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Sample records for hormone-related protein affect

  1. Parathyroid hormone-related protein blood test

    Science.gov (United States)

    ... ency/article/003691.htm Parathyroid hormone-related protein blood test To use the sharing features on this page, ... measures the level of a hormone in the blood, called parathyroid hormone-related protein. How the Test is Performed A blood sample is needed . How ...

  2. Parathyroid hormone-related protein in preterm human milk.

    Science.gov (United States)

    Lubetzky, Ronit; Weisman, Yosef; Dollberg, Shaul; Herman, Lea; Mandel, Dror

    2010-04-01

    Parathyroid hormone-related protein (PTHrP) has the ability to activate parathyroid hormone receptors and cause hypercalcemia. High concentrations of PTHrP are found in human breastmilk of mothers of term-infants. It is not known whether PTHrP is excreted in preterm human milk. This study tested the hypothesis that PTHrP concentrations in milk obtained from mothers of preterm infants are similar to those found in milk from mothers of term infants. We collected samples of expressed human milk obtained from 27 mothers of preterm infants (27-34 weeks' gestation) and from 16 mothers of full-term infants. Samples were collected within the first 72 hours postpartum (colostrum) and again at 1 and 2 weeks postpartum. PTHrP concentrations in these samples were measured by two-site immunoradiometric assay. PTHrP concentrations were significantly higher in samples obtained after 1 week postpartum than in samples obtained during the first 72 hours of life in breastmilk obtained from mothers of both term and preterm infants (P milk. PTHrP concentrations after 2 weeks of lactation were significantly higher in samples obtained from mothers of term infants (P milk expressed by mothers of preterm infants contains amounts of PTHrP similar to those measured in milk expressed by mothers of term infants.

  3. Parathyroid Hormone-Related Protein Promotes Rat Stem Leydig Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Tiantian Song

    2017-11-01

    Full Text Available The regulatory factors for stem Leydig cell development are largely unknown. Herein, we reported that parathyroid hormone-related protein (PTHrP may be a factor to regulate this process. The effects of PTHrP on rat stem Leydig cell proliferation and differentiation were investigated using a stem Leydig cell culture system and an ethane dimethane sulfonate (EDS-treated in vivo Leydig cell regeneration model. PTHrP (1,000 pg/ml significantly increased medium testosterone level and up-regulated STAR, CYP17A1, and 17β-HSD3 expressions. Co-treatment with PKA inhibitor H-89 or PKC inhibitor U73122 reversed PTHrP-mediated increase of testosterone production in vitro. Intratesticular injection of PTHrP (100 ng/testis into the Leydig cell-depleted testis from post-EDS day 7 to 21 significantly increased serum testosterone level, up-regulated LHCGR, SCARB1, CYP11A1, 11β-HSD1, and CYP17A1 expressions. It also enlarged Leydig cell size without affecting PCNA-labeled Leydig cell number. This indicates that PTHrP promotes stem Leydig cell differentiation. PTHrP in vivo increased CREB and p-CREB levels, suggesting that PTHrP acts via a PKA-CREB signaling pathway. In conclusion, PTHrP stimulates stem Leydig cell differentiation without affecting its proliferation, showing its novel action and mechanism on rat stem Leydig cell development.

  4. Parathyroid hormone-related protein is a gravisensor in lung and bone cell biology

    Science.gov (United States)

    Torday, J. S.

    2003-10-01

    Parathyroid Hormone-related Protein (PTHrP) has been shown to be essential for the development and homeostatic regulation of lung and bone. Since both lung and bone structure and function are affected by microgravity, we hypothesized that 0 × g down-regulates PTHrP signaling. To test this hypothesis, we suspended lung and bone cells in the simulated microgravity environment of a Rotating Wall Vessel Bioreactor, which simulates microgravity, for up to 72 hours. During the first 8 hours of exposure to simulated 0 × g, PTHrP expression fell precipitously, decreasing by 80-90%; during the subsequent 64 hours, PTHrP expression remained at this newly established level of expression. PTHrP production decreased from 12 pg/ml/hour to 1 pg/ml/hour in culture medium from microgravity-exposed cells. The cells were then recultured at unit gravity for 24hours, and PTHrP expression and production returned to normal levels. Based on these findings, we have obtained bones from rats flown in space for 2 weeks (Mission STS-58, SL-2). Analysis of PTHrP expression by femurs and tibias from these animals (n=5) revealed that PTHrP expression was 60% lower than in bones from control ground-based rats. Interestingly, there were no differences in PTHrP expression by parietal bone from space-exposed versus ground-based animals, indicating that the effect of weightlessness on PTHrP expression is due to the unweighting of weight-bearing bones. This finding is consistent with other studies of microgravity-induced osteoporosis. The loss of the PTHrP signaling mechanism may be corrected using chemical agents that up-regulate this pathway. In conclusion, PTHrP represents a stretch-sensitive paracrine signaling mechanism that may sense gravity.

  5. Parathyroid hormone-related protein (pthrp) is a gravisensor for lung and bone.

    Science.gov (United States)

    Torday, J.

    Parathyroid Hormone-related Protein (PTHrP) and its receptor represent a stretch- sensitive paracrine signaling mechanism (Torday, 1999) that may sense gravity. PTHrP has been shown to be essential for the development and homeostatic regulation of lung (Rubin et al, 2000) and bone (Kronenberg et al, 1994). Since both lung and bone structure and function are affected by microgravity, we hypothesized that microgravity down-regulates PTHrP signaling. To test this hypothesis, we suspended lung and bone cells in the microgravity environment of a rotating wall vessel apparatus, which simulates microgravity, for up to 72 hours. During the first 6-8 hours, PTHrP expression fell precipitously, decreasing by 80-90%; during the subsequent 64-66 hours, PTHrP expression remained at this newly established level. PTHrP production decreased from 5 pmol/ml/3hours to undetectable levels in culture medium from microgravity-exposed cells. The cells were then put back in culture at unit gravity for 24hours, and PTHrP expression and production returned to normal levels. Based on these findings, we have obtained bones from rats flown in space for 2 weeks (mission SLS-2, provided courtesy of the Biospecimen Facility, Ames Research Center, NASA, as a result of a peer-reviewed proposal). Analysis of PTHrP expression by femurs and tibias from these animals (n=5) revealed that PTHrP expression was 60% lower than in bones from ground-based rats. Interestingly, there were no differences in PTHrP exp ression by parietal bones, indicating that the effect of weightlessness on PTHrP expression is due to the unweighting of weight-bearing bones. This finding is consistent with other studies of microgravity-induced osteoporosis. The loss of the PTHrP signaling mechanism may be corrected using chemical agents that up-regulate this pathway.

  6. Prognostic implications of parathyroid hormone-related protein in males and females with non--small-cell lung cancer.

    Science.gov (United States)

    Montgrain, Philippe R; Deftos, Leonard J; Arenberg, Douglas; Tipps, Ann; Quintana, Rick; Carskadon, Shannon; Hastings, Randolph H

    2011-05-01

    Non-small-cell lung carcinoma immunoreactivity for parathyroid hormone-related protein has been associated with increased survival in female patients but not in male patients. The current investigation attempted to substantiate this finding in 2 new patient groups. Patients were divided into groups with and without immunoreactivity for a carboxyl-terminal parathyroid hormone-related protein epitope assessed in deparaffinized sections by a blinded observer. One group included 85 female patients with stage I lung cancer, and the second group had 48 female and 66 male patients with stage I-IV lung cancer. Survival times were compared by the log-rank test between groups separated by tumor parathyroid hormone-related protein status. Parathyroid hormone-related protein was present in 70%-80% of the patients, independent of sex, stage, and smoking history. In the females with stage I lung cancer, parathyroid hormone-related protein increased median survival from 25 to 60 months (P < .05). In the second group, parathyroid hormone-related protein expression increased 48-month disease-free survival of female lung cancer patients from 44% to 63% (P < .05), but had no effect in male patients. Parathyroid hormone-related protein remained a significant, independent predictor when evaluated together with other covariates by Cox multivariate regression. This study verifies that parathyroid hormone-related protein is a sex-dependent survival factor for non-small-cell lung carcinoma, that it correlates with disease-free survival, and that the association with survival holds for women with early-stage disease as well as more advanced cancer. Thus, the protein could find use as a prognostic indicator and could be a target for therapy. Copyright © 2011 Elsevier Inc. All rights reserved.

  7. Pancreatic neuroendocrine cell tumor secreting parathyroid hormone-related protein and gastrin: Report of a case.

    Science.gov (United States)

    Morita, Yoshifumi; Suzuki, Shohachi; Sakaguchi, Takanori; Oishi, Kosuke; Suzuki, Atsushi; Fukumoto, Kazuhiko; Inaba, Keisuke; Baba, Satoshi; Takehara, Yasuo; Konno, Hiroyuki

    2010-12-01

    This report presents a case of pancreatic neuroendocrine cell carcinoma with multiple liver metastases secreting gastrin and parathyroid hormone-related protein (PTHrP) related to lumbar bone fracture and hypercalcemia. A 58-year-old woman visited an affiliated hospital with a chief complaint of lumbago without any evidence of trauma. She was diagnosed with hepatic dysfunction and hypercalcemia as well as multiple lumbar compression fractures without osteolytic lesions. Abdominal computed tomography (CT) showed a hypervascular mass in the pancreatic tail and multiple liver tumors. Duodenal ulcers were found with gastrointestinal endoscopy. There was a marked increase in the serum gastrin level. She was diagnosed as gastrinoma with multiple liver metastases and was admitted to the hospital. She had an increase in serum PTHrP level without the elevation of intact parathyroid hormone at the time of admission. She underwent an extended right hepatectomy in addition to a distal pancreatectomy with a regional lymphadenectomy and splenectomy. The postoperative course was uneventful, and serum gastrin and PTHrP activities reduced to normal levels. She remained symptom-free, and serum calcium, gastrin, and PTHrP levels remain within the normal ranges 19 months after surgery without adjuvant therapy.

  8. Role of Parathyroid Hormone-Related Protein Signaling in Chronic Pancreatitis

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    Falzon, Miriam, E-mail: mfalzon@utmb.edu; Bhatia, Vandanajay [Department of Pharmacology and Toxicology, University of Texas Medical Branch, Galveston, TX 77555 (United States)

    2015-06-18

    Chronic pancreatitis (CP), a progressive inflammatory disease where acini are destroyed and replaced by fibrous tissue, increases the risk for pancreatic cancer. Risk factors include alcohol, smoking, and obesity. The effects of these risk factors are exacerbated in patients with mutations in genes that predispose to CP. The different environmental and genetic factors produce the same clinical phenotype; once CP develops, disease course is the same regardless of etiology. Critical questions still need to be answered to understand what modifies predisposition to develop CP in persons exposed to risk factors. We postulate that risk factors modulate endogenous pathways, with parathyroid hormone-related protein (PTHrP) signaling being one such pathway. In support, PTHrP levels are elevated in mice treated with alcohol, and in mouse models of cerulein- and pancreatic duct ligation-induced CP. Disrupting the Pthrp gene in acinar cells exerts protective effects (decreased edema, histological damage, amylase and cytokine release, and fibrosis) in these CP models. PTHrP levels are elevated in human CP. Currently, CP care lacks specific pharmacological interventions. Targeting PTHrP signaling may present a novel therapeutic strategy that inhibits pancreatic inflammation and fibrosis, especially since the risk of developing pancreatic cancer is strongly associated with duration of chronic inflammation.

