significantly, after 72-96 h. While membrane integrity values varied during the period of study, no significant changes in either membrane integrity or chromatin stability were, however, registered. This suggests a customary 96-day storage at 17 degrees C in these extenders was too short an interval to cause losses of integrity in nuclear DNA in the boar population studied.

  11. Molecular detection of Chlamydia trachomatis and other sexually transmitted bacteria in semen of male partners of infertile couples in Tunisia: the effect on semen parameters and spermatozoa apoptosis markers.

    Directory of Open Access Journals (Sweden)

    Hanen Sellami

    Full Text Available This study was undertaken to determine the prevalence of Chlamydia trachomatis, Mycoplasmas, and Ureaplasmas in semen samples of the male partners of infertile couples and to investigate whether Chlamydia trachomatis could initiate apoptosis in human spermatozoa. A total of 85 males partners of infertile couples undergoing routine semen analysis according to World Health Organization guidelines were included. Specimens were examined for the presence of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma urealyticum and Ureaplasma parvum by Real time PCR (qPCR. Semen specimens were analysed for the appearance of apoptotic markers (sperm DNA fragmentation, activated caspase 3 levels, mitochondrial membrane potential (ΔΨm using flow cytometry. C. trachomatis, N. gonorrhoeae, U. urealyticum, M genitalium were detected in semen samples of 13 (15.2%, 5 (5.8%, 5 (5.8% and 3 (3.5% male partners of infertile couples, respectively. M. hominis and U. parvum were detected in semen sample of only one patient (1.1%. The semen of infertile men positive for C. trachomatis showed lower mean of semen count and lower rapid progressive motility (category [a] of spermatozoa compared to uninfected men with statistically significances (p = 0.02 and p = 0.04, respectively. Flow cytometry analyses demonstrated a significant increase of the mean rate of semen with low ΔΨm and caspase 3 activation of infertile men positive for C. trachomatis compared to uninfected men (p = 0.006 and p = 0.001, respectively. DNA fragmentation was also increased in sperm of infertile men positive for C. trachomatis compared to uninfected men but without statistical significances (p = 0.62. Chlamydial infection was associated to loss of ΔΨm and caspase 3activation. Thus, C. trachomatis infection could be incriminated in apoptosis induction of spermatozoa. These effects may explain the negative direct impact of C

  12. Effect of post-thaw addition of seminal plasma on motility, viability and chromatin integrity of cryopreserved donkey jack (Equus asinus) spermatozoa.

    Science.gov (United States)

    Sabatini, C; Mari, G; Mislei, B; Love, Cc; Panzani, D; Camillo, F; Rota, A

    2014-12-01

    Pregnancy rates in donkeys after artificial insemination with cryopreserved semen are still low, compared to the horse species. Addition of autologous seminal plasma to frozen-thawed semen appeared to improve pregnancy rates. The aims of this study were to evaluate (1) sperm motility and plasma membrane integrity after thawing (T0) and after one and 2 h (T1 and T2) of post-thaw incubation in either 0% (SP0) or 70% (SP70) autologous seminal plasma and (2) sperm motility, plasma membrane integrity and DNA quality (%COMP-αt) after thawing (T0) and after 2 and 4 h (T2 and T4) of post-thaw incubation in either 0% (SP0), 5% (SP5) or 20% (SP20) homologous seminal plasma. In experiment 1, seminal plasma decreased total and progressive sperm motility and plasma membrane intact spermatozoa immediately after dilution and at all following time points (p < 0.05). In experiment 2, total and progressive motility did not differ between treatments immediately after dilution and between SP0 and SP5 at T2, while they were lower in both SP5 and SP20 than in SP0 at T4. Plasma membrane intact sperm cells did not differ between SP0 and SP5 and were lower in SP20 at all time points. DNA quality was not affected by treatment immediately after dilution and was significantly worse for SP20 after 4 h of incubation (p < 0.05). The post-thaw addition of seminal plasma at the tested concentrations did not improve donkey frozen semen characteristics in vitro over time. © 2014 Blackwell Verlag GmbH.