  9. Negative regulation of parathyroid hormone-related protein expression by steroid hormones

    Energy Technology Data Exchange (ETDEWEB)

    Kajitani, Takashi; Tamamori-Adachi, Mimi [Department of Biochemistry, Teikyo University School of Medicine, 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605 (Japan); Okinaga, Hiroko [Department of Internal Medicine, Teikyo University School of Medicine, 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605 (Japan); Chikamori, Minoru; Iizuka, Masayoshi [Department of Biochemistry, Teikyo University School of Medicine, 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605 (Japan); Okazaki, Tomoki, E-mail: okbgeni@med.teikyo-u.ac.jp [Department of Biochemistry, Teikyo University School of Medicine, 2-11-1 Kaga, Itabashi-ku, Tokyo 173-8605 (Japan)

    2011-04-15

    Highlights: {yields} Steroid hormones repress expression of PTHrP in the cell lines where the corresponding nuclear receptors are expressed. {yields} Nuclear receptors are required for suppression of PTHrP expression by steroid hormones, except for androgen receptor. {yields} Androgen-induced suppression of PTHrP expression appears to be mediated by estrogen receptor. -- Abstract: Elevated parathyroid hormone-related protein (PTHrP) is responsible for humoral hypercalcemia of malignancy (HHM), which is of clinical significance in treatment of terminal patients with malignancies. Steroid hormones were known to cause suppression of PTHrP expression. However, detailed studies linking multiple steroid hormones to PTHrP expression are lacking. Here we studied PTHrP expression in response to steroid hormones in four cell lines with excessive PTHrP production. Our study established that steroid hormones negatively regulate PTHrP expression. Vitamin D receptor, estrogen receptor {alpha}, glucocorticoid receptor, and progesterone receptor, were required for repression of PTHrP expression by the cognate ligands. A notable exception was the androgen receptor, which was dispensable for suppression of PTHrP expression in androgen-treated cells. We propose a pathway(s) involving nuclear receptors to suppress PTHrP expression.

  10. Role of Parathyroid Hormone-Related Protein Signaling in Chronic Pancreatitis.

    Science.gov (United States)

    Falzon, Miriam; Bhatia, Vandanajay

    2015-06-18

    Chronic pancreatitis (CP), a progressive inflammatory disease where acini are destroyed and replaced by fibrous tissue, increases the risk for pancreatic cancer. Risk factors include alcohol, smoking, and obesity. The effects of these risk factors are exacerbated in patients with mutations in genes that predispose to CP. The different environmental and genetic factors produce the same clinical phenotype; once CP develops, disease course is the same regardless of etiology. Critical questions still need to be answered to understand what modifies predisposition to develop CP in persons exposed to risk factors. We postulate that risk factors modulate endogenous pathways, with parathyroid hormone-related protein (PTHrP) signaling being one such pathway. In support, PTHrP levels are elevated in mice treated with alcohol, and in mouse models of cerulein- and pancreatic duct ligation-induced CP. Disrupting the Pthrp gene in acinar cells exerts protective effects (decreased edema, histological damage, amylase and cytokine release, and fibrosis) in these CP models. PTHrP levels are elevated in human CP. Currently, CP care lacks specific pharmacological interventions. Targeting PTHrP signaling may present a novel therapeutic strategy that inhibits pancreatic inflammation and fibrosis, especially since the risk of developing pancreatic cancer is strongly associated with duration of chronic inflammation.

  11. Role of Parathyroid Hormone-Related Protein Signaling in Chronic Pancreatitis

    Directory of Open Access Journals (Sweden)

    Miriam Falzon

    2015-06-01

    Full Text Available Chronic pancreatitis (CP, a progressive inflammatory disease where acini are destroyed and replaced by fibrous tissue, increases the risk for pancreatic cancer. Risk factors include alcohol, smoking, and obesity. The effects of these risk factors are exacerbated in patients with mutations in genes that predispose to CP. The different environmental and genetic factors produce the same clinical phenotype; once CP develops, disease course is the same regardless of etiology. Critical questions still need to be answered to understand what modifies predisposition to develop CP in persons exposed to risk factors. We postulate that risk factors modulate endogenous pathways, with parathyroid hormone-related protein (PTHrP signaling being one such pathway. In support, PTHrP levels are elevated in mice treated with alcohol, and in mouse models of cerulein- and pancreatic duct ligation-induced CP. Disrupting the Pthrp gene in acinar cells exerts protective effects (decreased edema, histological damage, amylase and cytokine release, and fibrosis in these CP models. PTHrP levels are elevated in human CP. Currently, CP care lacks specific pharmacological interventions. Targeting PTHrP signaling may present a novel therapeutic strategy that inhibits pancreatic inflammation and fibrosis, especially since the risk of developing pancreatic cancer is strongly associated with duration of chronic inflammation.

  12. Parathyroid hormone-related protein and calcium regulation in vitamin D-deficient sea bream (Sparus auratus).

    NARCIS (Netherlands)

    Abbink, W.; Hang, X.M.; Guerreiro, P.M.; Spanings, F.A.T.; Ross, H.A.; Canario, A.V.; Flik, G.

    2007-01-01

    Gilthead sea bream (Sparus auratus L.) were fed a vitamin D-deficient diet for 22 weeks. Growth rate, whole body mineral pools and calcium balance were determined. Plasma parathyroid hormone-related protein (PTHrP) and calcitriol levels were assessed. Expression of mRNA for pthrp and pth1r was

  13. Parathyroid hormone related-protein promotes epithelial-to-mesenchymal transition in prostate cancer.

    Directory of Open Access Journals (Sweden)

    Weg M Ongkeko

    Full Text Available Parathyroid hormone-related protein (PTHrP possesses a variety of physiological and developmental functions and is also known to facilitate the progression of many common cancers, notably their skeletal invasion, primarily by increasing bone resorption. The purpose of this study was to determine whether PTHrP could promote epithelial-to-mesenchymal transition (EMT, a process implicated in cancer stem cells that is critically involved in cancer invasion and metastasis. EMT was observed in DU 145 prostate cancer cells stably overexpressing either the 1-141 or 1-173 isoform of PTHrP, where there was upregulation of Snail and vimentin and downregulation of E-cadherin relative to parental DU 145. By contrast, the opposite effect was observed in PC-3 prostate cancer cells where high levels of PTHrP were knocked-down via lentiviral siRNA transduction. Increased tumor progression was observed in PTHrP-overexpressing DU 145 cells while decreased progression was observed in PTHrP-knockdown PC-3 cells. PTHrP-overexpressing DU 145 formed larger tumors when implanted orthoptopically into nude mice and in one case resulted in spinal metastasis, an effect not observed among mice injected with parental DU 145 cells. PTHrP-overexpressing DU 145 cells also caused significant bone destruction when injected into the tibiae of nude mice, while parental DU 145 cells caused little to no destruction of bone. Together, these results suggest that PTHrP may work through EMT to promote an aggressive and metastatic phenotype in prostate cancer, a pathway of importance in cancer stem cells. Thus, continued efforts to elucidate the pathways involved in PTHrP-induced EMT as well as to develop ways to specifically target PTHrP signaling may lead to more effective therapies for prostate cancer.

  14. The expression of parathyroid hormone-related protein mRNA and immunoreactive protein in human amnion and choriodecidua is increased at term compared with preterm gestation

    NARCIS (Netherlands)

    Curtis, NE; Ho, PWM; King, RG; Farrugia, W; Moses, EK; Gillespie, MT; Moseley, JM; Rice, GE; Wlodek, ME

    Parathyroid hormone-related protein (PTHrP) gene expression and/or immunoreactive protein have previously been identified in the uterus and intrauterine gestational tissues. The putative roles of PTHrP during pregnancy include vasodilatation, regulation of placental calcium transfer, uterine smooth

  15. [Seasonal rhythms of parathyroid hormone-related protein (PTHrP) expression in growing rats after functional mandibular protrusion].

    Science.gov (United States)

    Li, Jiang-Ning; Chen, Yang-Xi; Wang, Zheng-Rong

    2006-04-01

    To study parathyroid hormone-related protein (PTHrP) expression during forward mandibular positioning and compare it with the expression during natural growth in different seasons. Sixty-four SD rats were randomly divided into experimental and control groups. Each group were randomly divided into four groups according to seasons. Immunohistochemical (IHC) methods were used to test the protein expression of PTHrP. Macroscopic and microscopic approach were applied to analyze the results. PTHrP expressed in mandibular condylar cartilage (MCC), the expression was accelerated and enhanced when the mandible was positioned forward. Furthermore, there was a seasonal rhythm in the protein expression of PTHrP in both experimental and control groups. The protein expression in spring group rose more than other groups. The functional appliance therapy can enhance the protein expression of PTHrP. The enhancement has a seasonal rhythm, which indicates that for the functional treatment better results can be achieved in spring.

  16. Recombinant human parathyroid hormone related protein 1-34 and 1-84 and their roles in osteoporosis treatment.

    Directory of Open Access Journals (Sweden)

    Hua Wang

    Full Text Available Osteoporosis is a common disorder characterized by compromised bone strength that predisposes patients to increased fracture risk. Parathyroid hormone related protein (PTHrP is one of the candidates for clinical osteoporosis treatment. In this study, GST Gene Fusion System was used to express recombinant human PTHrP (hPTHrP 1-34 and 1-84. To determine whether the recombinant hPTHrP1-34 and 1-84 can enhance renal calcium reabsorption and promote bone formation, we examined effects of recombinant hPTHrP1-34 and 1-84 on osteogenic lineage commitment in a primary bone marrow cell culture system and on osteoporosis treatment. Results revealed that both of recombinant hPTHrP1-34 and 1-84 increased colony formation and osteogenic cell differentiation and mineralization in vitro; however, the effect of recombinant hPTHrP1-84 is a little stronger than that of hPTHrP1-34. Next, ovariectomy was used to construct osteoporosis animal model (OVX to test activities of these two recombinants in vivo. HPTHrP1-84 administration elevated serum calcium by up-regulating the expression of renal calcium transporters, which resulted in stimulation of osteoblastic bone formation. These factors contributed to augmented bone mass in hPTHrP1-84 treated OVX mice but did not affect bone resorption. There was no obvious bone mass alteration in hPTHrP1-34 treated OVX mice, which may be, at least partly, associated with shorter half-life of hPTHrP1-34 compared to hPTHrP1-84 in vivo. This study implies that recombinant hPTHrP1-84 is more effective than hPTHrP1-34 to enhance renal calcium reabsorption and to stimulate bone formation in vivo.

  17. Current Nomenclature of Pseudohypoparathyroidism: Inactivating Parathyroid Hormone/Parathyroid Hormone-Related Protein Signaling Disorder.

    Science.gov (United States)

    Turan, Serap

    2017-12-30

    Disorders related to parathyroid hormone (PTH) resistance and PTH signaling pathway impairment are historically classified under the term of pseudohypoparathyroidism (PHP). The disease was first described and named by Fuller Albright and colleagues in 1942. Albright hereditary osteodystrophy (AHO) is described as an associated clinical entity with PHP, characterized by brachydactyly, subcutaneous ossifications, round face, short stature and a stocky build. The classification of PHP is further divided into PHP-Ia, pseudo-PHP (pPHP), PHP-Ib, PHP-Ic and PHP-II according to the presence or absence of AHO, together with an in vivo response to exogenous PTH and the measurement of Gsα protein activity in peripheral erythrocyte membranes in vitro. However, PHP classification fails to differentiate all patients with different clinical and molecular findings for PHP subtypes and classification become more complicated with more recent molecular characterization and new forms having been identified. So far, new classifications have been established by the EuroPHP network to cover all disorders of the PTH receptor and its signaling pathway. Inactivating PTH/PTH-related protein signaling disorder (iPPSD) is the new name proposed for a group of these disorders and which can be further divided into subtypes - iPPSD1 to iPPSD6. These are termed, starting from PTH receptor inactivation mutation (Eiken and Blomstrand dysplasia) as iPPSD1, inactivating Gsα mutations (PHP-Ia, PHP-Ic and pPHP) as iPPSD2, loss of methylation of GNAS DMRs (PHP-Ib) as iPPSD3, PRKAR1A mutations (acrodysostosis type 1) as iPPSD4, PDE4D mutations (acrodysostosis type 2) as iPPSD5 and PDE3A mutations (autosomal dominant hypertension with brachydactyly) as iPPSD6. iPPSDx is reserved for unknown molecular defects and iPPSDn+1 for new molecular defects which are yet to be described. With these new classifications, the aim is to clarify the borders of each different subtype of disease and make the classification

  18. Parathyroid hormone-related protein: potential therapeutic target for melanoma invasion and metastasis.

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    Huang, Dao Chao; Yang, Xian Fang; Ochietti, Benoît; Fadhil, Ibtihal; Camirand, Anne; Kremer, Richard

    2014-10-01

    The role of PTHrP in the highly metastatic human melanoma disease is not known. This study investigates the mechanisms of action of this secreted factor through homozygous inactivation of the Pthrp gene in A375 human melanoma cells. In vitro, Pthrp-ablated cells (knockout [KO]-A375, -/-) showed decreased motility and anchorage-independent growth, rounder morphology, and a significant reduction in invasion capacity compared with nonablated A375 cells (wild-type [WT]-A375, +/+). PTHrP peptide 1-34 and conditioned medium from WT-A375 cells partially restored the invasive phenotype in KO-A375. Pthrp ablation substantially decreased actin polymerization, matrix metallopeptidase 9 expression and focal adhesion kinase phosphorylation. In vivo, green fluorescent protein-transduced ablated and nonablated A375 cells were injected intracardially or sc into nude mice to study proliferation and multiorgan metastasis. Dissemination of injected Pthrp-ablated cells to lung and liver was reduced by 85% and 50%, respectively, compared with nonablated controls (120 hours after injection). The number of metastatic lesions and the percentage of animals with metastasis were markedly lower in mice injected with Pthrp-ablated A375, and 45% of these animals survived a 7-week period compared with 15% of mice injected with nonablated WT-A375. When mice injected with WT-A375 were treated with our blocking anti-PTHrP monoclonal antibody raised against the first 33 amino acids of human PTHrP, tumor size was decreased by more than 80% over 4 weeks and survival was significantly improved over 8 months. This study provides direct evidence of the major role for PTHrP in melanoma invasion and metastasis and suggests that agents that suppress PTHrP may be beneficial against melanoma progression.

  19. Switching of G-protein Usage by the Calcium-sensing Receptor Reverses Its Effect on Parathyroid Hormone-related Protein Secretion in Normal Versus Malignant Breast Cells*

    OpenAIRE

    Mamillapalli, Ramanaiah; VanHouten, Joshua; Zawalich, Walter; Wysolmerski, John

    2008-01-01

    The calcium-sensing receptor (CaR) is a G-protein-coupled receptor that signals in response to extracellular calcium and regulates parathyroid hormone secretion. The CaR is also expressed on normal mammary epithelial cells (MMECs), where it has been shown to inhibit secretion of parathyroid hormone-related protein (PTHrP) and participate in the regulation of calcium and bone metabolism during lactation. In contrast to normal breast cells, the CaR has been reported to s...

  20. Structural requirements for the action of parathyroid hormone-related protein (PTHrP) on bone resorption by isolated osteoclasts

    Energy Technology Data Exchange (ETDEWEB)

    Evely, R.S.; Bonomo, A.; Schneider, H.G.; Moseley, J.M.; Gallagher, J.; Martin, T.J. (St. Vincent' s Institute of Medical Research, Melbourne (Australia))

    1991-01-01

    Parathyroid hormone-related protein (PTHrP) plays a major role in the syndrome of humoral hypercalcemia of malignancy (HHM) by its actions on bone and kidney. In this study an isolated osteoclast bone resorption assay was used to investigate the actions of this peptide and the structure-activity relationships for its resorption effect. As with PTH, neither synthetic nor recombinant PTHrP preparations stimulated resorption within highly purified osteoclast populations. Resorption was stimulated only in the presence of contaminating osteoblasts or in cocultures with the osteoblast-like cell line UMR-106. In the presence of osteoblasts PTHrP-(1-34) and PTHrP-(1-84) stimulated bone resorption in a dose-dependent manner with a potency comparable to that of PTH-(1-34) on a molar basis. The biologic activity of the PTHrP was shown to reside in the first 34 amino acids, and within that region the structural requirements for promotion of osteoclastic resorption resembled closely those for promotion of cyclic AMP formation in osteoblast-like cells. Using emulsion autoradiography with iodinated PTHrP-(1-34) and PTHrP-(1-84) on mixed bone cell preparations from neonatal rats, specific binding was demonstrated only to osteoblasts, not to osteoclasts. These results clearly demonstrate that PTHrP is a potent stimulator of bone resorption and that these effects are, like those of PTH, mediated by initial actions upon cells of the osteoblast lineage.

  1. Bone-Invasive Oral Squamous Cell Carcinoma in Cats: Pathology and Expression of Parathyroid Hormone-Related Protein

    Science.gov (United States)

    Martin, C. K.; Tannehill-Gregg, S. H.; Wolfe, T. D.; Rosol, T. J.

    2014-01-01

    Feline oral squamous cell carcinoma (OSCC) is the most common oral tumor in cats. There is no effective treatment, and the average duration of survival after diagnosis is only 2 months. Feline OSCC is frequently associated with osteolysis; however, the mechanisms responsible are unknown. The objective of this study was to characterize the epidemiology and pathology of bone-invasive OSCC in cats and to determine the expression of select bone resorption agonists. In sum, 451 cases of feline OSCC were evaluated. There was no sex or breed predisposition, although there were more intact cats in the OSCC group compared to the control group. Gingiva was the most common site, followed by the sublingual region and tongue. Cats with lingual OSCC were younger (mean, 11.9 years) compared to cats with gingival OSCC (mean, 13.6 years). In addition to osteolysis, there was periosteal new bone formation, osseous metaplasia of tumor stroma, and direct apposition of OSCC to fragments of bone, suggestive of bone-binding behavior. Eighty-two cases were selected for immunohistochemical detection of parathyroid hormone-related protein (PTHrP). Specimens with osteolysis had increased PTHrP expression and nuclear localization, compared to OSCC without osteolysis. Thirty-eight biopsies of OSCC with osteolysis were evaluated for tumor necrosis factor α expression, and only 4 biopsies had such expression in a small proportion of tumor cells. Increased tumor expression of PTHrP and increased localization of PTHrP to the nucleus were associated with osteolysis and may play an important role in bone resorption and tumor invasion in cats with OSCC. PMID:20940448

  2. A carboxyl leucine-rich region of parathyroid hormone-related protein is critical for nuclear export.

    Science.gov (United States)

    Pache, Jared C; Burton, Douglas W; Deftos, Leonard J; Hastings, Randolph H

    2006-02-01

    PTHrP is an oncofetal protein with distinct proliferative and antiapoptotic roles that are affected by nucleocytoplasmic shuttling. The protein's nuclear export is sensitive to leptomycin B, consistent with a chromosome region maintenance protein 1-dependent pathway. We determined that the 109-139 region of PTHrP was involved in its nuclear export by demonstrating that a C-terminal truncation mutant, residues 1-108, exports at a reduced rate, compared with the wild-type 139 amino acid isoform. We searched for potential nuclear export sequences within the 109-139 region, which is leucine rich. Comparisons with established nuclear export sequences identified a putative consensus signal at residues 126-136. Deletion of this region resulted in nuclear export characteristics that closely matched those of the C-terminal truncation mutant. Confocal microscopic analyses of transfected 293, COS-1, and HeLa cells showed that steady-state nuclear levels of the truncated and deletion mutants were significantly greater than levels of wild-type PTHrP and were unaffected by leptomycin B, unlike the wild-type protein. In addition, both mutants demonstrated greatly reduced nuclear export with assays using nuclear preparations and intact cells. Based on these results, we conclude that the 126-136 amino acid sequence closely approximates the structure of a chromosome region maintenance protein 1-dependent leucine-rich nuclear export signal and is critical for nuclear export of PTHrP.

  3. Parathyroid hormone-related protein is a hypertrophy factor for human mesangial cells: Implications for diabetic nephropathy.

    Science.gov (United States)

    Ortega, Arantxa; Romero, Montserrat; Izquierdo, Adriana; Troyano, Nuria; Arce, Yolanda; Ardura, Juan Antonio; Arenas, María Isabel; Bover, Jordi; Esbrit, Pedro; Bosch, Ricardo J

    2012-05-01

    Hypertrophy of human mesangial cells (HMC) is among the earliest characteristics in patients with diabetic nephropathy (DN). Recently, we observed the upregulation of parathyroid hormone (PTH)-related protein (PTHrP) in experimental DN, associated with renal hypertrophy. Herein, we first examined whether PTHrP was overexpressed in human DN, and next assessed the putative role of this protein on high glucose (HG)-induced HMC hypertrophy. As previously found in mice, kidneys from diabetic patients showed an increased tubular and glomerular immunostaining for PTHrP. In HMC, HG medium increased PTHrP protein expression associated with the development of hypertrophy as assessed by cell protein content. This effect was also induced by PTHrP(1-36). HG and PTHrP(1-36)-induced hypertrophy were associated with an increase in cyclin D1 and p27Kip1 protein expression, a decreased cyclin E expression, and the prevention of cyclin E/cdk2 complex activation. Both PTHrP neutralizing antiserum (α-PTHrP) and the PTH/PTHrP receptor antagonist (JB4250) were able to abolish HG induction of hypertrophy, the aforementioned changes in cell cycle proteins, and also TGF-β1 up-regulation. Moreover, the capability of both HG and PTHrP(1-36) to induce HMC hypertrophy was abolished by α-TGFβ1. These data show for the first time that PTHrP is upregulated in the kidney of patients with DN. Our findings also demonstrate that PTHrP acts as an important mediator of HG-induced HMC hypertrophy by modulating cell cycle regulatory proteins and TGF-β1. Copyright © 2011 Wiley Periodicals, Inc.