  13. Effects of post-mortem storage conditions of bovine epididymides on sperm characteristics: investigating a tool for preservation of sperm from endangered species

    DEFF Research Database (Denmark)

    Strand, Julie; Ragborg, Mette M; Pedersen, Hanne Skovsgaard

    2016-01-01

    The aim of this study was to establish and validate a reliable and efficient protocol for the recovery and cryopreservation of epididymal spermatozoa used for in vitro fertilization, using bulls of two different age classes. Testicles from 26 (37–51 weeks old, group 1) and 19 (52–115 weeks old, g...... that epididymal spermatozoa recovered from testicles kept in specific conditions can be used to preserve genetic material from endangered and threatened species or populations in nature as well as in domestic and zoo animals....

  14. Methylation status of imprinted genes DLK1-GTL2, MEST (PEG1), ZAC (PLAGL1), and LINE-1 elements in spermatozoa of normozoospermic men, unlike H19 imprinting control regions, is not associated with idiopathic recurrent spontaneous miscarriages.

    Science.gov (United States)

    Ankolkar, Mandar; Salvi, Vinita; Warke, Himangi; Vundinti, Babu Rao; Balasinor, N H

    2013-05-01

    To study methylation aberrations in spermatozoa at developmentally important imprinted regions to ascertain their role in early embryo loss in idiopathic recurrent spontaneous miscarriages (RSM). Case-control study. Academic research setting at National Institute for Research in Reproductive Health, Parel, Mumbai. Male partners of couples with a history of RSM and male partners of couples with proven fertility (control group). None. DNA methylation levels at imprinting control regions of DLK1-GTL2, MEST (PEG1), and ZAC (PLAGL1) by Epityper Massarray and global methylation levels as measured by LINE-1 methylation and anti-5-methyl cytosine antibody in spermatozoa of 23 men in control group and 23 men in RSM group. We did not observe any aberration in the total methylation levels in any of the imprinted genes or global methylation analyzed. Our results indicate that paternal methylation aberrations at imprinting control regions of DLK1-GTL2, MEST (PEG1), and ZAC (PLAGL1) and global methylation levels are not associated with idiopathic RSM and may not be good epigenetic markers (unlike the H-19 imprinting control region) for diagnosis of idiopathic RSM. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  15. Aminopurvalanol A, a Potent, Selective, and Cell Permeable Inhibitor of Cyclins/Cdk Complexes, Causes the Reduction of in Vitro Fertilizing Ability of Boar Spermatozoa, by Negatively Affecting the Capacitation-Dependent Actin Polymerization

    Directory of Open Access Journals (Sweden)

    Nicola Bernabò

    2017-12-01

    Full Text Available The adoption of high-througput technologies demonstrated that in mature spermatozoa are present proteins that are thought to be not present or active in sperm cells, such as those involved in control of cell cycle. Here, by using an in silico approach based on the application of networks theory, we found that Cyclins/Cdk complexes could play a central role in signal transduction active during capacitation. Then, we tested this hypothesis in the vitro model. With this approach, spermatozoa were incubated under capacitating conditions in control conditions (CTRL or in the presence of Aminopurvalanol A a potent, selective and cell permeable inhibitor of Cyclins/Cdk complexes at different concentrations (2, 10, and 20 μM. We found that this treatment caused dose-dependent inhibition of sperm fertilizing ability. We attribute this event to the loss of acrosome integrity due to the inhibition of physiological capacitation-dependent actin polymerization, rather than to a detrimental effect on membrane lipid remodeling or on other signaling pathways such as tubulin reorganization or MAPKs activation. In our opinion, these data could revamp the knowledge on biochemistry of sperm capacitation and could suggest new perspectives in studying male infertility.

  16. Viability of Timor deer stag (Cervus timorensis spermatozoa extended in tris egg yolk diluent with different sources of carbohydrate and storage at room temperature

    Directory of Open Access Journals (Sweden)

    W. Marlene Mesang-Nalley

    2007-12-01

    Full Text Available The successful sperm preservation, influenced by the capability of its extender on the maintenance the sperm quality during storage. The carbohydrate such as glucose and fructose were the common sugar added on the mammalian sperm extender to support their live and motility. The sucrose was the main carbohydrate in Timor deer stag seminal plasma. The experiment was conducted to evaluate the effect of carbohydrates in Tris egg yolk (TEY extender on the motility and viability of stag sperm, stored in room temperature (27-28 oC. The semen was collected using electro ejaculator from five Timor deer stags at hard antler stage, 3-5 years old, body weight of 64-102 kg with normal testes. The semen was than evaluated macro-and microscopically and divided into 3 aliquots. Each of them was diluted with TEY-glucose (TEYG, TEY-fructose (TEYF and TEY-Sucrose (TEYS with the concentration of spermatozoa 100 x 106 ml-1. The extended semen was than stored at room temperature. The sperm motility and viability were evaluated every 3 hours. Result of the experiment showed that the semen volume was 2.06 ± 0.63 ml, pH 7.03±0.13, yellow white until creamy in color and the consistency ranged from normal to thick. The mass movement between ++ to +++ and the sperm motility was 68.67 ± 7.4%. The average of sperm concentration was 842.35 ± 258.14x106 ml-1, the viable sperm was 78.11 ± 3.61%, the sperm abnormality was 7.31 ± 2.98%. The percentages of sperm motility on TEYG (18.00 ± 17.63% and TEYS (21.83 ± 15.92% were higher compare to TEYF (4,00 ± 0,00% extender in 24 hours observation. The percentage of sperm viability showed the same pattern. The sperm viability in TEYG (28.17 ± 20.06 and TEYS (24.00 ± 22.59% (P<0.05 were significantly higher compare to TEYF (4.00 ± 0.00%. It is concluded that the deer stag sperm can use the three sugars for their nutrition source. The diluted sperm still can be used for artificial insemination after 12 hour storage.

  17. Biogeographical plant-soil relations of invasive medusahead (Elymus caput-medusae)

    Science.gov (United States)

    Understanding the mechanisms that underlie the success of invasive plant species is integral to predicting and ameliorating their negative impacts. Many hypotheses have consequently been proposed to explain invasive behavior. This lack of consensus within invasion ecology can partially be attributed...

  18. Consequences of Gonadotropin Administration on Fertilization and Early Embryonic Development in the Domestic Cat and the In Vitro Fertilization of Feline Follicular Oocytes

    Science.gov (United States)

    1987-08-12

    embryo transfer is a major industry. In 1972, a total of 20 offspring were produced in the United States from embryo transfers in cattle whereas...accomplished by Chang and Yanagimachi (1963). Epididymal spermatozoa, collected after castration , were surgically placed into the ovarian capsule of...spermatozoa (0.2 to 1.8 X 106/ml), collected from the caudae epididymides of castrated males, were placed with the oocytes in a modified Kreb’s Ringer

  19. Sperm cell granuloma in a gobbler ( Meleagris Gallopavo ) | Ajayi ...

    African Journals Online (AJOL)

    Microscopically, there was severe diffuse testicular degeneration and necrosis of the germinal epithelial cells, extravasation of spermatozoa into the epididymal interstitium, inciting a granulomatous reaction with arteritis. Based on these findings, sperm cell granuloma was diagnosed. This is probably the first reported case ...

  20. Antifertility effects of Oldenlandia affinis in male rats - a preliminary ...

    African Journals Online (AJOL)

    Testis histology revealed fewer spermatozoa or azoospermic seminiferous tubules in treated animals compared to controls with no change in neither tubule thickness nor Sertoli cell structure. O. affinis treatment caused a 17% decrease in sperm motility but there was no change in cauda epididymal sperm counts. However ...