  4. Crystallization of the receptor-binding domain of parathyroid hormone-related protein in complex with a neutralizing monoclonal antibody Fab fragment

    Energy Technology Data Exchange (ETDEWEB)

    McKinstry, William J.; Polekhina, Galina; Diefenbach-Jagger, Hannelore; Sato, Koh; Onuma, Etsuro; Gillespie, Matthew T.; Martin, Thomas J.; Parker, Michael W.; (SVIMR-A); (Chugai); (Melbourne)

    2009-04-01

    Parathyroid hormone-related protein (PTHrP) plays an important role in regulating embryonic skeletal development and is abnormally regulated in the pathogenesis of skeletal complications observed with many cancers and osteoporosis. It exerts its action through binding to a G-protein-coupled seven-transmembrane cell-surface receptor (GPCR). Structurally, GPCRs are very difficult to study by X-ray crystallography. In this study, a monoclonal antibody Fab fragment which recognizes the same region of PTHrP as its receptor, PTH1R, was used to aid in the crystallization of PTHrP. The resultant protein complex was crystallized using the hanging-drop vapour-diffusion method with polyethylene glycol as a precipitant. The crystals belonged to the orthorhombic space group P2{sub 1}2{sub 1}2, with unit-cell parameters a = 72.6, b = 96.3, c = 88.5 {angstrom}, and diffracted to 2.0 {angstrom} resolution using synchrotron radiation. The crystal structure will shed light on the nature of the key residues of PTHrP that interact with the antibody and will provide insights into how the antibody is able to discriminate between PTHrP and the related molecule parathyroid homone.

  5. A Transgenic Mouse Model for Studying the Role of the Parathyroid Hormone-Related Protein System in Renal Injury

    Science.gov (United States)

    Bosch, Ricardo J.; Ortega, Arantxa; Izquierdo, Adriana; Arribas, Ignacio; Bover, Jordi; Esbrit, Pedro

    2011-01-01

    Parathyroid hormone- (PTH-) related protein (PTHrP) and its receptor, the PTH1 receptor (PTH1R), are widely expressed in the kidney, where PTHrP exerts a modulatory action on renal function. PTHrP is known to be upregulated in several experimental nephropathies such as acute renal failure (ARF), obstructive nephropathy (ON) as well as diabetic nephropathy (DN). In this paper, we will discuss the functional consequences of chronic PTHrP overexpression in the damaged kidney using a transgenic mouse strain overexpressing PTHrP in the renal proximal tubule. In both ARF and ON, PTHrP displays proinflammatory and profibrogenic actions including the induction of epithelia to mesenquima transition. Moreover, PTHrP participates in the mechanisms of renal hypertrophy as well as proteinuria in experimental DN. Angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be, at least in part, responsible for endogenous PTHrP upregulation in these pathophysiological settings. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches. PMID:21052497

  6. A Transgenic Mouse Model for Studying the Role of the Parathyroid Hormone-Related Protein System in Renal Injury

    Directory of Open Access Journals (Sweden)

    Ricardo J. Bosch

    2011-01-01

    Full Text Available Parathyroid hormone- (PTH- related protein (PTHrP and its receptor, the PTH1 receptor (PTH1R, are widely expressed in the kidney, where PTHrP exerts a modulatory action on renal function. PTHrP is known to be upregulated in several experimental nephropathies such as acute renal failure (ARF, obstructive nephropathy (ON as well as diabetic nephropathy (DN. In this paper, we will discuss the functional consequences of chronic PTHrP overexpression in the damaged kidney using a transgenic mouse strain overexpressing PTHrP in the renal proximal tubule. In both ARF and ON, PTHrP displays proinflammatory and profibrogenic actions including the induction of epithelia to mesenquima transition. Moreover, PTHrP participates in the mechanisms of renal hypertrophy as well as proteinuria in experimental DN. Angiotensin II (Ang II, a critical factor in the progression of renal injury, appears to be, at least in part, responsible for endogenous PTHrP upregulation in these pathophysiological settings. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.

  7. Novel Role of Parathyroid Hormone-Related Protein in the Pathophysiology of the Diabetic Kidney: Evidence from Experimental and Human Diabetic Nephropathy

    Science.gov (United States)

    Romero, Montserrat; Ortega, Arantxa; Olea, Nuria; Arenas, María Isabel; Izquierdo, Adriana; Bover, Jordi; Esbrit, Pedro

    2013-01-01

    Parathyroid hormone-related protein (PTHrP) and its receptor type 1 (PTH1R) are extensively expressed in the kidney, where they are able to modulate renal function. Renal PTHrP is known to be overexpressed in acute renal injury. Recently, we hypothesized that PTHrP involvement in the mechanisms of renal injury might not be limited to conditions with predominant damage of the renal tubulointerstitium and might be extended to glomerular diseases, such as diabetic nephropathy (DN). In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria. More recent data have shown, for the first time, that PTHrP is upregulated in the kidney from patients with DN. Collectively, animal and human studies have shown that PTHrP acts as an important mediator of diabetic renal cell hypertrophy by a mechanism which involves the modulation of cell cycle regulatory proteins and TGF-β1. Furthermore, angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be responsible for PTHrP upregulation in these conditions. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches. PMID:23984429

  8. Novel Role of Parathyroid Hormone-Related Protein in the Pathophysiology of the Diabetic Kidney: Evidence from Experimental and Human Diabetic Nephropathy

    Directory of Open Access Journals (Sweden)

    Montserrat Romero

    2013-01-01

    Full Text Available Parathyroid hormone-related protein (PTHrP and its receptor type 1 (PTH1R are extensively expressed in the kidney, where they are able to modulate renal function. Renal PTHrP is known to be overexpressed in acute renal injury. Recently, we hypothesized that PTHrP involvement in the mechanisms of renal injury might not be limited to conditions with predominant damage of the renal tubulointerstitium and might be extended to glomerular diseases, such as diabetic nephropathy (DN. In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria. More recent data have shown, for the first time, that PTHrP is upregulated in the kidney from patients with DN. Collectively, animal and human studies have shown that PTHrP acts as an important mediator of diabetic renal cell hypertrophy by a mechanism which involves the modulation of cell cycle regulatory proteins and TGF-β1. Furthermore, angiotensin II (Ang II, a critical factor in the progression of renal injury, appears to be responsible for PTHrP upregulation in these conditions. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.

  9. Novel role of parathyroid hormone-related protein in the pathophysiology of the diabetic kidney: evidence from experimental and human diabetic nephropathy.

    Science.gov (United States)

    Romero, Montserrat; Ortega, Arantxa; Olea, Nuria; Arenas, María Isabel; Izquierdo, Adriana; Bover, Jordi; Esbrit, Pedro; Bosch, Ricardo J

    2013-01-01

    Parathyroid hormone-related protein (PTHrP) and its receptor type 1 (PTH1R) are extensively expressed in the kidney, where they are able to modulate renal function. Renal PTHrP is known to be overexpressed in acute renal injury. Recently, we hypothesized that PTHrP involvement in the mechanisms of renal injury might not be limited to conditions with predominant damage of the renal tubulointerstitium and might be extended to glomerular diseases, such as diabetic nephropathy (DN). In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria. More recent data have shown, for the first time, that PTHrP is upregulated in the kidney from patients with DN. Collectively, animal and human studies have shown that PTHrP acts as an important mediator of diabetic renal cell hypertrophy by a mechanism which involves the modulation of cell cycle regulatory proteins and TGF- β 1. Furthermore, angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be responsible for PTHrP upregulation in these conditions. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.

  10. Role of parathyroid hormone-related protein in the pro-inflammatory and pro-fibrogenic response associated with acute pancreatitis.

    Science.gov (United States)

    Bhatia, Vandanajay; Kim, Sung O K; Aronson, Judith F; Chao, Celia; Hellmich, Mark R; Falzon, Miriam

    2012-04-10

    Pancreatitis is a common and potentially lethal necro-inflammatory disease with both acute and chronic manifestations. Current evidence suggests that the accumulated damage incurred during repeated bouts of acute pancreatitis (AP) can lead to chronic disease, which is associated with an increased risk of pancreatic cancer. While parathyroid hormone-related protein (PTHrP) exerts multiple effects in normal physiology and disease states, its function in pancreatitis has not been previously addressed. Here we show that PTHrP levels are transiently elevated in a mouse model of cerulein-induced AP. Treatment with alcohol, a risk factor for both AP and chronic pancreatitis (CP), also increases PTHrP levels. These effects of cerulein and ethanol are evident in isolated primary acinar and stellate cells, as well as in the immortalized acinar and stellate cell lines AR42J and irPSCc3, respectively. Ethanol sensitizes acinar and stellate cells to the PTHrP-modulating effects of cerulein. Treatment of acinar cells with PTHrP (1-36) increases expression of the inflammatory mediators interleukin-6 (IL-6) and intracellular adhesion protein (ICAM-1), suggesting a potential autocrine loop. PTHrP also increases apoptosis in AR42J cells. Stellate cells mediate the fibrogenic response associated with pancreatitis; PTHrP (1-36) increases procollagen I and fibronectin mRNA levels in both primary and immortalized stellate cells. The effects of cerulein and ethanol on levels of IL-6 and procollagen I are suppressed by the PTH1R antagonist, PTHrP (7-34). Together these studies identify PTHrP as a potential mediator of the inflammatory and fibrogenic responses associated with alcoholic pancreatitis. Copyright © 2011 Elsevier B.V. All rights reserved.

  11. Treatment with N- and C-Terminal Peptides of Parathyroid Hormone-Related Protein Partly Compensate the Skeletal Abnormalities in IGF-I Deficient Mice

    Science.gov (United States)

    Portal-Núñez, Sergio; Murillo-Cuesta, Silvia; Lozano, Daniel; Cediel, Rafael; Esbrit, Pedro

    2014-01-01

    Insulin-like growth factor-I (IGF-I) deficiency causes growth delay, and IGF-I has been shown to partially mediate bone anabolism by parathyroid hormone (PTH). PTH-related protein (PTHrP) is abundant in bone, and has osteogenic features by poorly defined mechanisms. We here examined the capacity of PTHrP (1–36) and PTHrP (107–111) (osteostatin) to reverse the skeletal alterations associated with IGF-I deficiency. Igf1-null mice and their wild type littermates were treated with each PTHrP peptide (80 µg/Kg/every other day/2 weeks; 2 males and 4 females for each genotype) or saline vehicle (3 males and 3 females for each genotype). We found that treatment with either PTHrP peptide ameliorated trabecular structure in the femur in both genotypes. However, these peptides were ineffective in normalizing the altered cortical structure at this bone site in Igf1-null mice. An aberrant gene expression of factors associated with osteoblast differentiation and function, namely runx2, osteoprotegerin/receptor activator of NF-κB ligand ratio, Wnt3a, cyclin D1, connexin 43, catalase and Gadd45, as well as in osteocyte sclerostin, was found in the long bones of Igf1-null mice. These mice also displayed a lower amount of trabecular osteoblasts and osteoclasts in the tibial metaphysis than those in wild type mice. These alterations in Igf1-null mice were only partially corrected by each PTHrP peptide treatment. The skeletal expression of Igf2, Igf1 receptor and Irs2 was increased in Igf1-null mice, and this compensatory profile was further improved by treatment with each PTHrP peptide related to ERK1/2 and FoxM1 activation. In vitro, PTHrP (1–36) and osteostatin were effective in promoting bone marrow stromal cell mineralization in normal mice but not in IGF-I-deficient mice. Collectively, these findings indicate that PTHrP (1–36) and osteostatin can exert several osteogenic actions even in the absence of IGF-I in the mouse bone. PMID:24503961

  12. Can Supersaturation Affect Protein Crystal Quality?

    Science.gov (United States)

    Gorti, Sridhar

    2013-01-01

    In quiescent environments (microgravity, capillary tubes, gels) formation of a depletion zone is to be expected, due either to limited sedimentation, density driven convection or a combination of both. The formation of a depletion zone can: Modify solution supersaturation near crystal; Give rise to impurity partitioning. It is conjectured that both supersaturation and impurity partitioning affect protein crystal quality and size. Further detailed investigations on various proteins are needed to assess above hypothesis.