  1. Alterações de cromatina em espermatozóides de ovinos e caprinos avaliadas por azul de toluidina e alaranjado de acridina Chromatin alterations in ram and goat spermatozoa evaluated by toluidine blue and acridine orange

    Directory of Open Access Journals (Sweden)

    Cristina de Figueiredo Kamimura

    2010-02-01

    Full Text Available Reprodutores com espermograma normal podem se comportar como subférteis ou passarem por períodos de subfertilidade. As alterações na cromatina dos espermatozóides são possíveis explicações encontradas para tais comportamentos. Conduziu-se este trabalho, com o objetivo de testar a eficiência de azul de toluidina (AT e do alaranjado de acridina (AA na identificação de alterações na compactação de cromatina em espermatozóides de ovinos e caprinos, além de avaliar a correlação entre essas alterações e as de morfologia espermática. Para tal, foram avaliadas amostras de sêmen de 15 ovinos e de 15 caprinos, com dez repetições para cada método por animal. Calcularam-se a média, o desvio padrão (DP e o coeficiente de variação (CV para cada técnica e animal. Utilizou-se o teste t-Student para avaliar diferença entre as médias obtidas nos dois métodos. Também foram calculados a correlação de Pearson e os coeficientes kappa ponderado e não ponderado para avaliar a concordância entre os métodos com AT e AA. Foi verificado que nem sempre as anomalias morfológicas de cabeça são acompanhadas por alterações na cromatina identificáveis pelos métodos utilizados neste trabalho. O método AT é mais estável e possui maior sensibilidade do que AA para ambas as espécies, sendo o mais indicado para caprinos. Contudo, em razão de apresentar repetibilidade muito baixa, ambos os métodos não são indicados para avaliação espermática em ovinos.Males with normal spermogram can behave as subfertile or pass for periods of subfertility. Chromatin alterations of spermatozoa can account for such behavior. The objective of the present work was to test the efficacy of toluidine blue (TB and acridine orange (AO in the identification of alterations in chromatin compaction in spermatozoa from rams and goats, in addition to evaluate the correlation between those alterations and the ones of spermatic morphology. In order to do that

  2. In vitro evaluation of canine spermatozoa cryopreserved in different extenders Avaliação in vitro do sêmen canino criopreservado em diferentes diluidores de congelação

    Directory of Open Access Journals (Sweden)

    E.C.S. Oliveira

    2006-12-01

    Full Text Available The efficacy of three extenders, tris-egg yolk-5% ethylene glycol (T1, lactose-egg yolk-5% ethylene glycol (T2 and lactose-egg yolk-5% dimethyl formamide (T3 on preserving the viability of post-thawing canine spermatozoa was evaluated. Three ejaculates per dog were obtained of five animals. The semen was packaged in 0.5ml straws and cooled to 4°C for 120min. The straws were frozen 4cm above the nitrogen level for 15min and thawed in water-bath at 37°C for 60sec and at 75°C for 7sec. Progressive motility and vigour were evaluated immediately after thawing (time 0 and at 30, 60, 90 and 120min. Structural and functional integrity of plasma membrane of the spermatozoa were evaluated, respectively, by fluorescent staining probes and hypoosmotic swelling test. Lactose-egg yolk based extenders showed better cryoprotectant capability and dimethyl formamide was an alternative cryoprotectant agent for dog sperm cells.Avaliou-se a eficácia de três diluidores, tris-gema com 5% de etileno glycol (T1, lactose-gema com 5% de etileno glicol (T2 e lactose-gema com 5% de dimetil-formamida (T3 na criopreservação do sêmen de cães. Foram obtidos três ejaculados por cão de um total de cinco animais. O sêmen foi envasado em palhetas de 0,5ml e resfriado até 4°C por 120min. As palhetas foram congeladas 4cm acima do nitrogênio líquido por 15min e descongeladas em banho-maria a 37°C por 60seg e 75°C por 7seg. A motilidade progressiva e o vigor foram avaliados imediatamente após a descongelação (tempo 0 e aos 30, 60, 90 e 120min. A integridade estrutural e funcional da membrana plasmática do espermatozóide foi avaliada, respectivamente, por meio da coloração de fluorescência e pelo teste hiposmótico. Os diluidores à base de lactose gema foram mais eficazes em preservar a viabilidade espermática pós-descongelação e a dimetil-formamida é um crioprotetor eficaz para espermatozóides de cães.

  3. The oviducal protein, heat-shock 70-kDa protein 8, improves the long-term survival of ram spermatozoa during storage at 17°C in a commercial extender.

    Science.gov (United States)

    Lloyd, R E; Fazeli, A; Watson, P F; Holt, W V

    2012-01-01

    Poor fertility rates are often observed when fresh ram semen stored in conventional extenders is used for cervical artificial insemination (AI). Heat-shock 70-kDa protein 8 (HSPA8), found within the oviduct, prolongs boar, ram and bull sperm survival at body temperatures in vitro. Here, we aimed to determine whether supplementing extenders (INRA-96 and RSD-1) with HSPA8 (4 µg mL⁻¹) would improve their performance in maintaining freshly collected ram sperm viability and sperm nuclear DNA integrity during storage over 48 h at 17°C. Sperm function was assessed at 1, 6, 24 and 48h and this experiment was repeated using 25 × 10⁶ and 800 × 10⁶ spermatozoa mL⁻¹. INRA96 supplemented with HSPA8 maintained sperm viability significantly better than INRA96 alone at both sperm concentrations. However, sperm nuclear DNA fragmentation (DF) increased significantly during storage using the higher sperm concentration, irrespective of the extender and the protein treatment used. Increasing levels of sperm nuclear DF over time could explain why poor fertility rates are often observed following cervical AI using stored ram semen. However, further research is required to ascertain whether supplementing the commercially available INRA96 extender with HSPA8 will improve fertility rates following cervical AI using stored ram semen.

  4. In vivo serial sampling of epididymal sperm in mice.

    Science.gov (United States)

    Del Val, Gonzalo Moreno; Robledano, Patricia Muñoz

    2013-07-01

    This study was undertaken to refine the techniques of in vivo collection of sperm in the mouse. The principal objective was to offer a viable, safe and reliable method for serial collection of in vivo epididimary sperm through the direct puncture of the epididymis. Six C57Bl/6J males were subjected to the whole experiment. First we obtain a sperm sample of the right epididymis, and perform a vasectomy on the left side. This sample was used in an in vitro fertilization (IVF) experiment while the males were individually housed for 10 days to let them recover from the surgery, and then their fertility was tested with natural matings until we obtained a litter of each one. After that, the animals were subjected another time to the same process (sampling, recover and natural mating). The results of these experiments were a fertilization average value of 56.7%, and that all the males had a litter in the first month after the natural matings. This study documented the feasibility of the epididimary puncture technique to in vivo serial sampling of sperm in the mouse.

  5. Effect of Maternal Alcohol Consumption on Epididymal Growth in ...

    African Journals Online (AJOL)

    A study was conducted to determine the effect of maternal alcohol consumption on the growth of epididymis in neonatal mice. Three groups of adult female mice were used. The pups of group 1 served as control while the pups of groups 2 and 3 were given 30% ethanol (v/v) during pregnancy and during pregnancy and ...

  6. Efecto del levonorgestrel como anticonceptivo oral de emergencia en la ovulación, el endometrio y los espermatozoides Effect of levonorgestrel in the ovulation, endometrium, and spermatozoa for emergency oral contraception

    Directory of Open Access Journals (Sweden)

    Víctor J. Suárez

    2010-06-01

    doesn’t affect the spermatozoa in their migration or egg-penetration capacities. No morphological or molecular alterations in the endometrium that could interfere with the implantation of the fertilized egg have been demonstrated. There is no actual scientific evidence available supporting that the use of levonorgestrel for EOC is abortive.

  7. Scattering of spermatozoa off cylindrical pillars

    Science.gov (United States)

    Bukatin, Anton; Lushi, Enkeleida; Kantsler, Vasily

    2017-11-01

    The motion of micro-swimmers in structured environments, even though crucial in processes such as in vivo and in vitro egg fertilization, is still not completely understood. We combine microfluidic experiments with mathematical modeling of 3D swimming near convex surfaces to quantify the dynamics of individual sperm cells in the proximity of cylindrical pillars. Our results show that the hydrodynamic and contact forces that account for the shape asymmetry and flagellar motion, are crucial in correctly describing the dynamics observed in the experiments. Last, we discuss how the size of the cylindrical obstacles determines whether the swimmers scatter off or get trapped circling the pillar.