  13. Can Solution Supersaturation Affect Protein Crystal Quality?

    Science.gov (United States)

    Gorti, Sridhar

    2013-01-01

    The formation of large protein crystals of "high quality" is considered a characteristic manifestation of microgravity. The physical processes that predict the formation of large, high quality protein crystals in the microgravity environment of space are considered rooted in the existence of a "depletion zone" in the vicinity of crystal. Namely, it is considered reasonable that crystal quality suffers in earth-grown crystals as a result of the incorporation of large aggregates, micro-crystals and/or large molecular weight "impurities", processes which are aided by density driven convective flow or mixing at the crystal-liquid interface. Sedimentation and density driven convection produce unfavorable solution conditions in the vicinity of the crystal surface, which promotes rapid crystal growth to the detriment of crystal size and quality. In this effort, we shall further present the hypothesis that the solution supersaturatoin at the crystal surface determines the growth mechanism, or mode, by which protein crystals grow. It is further hypothesized that protein crystal quality is affected by the mechanism or mode of crystal growth. Hence the formation of a depletion zone in microgravity environment is beneficial due to inhibition of impurity incorporatoin as well as preventing a kinetic roughening transition. It should be noted that for many proteins the magnitude of neither protein crystal growth rates nor solution supersaturation are predictors of a kinetic roughening transition. That is, the kinetic roughening transition supersaturation must be dtermined for each individual protein.

  14. DROUGHT AFFECTS PROTEIN AND PHENOLIC CONTENT IN ...

    African Journals Online (AJOL)

    userpc

    ABSTRACT. Bambara groundnut (Vigna subterranea L. Verdc.) is a legume crop, which has long been recognised as a protein-rich and drought-tolerant crop, used extensively in sub-Saharan. Africa. This study evaluates the effect of experimental water deficit stress on total protein concentration, secondary protein structure ...

  15. Circulating forms of immunoreactive parathyroid hormone-related protein for identifying patients with humoral hypercalcemia of malignancy. A comparative study with C-terminal (109-141)- and N-terminal (1-86)-region-specific PTHrP radioassay

    Energy Technology Data Exchange (ETDEWEB)

    Suehiro, Mitsuko; Murakami, Minoru; Fukuchi, Minoru (Hyogo Coll. of Medicine, Nishinomiya (Japan))

    1994-11-01

    We evaluated the circulating forms of immunoreactive parathyroid hormone-related protein(PTHrP) in 115 healthy subjects and 122 patients with malignant diseases by using radioassay systems (RAS) specific for the C-terminal (109-141) fragment of PTHrP (C-RAS) and for the N-terminal(1-86) (N-RAS). PTHrP levels in healthy controls ranged from 1.5 to 38.2 (mean: 24.5) pmol/L with the C-RAS and from 0.9 to 2.5 (mean: 1.7) pmol/L with the N-RAS. The ratio of circulating N-terminal fragment (N) to C-terminal fragment (C) of PTHrP was calculated to be about 1 : 14.4 in the healthy subjects. Of the 122 patients with malignant diseases, 40 (32.8%) had circulating PTHrP levels undetectable with the N-RAS, but only 11 (9.0%) patients had levels undetectable with the C-RAS. Of the former 122 patients, 41 (33.6%) had high PTHrP as determined with the C-RAS, and 10 (8.2%) had high PTHrP as determined with the N-RAS. The former of these included only 8 (19.5%) humoral hypercalcemia malignancy(HHM) patients, while the latter included 8 (80.0%) HHM patients. The circulating N to C ratio was about 1 : 70.7 in the HHM patients. The N and C obtained with the different RASs showed a close correlation (r=0.86). The values also showed a close correlation with serum Ca; r=0.75 for C-RAS and r=0.81 for N-RAS. In addition, the correlation between the PTHrP reading obtained with the different RASs and serum Cr were: r=0.42 with C-RAS and r=0.26 with N-RAS. The circulating form of immunoreactive PTHrP fragments is therefore comprised mainly of PTHrP (109-141). In contrast, circulating concentrations of the PTHrP (1-86) fragment are very low, but detection of the PTHrP (1-86) fragment with the N-RAS is a more useful indicator of HHM with fewer false positive results and is less likely to be influenced by renal function than the detection of the PHPrP (109-141) fragment with C-RAS. (author).

  16. Inactivation of Tor proteins affects the dynamics of endocytic proteins ...

    Indian Academy of Sciences (India)

    Tor2 is an activator of the Rom2/Rho1 pathway that regulates -factor internalization. Since the recruitment of endocytic proteins such as actin-binding proteins and the amphiphysins precedes the internalization of -factor, we hypothesized that loss of Tor function leads to an alteration in the dynamics of the endocytic ...

  17. Phosphatidylcholine Affects Inner Membrane Protein Translocases of Mitochondria.

    Science.gov (United States)

    Schuler, Max-Hinderk; Di Bartolomeo, Francesca; Mårtensson, Christoph U; Daum, Günther; Becker, Thomas

    2016-09-02

    Two protein translocases transport precursor proteins into or across the inner mitochondrial membrane. The presequence translocase (TIM23 complex) sorts precursor proteins with a cleavable presequence either into the matrix or into the inner membrane. The carrier translocase (TIM22 complex) inserts multispanning proteins into the inner membrane. Both protein import pathways depend on the presence of a membrane potential, which is generated by the activity of the respiratory chain. The non-bilayer-forming phospholipids cardiolipin and phosphatidylethanolamine are required for the activity of the respiratory chain and therefore to maintain the membrane potential for protein import. Depletion of cardiolipin further affects the stability of the TIM23 complex. The role of bilayer-forming phospholipids like phosphatidylcholine (PC) in protein transport into the inner membrane and the matrix is unknown. Here, we report that import of presequence-containing precursors and carrier proteins is impaired in PC-deficient mitochondria. Surprisingly, depletion of PC does not affect stability and activity of respiratory supercomplexes, and the membrane potential is maintained. Instead, the dynamic TIM23 complex is destabilized when the PC levels are reduced, whereas the TIM22 complex remains intact. Our analysis further revealed that initial precursor binding to the TIM23 complex is impaired in PC-deficient mitochondria. We conclude that reduced PC levels differentially affect the TIM22 and TIM23 complexes in mitochondrial protein transport. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Actions of parathyroid hormone related peptide in mouse parietal endoderm formation

    NARCIS (Netherlands)

    Veltmaat, J.M.

    2001-01-01

    Summary Since about a decade, several reports have strongly suggested a role for parathyroid hormone related peptide (PTHrP) in the formation of parietal endoderm (PE) in the mouse embryo. This thesis is aimed first at elucidating the biological significance of parathyroid hormone related peptide

  19. Rapid formation of plasma protein corona critically affects nanoparticle pathophysiology

    Science.gov (United States)

    Tenzer, Stefan; Docter, Dominic; Kuharev, Jörg; Musyanovych, Anna; Fetz, Verena; Hecht, Rouven; Schlenk, Florian; Fischer, Dagmar; Kiouptsi, Klytaimnistra; Reinhardt, Christoph; Landfester, Katharina; Schild, Hansjörg; Maskos, Michael; Knauer, Shirley K.; Stauber, Roland H.

    2013-10-01

    In biological fluids, proteins bind to the surface of nanoparticles to form a coating known as the protein corona, which can critically affect the interaction of the nanoparticles with living systems. As physiological systems are highly dynamic, it is important to obtain a time-resolved knowledge of protein-corona formation, development and biological relevancy. Here we show that label-free snapshot proteomics can be used to obtain quantitative time-resolved profiles of human plasma coronas formed on silica and polystyrene nanoparticles of various size and surface functionalization. Complex time- and nanoparticle-specific coronas, which comprise almost 300 different proteins, were found to form rapidly (<0.5 minutes) and, over time, to change significantly in terms of the amount of bound protein, but not in composition. Rapid corona formation is found to affect haemolysis, thrombocyte activation, nanoparticle uptake and endothelial cell death at an early exposure time.

  20. Prediction of disease-related mutations affecting protein localization

    Directory of Open Access Journals (Sweden)

    Laurila Kirsti

    2009-03-01

    Full Text Available Abstract Background Eukaryotic cells contain numerous compartments, which have different protein constituents. Proteins are typically directed to compartments by short peptide sequences that act as targeting signals. Translocation to the proper compartment allows a protein to form the necessary interactions with its partners and take part in biological networks such as signalling and metabolic pathways. If a protein is not transported to the correct intracellular compartment either the reaction performed or information carried by the protein does not reach the proper site, causing either inactivation of central reactions or misregulation of signalling cascades, or the mislocalized active protein has harmful effects by acting in the wrong place. Results Numerous methods have been developed to predict protein subcellular localization with quite high accuracy. We applied bioinformatics methods to investigate the effects of known disease-related mutations on protein targeting and localization by analyzing over 22,000 missense mutations in more than 1,500 proteins with two complementary prediction approaches. Several hundred putative localization affecting mutations were identified and investigated statistically. Conclusion Although alterations to localization signals are rare, these effects should be taken into account when analyzing the consequences of disease-related mutations.

  1. Digestion and microbial protein synthesis in sheep as affected by ...

    African Journals Online (AJOL)

    Useni , Alain

    sajas.v42i5.9. Digestion and microbial protein synthesis in sheep as affected by exogenous fibrolytic enzymes. W.F.J. van de Vyver. #. & B.A. Useni. Department of Animal Sciences, Stellenbosch University, Private Bag X1, Matieland 7602, South ...

  2. The Arabidopsis NIMIN proteins affect NPR1 differentially

    Directory of Open Access Journals (Sweden)

    Meike eHermann

    2013-04-01

    Full Text Available NON-EXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1 is the central regulator of the pathogen defense reaction systemic acquired resistance (SAR. NPR1 acts by sensing the SAR signal molecule salicylic acid (SA to induce expression of PATHOGENESIS-RELATED (PR genes. Mechanistically, NPR1 is the core of a transcription complex interacting with TGA transcription factors and NIM1 INTERACTING (NIMIN proteins. Arabidopsis NIMIN1 has been shown to suppress NPR1 activity in transgenic plants. The Arabidopsis NIMIN family comprises four structurally related, yet distinct members. Here, we show that NIMIN1, NIMIN2 and NIMIN3 are expressed differentially, and that the encoded proteins affect expression of the SAR marker PR-1 differentially. NIMIN3 is expressed constitutively at a low level, but NIMIN2 and NIMIN1 are both responsive to SA. While NIMIN2 is an immediate early SA-induced and NPR1-independent gene, NIMIN1 is activated after NIMIN2, but clearly before PR-1. Notably, NIMIN1, like PR-1, depends on NPR1. In a transient assay system, NIMIN3 suppresses SA-induced PR-1 expression, albeit to a lesser extent than NIMIN1, whereas NIMIN2 does not negatively affect PR-1 gene activation. Furthermore, although binding to the same domain in the C-terminus, NIMIN1 and NIMIN2 interact differentially with NPR1, thus providing a molecular basis for their opposing effects on NPR1. Together, our data suggest that the Arabidopsis NIMIN proteins are regulators of the SAR response. We propose that NIMINs act in a strictly consecutive and SA-regulated manner on the SA sensor protein NPR1, enabling NPR1 to monitor progressing threat by pathogens and to promote appropriate defense gene activation at distinct stages of SAR. In this scenario, the defense gene PR-1 is repressed at the onset of SAR by SA-induced, yet instable NIMIN1.

  3. Hormone-Related Migraine Headaches and Mood Disorders: Treatment with Estrogen Stabilization.

    Science.gov (United States)

    Warnock, Julia K; Cohen, Lawrence J; Blumenthal, Harvey; Hammond, Jordan E

    2017-01-01

    Because estrogens and the trigeminal system are inherently linked, prescribers who are treating a woman with a hormonally related mood disorder and migraine headaches should consider hormonal options to optimize the patient's treatment. This article discusses the interrelationships of estrogen, serotonin, and the trigeminal system as they relate to menstrual migraine occurrence and hormone-related mood symptoms. In addition, clinical examples are provided to facilitate the prescribers treating women during reproductive transitions in which declining estrogens are related to their suffering. © 2016 Pharmacotherapy Publications, Inc.

  4. Parathyroid Hormone-Related Peptide Elicits Peripheral TRPV1-dependent Mechanical Hypersensitivity

    Science.gov (United States)

    Shepherd, Andrew J.; Mickle, Aaron D.; Kadunganattil, Suraj; Hu, Hongzhen; Mohapatra, Durga P.

    2018-01-01

    Bone metastasis in breast, prostate and lung cancers often leads to chronic pain, which is poorly managed by existing analgesics. The neurobiological mechanisms that underlie chronic pain associated with bone-metastasized cancers are not well understood, but sensitization of peripheral nociceptors by tumor microenvironment factors has been demonstrated to be important. Parathyroid hormone-related peptide (PTHrP) is highly expressed in bone-metastasized breast and prostate cancers, and is critical to growth and proliferation of these tumors in the bone tumor microenvironment. Previous studies have suggested that PTHrP could sensitize nociceptive sensory neurons, resulting in peripheral pain hypersensitivity. In this study, we found that PTHrP induces both heat and mechanical hypersensitivity, that are dependent on the pain-transducing transient receptor potential channel family vanilloid, member-1 (TRPV1), but not the mechano-transducing TRPV4 and TRPA1 ion channels. Functional ratiometric Ca2+ imaging and voltage-clamp electrophysiological analysis of cultured mouse DRG neurons show significant potentiation of TRPV1, but not TRPA1 or TRPV4 channel activation by PTHrP. Interestingly, PTHrP exposure led to the slow and sustained activation of TRPV1, in the absence of any exogenous channel agonist, and is dependent on the expression of the type-1 parathyroid hormone receptor (PTH1), as well as on downstream phosphorylation of the channel by protein kinase C (PKC). Accordingly, local administration of specific small-molecule antagonists of TRPV1 to mouse hindpaws after the development of PTHrP-induced mechanical hypersensitivity led to its significant attenuation. Collectively, our findings suggest that PTHrP/PTH1-mediated flow activation of TRPV1 channel contributes at least in part to the development and maintenance of peripheral mechanical pain hypersensitivity, and could therefore constitute a mechanism for nociceptor sensitization in the context of metastatic bone

  5. Plant protein and animal proteins: do they differentially affect cardiovascular disease risk?

    Science.gov (United States)

    Richter, Chesney K; Skulas-Ray, Ann C; Champagne, Catherine M; Kris-Etherton, Penny M

    2015-11-01

    Proteins from plant-based compared with animal-based food sources may have different effects on cardiovascular disease (CVD) risk factors. Numerous epidemiologic and intervention studies have evaluated their respective health benefits; however, it is difficult to isolate the role of plant or animal protein on CVD risk. This review evaluates the current evidence from observational and intervention studies, focusing on the specific protein-providing foods and populations studied. Dietary protein is derived from many food sources, and each provides a different composite of nonprotein compounds that can also affect CVD risk factors. Increasing the consumption of protein-rich foods also typically results in lower intakes of other nutrients, which may simultaneously influence outcomes. Given these complexities, blanket statements about plant or animal protein may be too general, and greater consideration of the specific protein food sources and the background diet is required. The potential mechanisms responsible for any specific effects of plant and animal protein are similarly multifaceted and include the amino acid content of particular foods, contributions from other nonprotein compounds provided concomitantly by the whole food, and interactions with the gut microbiome. Evidence to date is inconclusive, and additional studies are needed to further advance our understanding of the complexity of plant protein vs. animal protein comparisons. Nonetheless, current evidence supports the idea that CVD risk can be reduced by a dietary pattern that provides more plant sources of protein compared with the typical American diet and also includes animal-based protein foods that are unprocessed and low in saturated fat. © 2015 American Society for Nutrition.

  6. Plant Protein and Animal Proteins: Do They Differentially Affect Cardiovascular Disease Risk?12

    Science.gov (United States)

    Richter, Chesney K; Skulas-Ray, Ann C; Champagne, Catherine M; Kris-Etherton, Penny M

    2015-01-01

    Proteins from plant-based compared with animal-based food sources may have different effects on cardiovascular disease (CVD) risk factors. Numerous epidemiologic and intervention studies have evaluated their respective health benefits; however, it is difficult to isolate the role of plant or animal protein on CVD risk. This review evaluates the current evidence from observational and intervention studies, focusing on the specific protein-providing foods and populations studied. Dietary protein is derived from many food sources, and each provides a different composite of nonprotein compounds that can also affect CVD risk factors. Increasing the consumption of protein-rich foods also typically results in lower intakes of other nutrients, which may simultaneously influence outcomes. Given these complexities, blanket statements about plant or animal protein may be too general, and greater consideration of the specific protein food sources and the background diet is required. The potential mechanisms responsible for any specific effects of plant and animal protein are similarly multifaceted and include the amino acid content of particular foods, contributions from other nonprotein compounds provided concomitantly by the whole food, and interactions with the gut microbiome. Evidence to date is inconclusive, and additional studies are needed to further advance our understanding of the complexity of plant protein vs. animal protein comparisons. Nonetheless, current evidence supports the idea that CVD risk can be reduced by a dietary pattern that provides more plant sources of protein compared with the typical American diet and also includes animal-based protein foods that are unprocessed and low in saturated fat. PMID:26567196

  7. Glycolipid transfer protein expression is affected by glycosphingolipid synthesis.

    Directory of Open Access Journals (Sweden)

    Matti A Kjellberg

    Full Text Available Members of the glycolipid transfer protein superfamily (GLTP are found from animals and fungi to plants and red micro-alga. Eukaryotes that encode the glucosylceramide synthase responsible for the synthesis of glucosylceramide, the precursor for most glycosphingolipids, also produce GLTPs. Cells that does not synthesize glucosylceramide neither express GLTPs. Based on this genetic relationship there must be a strong correlation between the synthesis of glucosylceramide and GLTPs. To regulate the levels of glycolipids we have used inhibitors of intracellular trafficking, glycosphingolipid synthesis and degradation, and small interfering RNA to down-regulate the activity of glucosylceramide synthase activity. We found that GLTP expression, both at the mRNA and protein levels, is elevated in cells that accumulate glucosylceramide. Monensin and brefeldin A block intracellular vesicular transport mechanisms. Brefeldin A treatment leads to accumulation of newly synthesized glucosylceramide, galactosylceramide and lactosylceramide in a fused endoplasmic reticulum-Golgi complex. On the other hand, inhibiting glycosphingolipid degradation with conduritol-B-epoxide, that generates glucosylceramide accumulation in the lysosomes, did not affect the levels of GLTP. However, glycosphingolipid synthesis inhibitors like PDMP, NB-DNJ and myriocin, all decreased glucosylceramide and GLTP below normal levels. We also found that an 80% loss of glucosylceramide due to glucosylceramide synthase knockdown resulted in a significant reduction in the expression of GLTP. We show here that interfering with membrane trafficking events and simple neutral glycosphingolipid synthesis will affect the expression of GLTP. We postulate that a change in the glucosylceramide balance causes a response in the GLTP expression, and put forward that GLTP might play a role in lipid directing and sensing of glucosylceramide at the ER-Golgi interface.

  8. A protein aggregation based test for screening of the agents affecting thermostability of proteins.

    Directory of Open Access Journals (Sweden)

    Tatyana Eronina

    Full Text Available To search for agents affecting thermal stability of proteins, a test based on the registration of protein aggregation in the regime of heating with a constant rate was used. The initial parts of the dependences of the light scattering intensity (I on temperature (T were analyzed using the following empiric equation: I = K(agg(T-T(0(2, where K(agg is the parameter characterizing the initial rate of aggregation and T(0 is a temperature at which the initial increase in the light scattering intensity is registered. The aggregation data are interpreted in the frame of the model assuming the formation of the start aggregates at the initial stages of the aggregation process. Parameter T(0 corresponds to the moment of the origination of the start aggregates. The applicability of the proposed approach was demonstrated on the examples of thermal aggregation of glycogen phosphorylase b from rabbit skeletal muscles and bovine liver glutamate dehydrogenase studied in the presence of agents of different chemical nature. The elaborated approach to the study of protein aggregation may be used for rapid identification of small molecules that interact with protein targets.

  9. Role of Parathyroid Hormone-Related Protein Signaling in Chronic Pancreatitis

    OpenAIRE

    Miriam Falzon; Vandanajay Bhatia

    2015-01-01

    Chronic pancreatitis (CP), a progressive inflammatory disease where acini are destroyed and replaced by fibrous tissue, increases the risk for pancreatic cancer. Risk factors include alcohol, smoking, and obesity. The effects of these risk factors are exacerbated in patients with mutations in genes that predispose to CP. The different environmental and genetic factors produce the same clinical phenotype; once CP develops, disease course is the same regardless of etiology. Critical questions s...

  10. Protein and lipid oxidation affect the viscoelasticity of whey protein layers at the oil-water interface

    NARCIS (Netherlands)

    Berton-Carabin, Claire C.; Schroder, Anja; Rovalino-Cordova, Ana; Schroën, Karin; Sagis, Leonard

    2016-01-01

    Protein and lipid oxidation are prevailing issues that negatively affect the nutritional and sensory quality of food emulsions. It is probable that such oxidative modifications affect the functional properties of proteins, and in particular their ability to form densely packed, interconnected

  11. Conformational fluctuations affect protein alignment in dilute liquid crystal media

    NARCIS (Netherlands)

    Louhivuori, M; Otten, R; Lindorff-Larsen, K; Annila, A

    2006-01-01

    The discovery of dilute liquid crystalline media to align biological macromolecules has opened many new possibilities to study protein and nucleic acid structures by NMR spectroscopy. We inspect the basic alignment phenomenon for an ensemble of protein conformations to deduce relative contributions

  12. Conformational fluctuations affect protein alignment in dilute liquid crystal media

    DEFF Research Database (Denmark)

    Louhivuori, M.; Otten, R.; Lindorff-Larsen, Kresten

    2006-01-01

    The discovery of dilute liquid crystalline media to align biological macromolecules has opened many new possibilities to study protein and nucleic acid structures by NMR spectroscopy. We inspect the basic alignment phenomenon for an ensemble of protein conformations to deduce relative contributions...