  8. Características gerais dos espermatozóides em homens varicocele clínica General characteristics of the spermatozoa in oligozoospermic men with and without clinical varicocele

    Directory of Open Access Journals (Sweden)

    F.T. Andrade-Rocha

    1997-03-01

    Full Text Available Pacientes com oligozoospermia apresentam uma redução na qualidade do sêmen, independente da etiologia do distúrbio. OBJETIVO. Investigar o papel da varicocele na redução da qualidade do sêmen, em homens oligozoospérmicos. MÉTODOS. Foram estudados dez pacientes portadores de varicocele clínica esquerda (PCV e 21 pacientes não-portadores desta entidade (PSV, atendidos em um laboratório privado, em Petrópolis, RJ. Foram analisados os seguintes parâmetros seminais: contagem de espermatozóides, vitalidade, motilidade e morfologia espermática. PCV e PSV foram comparados entre si e com um grupo-controle (n=15 de pacientes sem varicocele clínica e com espermograma normal. RESULTADOS. PCV e PSV apresentaram uma redução significativa na vitalidade (43,9% e 34,9% versus 73,0% no grupo-controle, nos graus (a (5,3% e 2,4% versus 32,4 % e (d (76,7% e 83,8% versus 44,9% de progressão espermática e nos percentuais de espermatozóides ovais (25,5% e 22,9% versus 61,2%, amorfos (25,4% e 23,8% versus 12,5% e outras anomalias (23,8% e 30,5% versus 13,0%. PCV ainda apresentou diferença significativa no percentual de espermatozóides fusiformes (10,9% versus 1,3% e PSV no grau (b de progressão espermática (11,0% versus 22,0%, em relação ao grupo-controle. Entre PCV e PSV, não foram encontradas diferenças significativas. CONCLUSÃO. A varicocele reduz a qualidade de semens oligozoospérmicos, mas esta redução também ocorre em oligozoospermias de outras etiologias.Patients with oligozoospermia show a reduction in the semen quality, independent of the etiology of the disturbance. PURPOSE - To investigate the role of the varicocele in the decrease of the semen quality in oligozoospermic men. METHODS - Ten patients with left clinical varicocele (termed PCV and 21 patients without this entity (termed PSV attended in a private laboratory from Petrópolis, RJ, were investigated. Sperm count, vitality, motility, and morphology of spermatozoa

  9. Digital image analysis of testicular and prostatic ultrasonographic echogencity and heterogeneity in dogs and the relation to semen quality

    OpenAIRE

    Moxon, Rachel; Bright, Lucy; Pritchard, Beth; Bowen, I. Mark; Souza, Mírley Barbosa de; Silva, Lúcia Daniel Machado da; England, Gary C.W.

    2015-01-01

    A semi-automated ultrasonographic method was developed to measure echogenicity and heterogeneity of the testes and prostate gland and relationships of these measures with semen quality were assessed in 43 fertile dogs. The relationship between animal age and body weight upon the volume of the testes, epididymal tail volume and prostate volume were also established.\\ud \\ud Mean testicular echogenicity was negatively correlated with the percentage of morphologically normal live spermatozoa (mor...

  10. Epididymosomes: transfer of fertility-modulating proteins to the sperm surface

    OpenAIRE

    Patricia A Martin-DeLeon

    2015-01-01

    A variety of glycosylphosphatidylinositol (GPI)-linked proteins are acquired on spermatozoa from epididymal luminal fluids (ELF) during sperm maturation. These proteins serve roles in immunoprotection and in key steps of fertilization such as capacitation, acrosomal exocytosis and sperm-egg interactions. Their acquisition on sperm cells is mediated both by membrane vesicles (epididymosomes, EP) which were first reported to dock on the sperm surface, and by lipid carriers which facilitate the ...

  11. EFSA NDA Panel (EFSA Panel on Dietetic Products, Nutrition and Allergies), 2015. Scientific Opinion on the substantiation of a health claim related to a combination of pomegranate pomace extract and greater galangal rhizome powder and an increase in the number of motile spermatozoa in semen

    DEFF Research Database (Denmark)

    Tetens, Inge

    2015-01-01

    on the scientific substantiation of a health claim related to a combination of pomegranate pomace extract (standardised by its content of punicalagins) and greater galangal rhizome powder (standardised by its content of acetoxychavicol acetate) and an increase in the number of motile spermatozoa in semen. The Panel...... the combination of pomegranate pomace extract and greater galangal rhizome powder was consumed for three months, that no other human studies in which these results have been replicated were provided, and that no evidence was provided for a mechanism by which the food could exert the claimed effect. The Panel...... concludes that a cause and effect relationship has not been established between the consumption of the combination of pomegranate pomace extract (standardised by its content of punicalagins) and greater galangal rhizome powder (standardised by its content of acetoxychavicol acetate) and an increase...

  12. Regional differences in steroidogenesis and hormone levels in the epididymis and vas deferens of adult rats.

    Science.gov (United States)

    Kumari, G L; Allag, I S; Das, R P; Datta, J K

    1980-06-01

    In vivo and in vitro studies with different parts of the epididymis and vas deferens were carried out to determine their inherent capacity to synthesize steroids and to correlate with the endogenous levels with or without the administration of hCG. Incubation with 14C-labelled pregnenolone and testosterone demonstrated that caput epididymidis was more active than other parts in synthesizing testosterone from 14C-pregnenolone and in converting labelled testosterone to 5 alpha-dihydrotestosterone (DHT). The cauda epididymidis and vas deferens accumulated more radioactivity in progesterone and dehydroepiandrosterone (DHEA) than the caput epididymidis. The levels of DHT, testosterone and 4-androstene-3, 17-dione in the caput epididymidis were reduced after ligation of ipselateral efferent ductules indicating the testicular origin of these steroids. The cauda epididymidis and vas deferens had higher levels of progesterone as compared to the other regions of the epididymis, which were decreased after the ligation. Intravenous injection of hCG increased the levels of oestradiol-17 beta in all tissues and markedly in the cauda epididymidis and vas deferens. The high levels of progesterone and oestradiol-17 beta present in these organs may be of importance in maintaining fertilizing ability of spermatozoa stored in the cauda epididymidis and vas deferens and their transport.

  13. Identification and changes in the seasonal concentrations of heat shock proteins in roe deer (Capreolus capreolus) epididymides.

    Science.gov (United States)

    Majewska, A M; Kordan, W; Koziorowska-Gilun, M; Wysocki, P

    2017-02-01

    Heat shock proteins (HSPs) act as molecular chaperones with important regulatory functions. HSPs are considered to be essential factors in animal reproduction. In view of seasonal variations in the secretory activity of the reproductive tract of mature roe deer (Capreolus capreolus), the aims of this study were to identify HSPs in the epididymides and compare the expression of the identified proteins in three periods of the reproductive season. Two-dimensional polyacrylamide gel electrophoresis revealed the highest number of polypeptides in homogenates of epididymal tissues and in caput, corpus and cauda epididymal fluids throughout the reproductive season. Epididymal tissue homogenates and epididymal fluids were analysed by tandem mass spectrometry (MS/MS) to reveal 31 polypeptides with enzymatic activity, including polypeptides with antioxidant properties, structural and cell signalling functions. Moreover, among the identified polypeptides, five of them were similar to heat shock proteins: endoplasmin (Grp94); heat shock protein 90 kDa (HSP90); 78-kDa glucose-regulated protein (Grp78); chain A, the crystal structure of the human HSP70 ATPase domain and heat shock protein beta-1 isoform X. The concentrations of the analysed polypeptides, expressed in optical density units (ODU), differed significantly (p ≤ .05) across the examined periods of the reproductive season. The highest ODU values for almost all analysed proteins were observed during the rutting period. The presence of HSPs in the epididymal tissues and fluids of roe deer in different periods of the reproductive season could indicate that those proteins play an important role in sperm maturation in the epididymis. © 2016 Blackwell Verlag GmbH.

  14. Determinação da razão ótima de espermatozóides por ovócitos de piabanha Brycon insignis (pisces - characidae Determination of the optimum ratio of spermatozoa per oocyte of the piabanha Brycon insignis

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