  13. Lipid Bilayer Composition Affects Transmembrane Protein Orientation and Function

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    Katie D. Hickey

    2011-01-01

    Full Text Available Sperm membranes change in structure and composition upon ejaculation to undergo capacitation, a molecular transformation which enables spermatozoa to undergo the acrosome reaction and be capable of fertilization. Changes to the membrane environment including lipid composition, specifically lipid microdomains, may be responsible for enabling capacitation. To study the effect of lipid environment on proteins, liposomes were created using lipids extracted from bull sperm membranes, with or without a protein (Na+ K+-ATPase or -amylase. Protein incorporation, function, and orientation were determined. Fluorescence resonance energy transfer (FRET confirmed protein inclusion in the lipid bilayer, and protein function was confirmed using a colourometric assay of phosphate production from ATP cleavage. In the native lipid liposomes, ATPase was oriented with the subunit facing the outer leaflet, while changing the lipid composition to 50% native lipids and 50% exogenous lipids significantly altered this orientation of Na+ K+-ATPase within the membranes.

  14. A novel family of small proteins that affect plant development

    Energy Technology Data Exchange (ETDEWEB)

    John Charles Walker

    2011-04-29

    The DVL genes represent a new group of plant proteins that influence plant growth and development. Overexpression of DVL1, and other members of the DVL family, causes striking phenotypic changes. The DVL proteins share sequence homology in their C-terminal half. Point mutations in the C-terminal domain show it is necessary and deletion studies demonstrate the C-terminal domain is sufficient to confer the overexpression phenotypes. The phenotypes observed, and the conservation of the protein sequence in the plant kingdom, does suggest the DVL proteins have a role in modulating plant growth and development. Our working hypothesis is the DVL proteins function as regulators of cellular signaling pathways that control growth and development.

  15. The HIV Tat protein affects processing of ribosomal RNA precursor

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    Bellenchi Gian

    2008-06-01

    Full Text Available Abstract Background Inside the cell, the HIV Tat protein is mainly found in the nucleus and nucleolus. The nucleolus, the site of ribosome biogenesis, is a highly organized, non-membrane-bound sub-compartment where proteins with a high affinity for nucleolar components are found. While it is well known that Tat accumulates in the nucleolus via a specific nucleolar targeting sequence, its function in this compartment it still unknown. Results To clarify the significance of the Tat nucleolar localization, we induced the expression of the protein during oogenesis in Drosophila melanogaster strain transgenic for HIV-tat gene. Here we show that Tat localizes in the nucleoli of Drosophila oocyte nurse cells, where it specifically co-localizes with fibrillarin. Tat expression is accompanied by a significant decrease of cytoplasmic ribosomes, which is apparently related to an impairment of ribosomal rRNA precursor processing. Such an event is accounted for by the interaction of Tat with fibrillarin and U3 snoRNA, which are both required for pre-rRNA maturation. Conclusion Our data contribute to understanding the function of Tat in the nucleolus, where ribosomal RNA synthesis and cell cycle control take place. The impairment of nucleolar pre-rRNA maturation through the interaction of Tat with fibrillarin-U3snoRNA complex suggests a process by which the virus modulates host response, thus contributing to apoptosis and protein shut-off in HIV-uninfected cells.

  16. REEPs Are Membrane Shaping Adapter Proteins That Modulate Specific G Protein-Coupled Receptor Trafficking by Affecting ER Cargo Capacity

    OpenAIRE

    Susann Björk; Hurt, Carl M.; Ho, Vincent K.; Timothy Angelotti

    2013-01-01

    Receptor expression enhancing proteins (REEPs) were identified by their ability to enhance cell surface expression of a subset of G protein-coupled receptors (GPCRs), specifically GPCRs that have proven difficult to express in heterologous cell systems. Further analysis revealed that they belong to the Yip (Ypt-interacting protein) family and that some REEP subtypes affect ER structure. Yip family comparisons have established other potential roles for REEPs, including regulation of ER-Golgi t...

  17. Does oxidation affect the water functionality of myofibrillar proteins?

    Science.gov (United States)

    Bertram, Hanne Christine; Kristensen, Mette; Østdal, Henrik; Baron, Caroline P; Young, Jette F; Andersen, Henrik Jørgen

    2007-03-21

    Water-binding properties of myofibrils extracted from porcine muscle, and added hemoglobin with and without exposure to H2O2, were characterized using low-field proton NMR T2 relaxometry. The effects of pH and ionic strength in the samples were investigated as pH was adjusted to 5.4, 6.2, and 7.0 and ionic strength was adjusted to 0.29, 0.46, and 0.71 M, respectively. The formation of dityrosine as a measure of oxidative protein cross-linking revealed a significant increase in dityrosine concentrations upon H2O2 activation. The formation of dityrosine was strongly pH-dependent and increased with decreasing pH. In addition, increased levels of thiobarbituric acid reactive substances were observed upon addition of H2O2, implying that lipid oxidation was enhanced, however, with a different oxidation pattern as compared to the myofibrillar proteins. Low-field NMR relaxation measurements revealed reduced T2 relaxation times upon H2O2 activation, which corresponds to reduced water-holding capacity upon oxidation. However, a direct relationship between degree of oxidation and T2 relaxation time was not observed with various pH values and ionic strengths, and further studies are needed for a complete understanding of the effect of oxidation on myofibrillar functionality.

  18. Prion protein polymorphisms affect chronic wasting disease progression.

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    Chad J Johnson

    Full Text Available Analysis of the PRNP gene in cervids naturally infected with chronic wasting disease (CWD suggested that PRNP polymorphisms affect the susceptibility of deer to infection. To test this effect, we orally inoculated 12 white-tailed deer with CWD agent. Three different PRNP alleles, wild-type (wt; glutamine at amino acid 95 and glycine at 96, Q95H (glutamine to histidine at amino acid position 95 and G96S (glycine to serine at position 96 were represented in the study cohort with 5 wt/wt, 3 wt/G96S, and 1 each wt/Q95H and Q95H/G96S. Two animals were lost to follow-up due to intercurrent disease. The inoculum was prepared from Wisconsin hunter-harvested homozygous wt/wt animals. All infected deer presented with clinical signs of CWD; the orally infected wt/wt had an average survival period of 693 days post inoculation (dpi and G96S/wt deer had an average survival period of 956 dpi. The Q95H/wt and Q95H/G96S deer succumbed to CWD at 1,508 and 1,596 dpi respectively. These data show that polymorphisms in the PRNP gene affect CWD incubation period. Deer heterozygous for the PRNP alleles had extended incubation periods with the Q95H allele having the greatest effect.

  19. Protein leverage affects energy intake of high-protein diets in humans.

    Science.gov (United States)

    Martens, Eveline A; Lemmens, Sofie G; Westerterp-Plantenga, Margriet S

    2013-01-01

    The protein leverage hypothesis requires specific evidence that protein intake is regulated more strongly than energy intake. The objective was to determine ad libitum energy intake, body weight changes, and appetite profile in response to protein-to-carbohydrate + fat ratio over 12 consecutive days and in relation to age, sex, BMI, and type of protein. A 12-d randomized crossover study was performed in 40 men and 39 women [mean ± SD age: 34.0 ± 17.6 y; BMI (in kg/m(2)): 23.7 ± 3.4] with the use of diets containing 5%, 15%, and 30% of energy from protein from a milk or plant source. Protein-content effects did not differ by age, sex, BMI, or type of protein. Total energy intake was significantly lower in the high-protein (7.21 ± 3.08 MJ/d) condition than in the low-protein (9.33 ± 3.52 MJ/d) and normal-protein (9.62 ± 3.51 MJ/d) conditions (P = 0.001), which was predominantly the result of a lower energy intake from meals (P = 0.001). Protein intake varied directly according to the amount of protein in the diet (P = 0.001). The AUC of visual analog scale appetite ratings did not differ significantly, yet fluctuations in hunger (P = 0.019) and desire to eat (P = 0.026) over the day were attenuated in the high-protein condition compared with the normal-protein condition. We found evidence to support the protein leverage hypothesis in that individuals underate relative to energy balance from diets containing a higher protein-to-carbohydrate + fat ratio. No evidence for protein leverage effects from diets containing a lower ratio of protein to carbohydrate + fat was obtained. It remains to be shown whether a relatively low protein intake would cause overeating or would be the effect of overeating of carbohydrate and fat. The study was registered at clinicaltrials.gov as NCT01320189.

  20. Protein source and choice of anticoagulant decisively affect nanoparticle protein corona and cellular uptake

    Science.gov (United States)

    Schöttler, S.; Klein, Katja; Landfester, K.; Mailänder, V.

    2016-03-01

    Protein adsorption on nanoparticles has been a focus of the field of nanocarrier research in the past few years and more and more papers are dealing with increasingly detailed lists of proteins adsorbed to a plethora of nanocarriers. While there is an urgent need to understand the influence of this protein corona on nanocarriers' interactions with cells the strong impact of the protein source on corona formation and the consequence for interaction with different cell types are factors that are regularly neglected, but should be taken into account for a meaningful analysis. In this study, the importance of the choice of protein source used for in vitro protein corona analysis is concisely investigated. Major and decisive differences in cellular uptake of a polystyrene nanoparticle incubated in fetal bovine serum, human serum, human citrate and heparin plasma are reported. Furthermore, the protein compositions are determined for coronas formed in the respective incubation media. A strong influence of heparin, which is used as an anticoagulant for plasma generation, on cell interaction is demonstrated. While heparin enhances the uptake into macrophages, it prevents internalization into HeLa cells. Taken together we can give the recommendation that human plasma anticoagulated with citrate seems to give the most relevant results for in vitro studies of nanoparticle uptake.Protein adsorption on nanoparticles has been a focus of the field of nanocarrier research in the past few years and more and more papers are dealing with increasingly detailed lists of proteins adsorbed to a plethora of nanocarriers. While there is an urgent need to understand the influence of this protein corona on nanocarriers' interactions with cells the strong impact of the protein source on corona formation and the consequence for interaction with different cell types are factors that are regularly neglected, but should be taken into account for a meaningful analysis. In this study, the importance

  1. Intrinsically Disordered Segments Affect Protein Half-Life in the Cell and during Evolution

    Directory of Open Access Journals (Sweden)

    Robin van der Lee

    2014-09-01

    Full Text Available Precise control of protein turnover is essential for cellular homeostasis. The ubiquitin-proteasome system is well established as a major regulator of protein degradation, but an understanding of how inherent structural features influence the lifetimes of proteins is lacking. We report that yeast, mouse, and human proteins with terminal or internal intrinsically disordered segments have significantly shorter half-lives than proteins without these features. The lengths of the disordered segments that affect protein half-life are compatible with the structure of the proteasome. Divergence in terminal and internal disordered segments in yeast proteins originating from gene duplication leads to significantly altered half-life. Many paralogs that are affected by such changes participate in signaling, where altered protein half-life will directly impact cellular processes and function. Thus, natural variation in the length and position of disordered segments may affect protein half-life and could serve as an underappreciated source of genetic variation with important phenotypic consequences.

  2. Protein quality affects bone status during moderate protein restriction in growing mice.

    Science.gov (United States)

    Rouy, Emilien; Vico, Laurence; Laroche, Norbert; Benoit, Valérie; Rousseau, Brigitte; Blachier, François; Tomé, Daniel; Blais, Anne

    2014-02-01

    Adequate protein intake during development is critical to ensure optimal bone gain and to attain a higher peak bone mass later on. We hypothesized that the quality of the dietary protein is of prime importance for bone physiology during moderate protein restriction. The target population was growing Balb/C mice. We compared two protein restricted diets (6% of total energy as protein), one based on soy (LP-SOY) and one based on casein (LP-CAS). For comparison, a normal protein soy-based control group (NP-SOY) and a low protein group receiving an anabolic daily parathyroid hormone (PTH) 1-34 injection (LP-SOY+PTH) were included in the protocol. After 8weeks, LP-SOY mice had reduced body weights related to a lower lean mass whereas LP-CAS mice were not different from the NP-SOY group. LP-SOY mice were characterized by lower femoral cortical thickness, bone volume, trabecular number and thickness and increased medullar adiposity when compared to both the LP-CAS and NP-SOY groups. However, the dietary intervention had no effect on the vertebral parameters. The negative effect of the LP-SOY diet was correlated to an impaired bone formation as shown by the reduced P1NP serum level as well as the reduced osteoid surfaces and bone formation rate in the femur. PTH injection in LP-SOY mice had no effect on total weight or lean mass, but improved all bone parameters at both femoral and vertebral sites, suggesting that amino acid deficiency was not the primary reason for degraded bone status in mice consuming soy protein. In conclusion, our study showed that under the same protein restriction (6% of energy), a soy diet leads to impaired bone health whereas a casein diet has little effect when compared to a normal protein control. © 2013.

  3. Interactions affected by arginine methylation in the yeast protein-protein interaction network.

    Science.gov (United States)

    Erce, Melissa A; Abeygunawardena, Dhanushi; Low, Jason K K; Hart-Smith, Gene; Wilkins, Marc R

    2013-11-01

    Protein-protein interactions can be modulated by the methylation of arginine residues. As a means of testing this, we recently described a conditional two-hybrid system, based on the bacterial adenylate cyclase (BACTH) system. Here, we have used this conditional two-hybrid system to explore the effect of arginine methylation in modulating protein-protein interactions in a subset of the Saccharomyces cerevisiae arginine methylproteome network. Interactions between the yeast hub protein Npl3 and yeast proteins Air2, Ded1, Gbp2, Snp1, and Yra1 were first validated in the absence of methylation. The major yeast arginine methyltransferase Hmt1 was subsequently included in the conditional two-hybrid assay, initially to determine the degree of methylation that occurs. Proteins Snp1 and Yra1 were confirmed as Hmt1 substrates, with five and two novel arginine methylation sites mapped by ETD LC-MS/MS on these proteins, respectively. Proteins Ded1 and Gbp2, previously predicted but not confirmed as substrates of Hmt1, were also found to be methylated with five and seven sites mapped respectively. Air2 was found to be a novel substrate of Hmt1 with two sites mapped. Finally, we investigated the interactions of Npl3 with the five interaction partners in the presence of active Hmt1 and in the presence of Hmt1 with a G68R inactivation mutation. We found that the interaction between Npl3 and Air2, and Npl3 and Ded1, were significantly increased in the presence of active Hmt1; the interaction of Npl3 and Snp1 showed a similar degree of increase in interaction but this was not statistically significant. The interactions of Npl3 and Gbp2, along with Npl3 and Yra1, were not significantly increased or decreased by methylation. We conclude that methylarginine may be a widespread means by which the interactions of proteins are modulated.

  4. Parathyroid hormone-related peptide is a downstream target for ras and src activation.

    Science.gov (United States)

    Li, X; Drucker, D J

    1994-03-04

    Neoplastic transformation may be associated with the development of paraneoplastic syndromes that complicate the management of patients with malignant disease. One of these syndromes, humoral hypercalcemia of malignancy, has been shown to be attributable to increased secretion of a parathyroid hormone-related peptide (PTHrP). The gene encoding PTHrP is expressed at low levels in a growth factor-dependent manner in normal tissues, and the mechanisms underlying selective tumor-associated induction of PTHrP gene expression remain unclear. Here we show that cellular transformation by the EJ-Ha-ras and v-src oncogenes is associated with a marked increase in PTHrP gene expression. Ha-ras- and v-src-transfected NRK 49F cells and ras-transfected RCB 2.2 cell lines secreted increased amounts of immunoreactive PTHrP. Northern blot analysis also demonstrated increased levels of PTHrP mRNA transcripts in oncogene-transfected stable cell lines. No significant change in the half-life of PTHrP mRNA transcripts was detected in Ha-ras- and v-src-transformed cells. In contrast, nuclear run-on assays demonstrated an increased rate of PTHrP gene transcription in oncogene-transformed cells. These observations demonstrate that the gene encoding PTHrP is a downstream target for ras and src, potentially providing a molecular basis for understanding the activation of PTHrP gene expression in malignancy-associated hypercalcemia.

  5. Assessment of Multifactor Gene-Environment Interactions and Ovarian Cancer Risk: Candidate Genes, Obesity, and Hormone-Related Risk Factors

    NARCIS (Netherlands)

    Usset, J.L.; Raghavan, R.; Tyrer, J.P.; McGuire, V.; Sieh, W.; Webb, P.; Chang-Claude, J.; Rudolph, A.; Anton-Culver, H.; Berchuck, A.; Brinton, L.; Cunningham, J.M.; Defazio, A.; Doherty, J.A.; Edwards, R.P.; Gayther, S.A.; Gentry-Maharaj, A.; Goodman, M.T.; Hogdall, E.; Jensen, A.; Johnatty, S.E.; Kiemeney, L.A.L.M.; Kjaer, S.K.; Larson, M.C.; Lurie, G.; Massuger, L.F.; Menon, U.; Modugno, F.; Moysich, K.B.; Ness, R.B.; Pike, M.C.; Ramus, S.J.; Rossing, M.A.; Rothstein, J.; Song, H.; Thompson, P.J.; Berg, D.J. Van Den; Vierkant, R.A.; Wang-Gohrke, S.; Wentzensen, N.; Whittemore, A.S.; Wilkens, L.R.; Wu, A.H.; Yang, H.; Pearce, C.L.; Schildkraut, J.M.; Pharoah, P.; Goode, E.L.; Fridley, B.L.

    2016-01-01

    BACKGROUND: Many epithelial ovarian cancer (EOC) risk factors relate to hormone exposure and elevated estrogen levels are associated with obesity in postmenopausal women. Therefore, we hypothesized that gene-environment interactions related to hormone-related risk factors could differ between obese

  6. Efficacy and safety of prolonged-release lanreotide in patients with gastrointestinal neuroendocrine tumors and hormone-related symptoms

    NARCIS (Netherlands)

    Wymenga, ANM; Eriksson, B; Salmela, PI; Jacobsen, MB; Van Cutsem, EJDG; Fiasse, RH; Valimaki, MJ; Renstrup, J; de Vries, EGE; Oberg, KE

    Purpose: To evaluate the prolonged release (PR) of the long-acting somatostatin analog lanreotide in patients with gastrointestinal neuroendocrine tumors and its effect on hormone-related symptomatology, tumor markers, tumor size, tolerability, and quality of life (QOL), Patients and Methods:

  7. Feeding modality affects muscle protein deposition by influencing protein synthesis, but not degradation in muscle of neonatal pigs

    Science.gov (United States)

    Neonatal pigs can serve as dual-use models for nutrition research in animal agriculture and biomedical fields. To determine how feeding modality by either intermittent bolus or continuous schedule affects protein anabolism and catabolism, neonatal pigs (n = 6/group, 9-d-old) were overnight fasted (F...

  8. Arginine depletion by arginine deiminase does not affect whole protein metabolism or muscle fractional protein synthesis rate in mice.

    Directory of Open Access Journals (Sweden)

    Juan C Marini

    Full Text Available Due to the absolute need for arginine that certain cancer cells have, arginine depletion is a therapy in clinical trials to treat several types of cancers. Arginine is an amino acids utilized not only as a precursor for other important molecules, but also for protein synthesis. Because arginine depletion can potentially exacerbate the progressive loss of body weight, and especially lean body mass, in cancer patients we determined the effect of arginine depletion by pegylated arginine deiminase (ADI-PEG 20 on whole body protein synthesis and fractional protein synthesis rate in multiple tissues of mice. ADI-PEG 20 successfully depleted circulating arginine (<1 μmol/L, and increased citrulline concentration more than tenfold. Body weight and body composition, however, were not affected by ADI-PEG 20. Despite the depletion of arginine, whole body protein synthesis and breakdown were maintained in the ADI-PEG 20 treated mice. The fractional protein synthesis rate of muscle was also not affected by arginine depletion. Most tissues (liver, kidney, spleen, heart, lungs, stomach, small and large intestine, pancreas were able to maintain their fractional protein synthesis rate; however, the fractional protein synthesis rate of brain, thymus and testicles was reduced due to the ADI-PEG 20 treatment. Furthermore, these results were confirmed by the incorporation of ureido [14C]citrulline, which indicate the local conversion into arginine, into protein. In conclusion, the intracellular recycling pathway of citrulline is able to provide enough arginine to maintain protein synthesis rate and prevent the loss of lean body mass and body weight.

  9. Mild hypothermic culture conditions affect residual host cell protein composition post-Protein A chromatography.

    Science.gov (United States)

    Goey, Cher Hui; Bell, David; Kontoravdi, Cleo

    2018-01-30

    Host cell proteins (HCPs) are endogenous impurities, and their proteolytic and binding properties can compromise the integrity, and, hence, the stability and efficacy of recombinant therapeutic proteins such as monoclonal antibodies (mAbs). Nonetheless, purification of mAbs currently presents a challenge because they often co-elute with certain HCP species during the capture step of protein A affinity chromatography. A Quality-by-Design (QbD) strategy to overcome this challenge involves identifying residual HCPs and tracing their source to the harvested cell culture fluid (HCCF) and the corresponding cell culture operating parameters. Then, problematic HCPs in HCCF may be reduced by cell engineering or culture process optimization. Here, we present experimental results linking cell culture temperature and post-protein A residual HCP profile. We had previously reported that Chinese hamster ovary cell cultures conducted at standard physiological temperature and with a shift to mild hypothermia on day 5 produced HCCF of comparable product titer and HCP concentration, but with considerably different HCP composition. In this study, we show that differences in HCP variety at harvest cascaded to downstream purification where different residual HCPs were present in the two sets of samples post-protein A purification. To detect low-abundant residual HCPs, we designed a looping liquid chromatography-mass spectrometry method with continuous expansion of a preferred, exclude, and targeted peptide list. Mild hypothermic cultures produced 20% more residual HCP species, especially cell membrane proteins, distinct from the control. Critically, we identified that half of the potentially immunogenic residual HCP species were different between the two sets of samples.

  10. Common soil of smoking-associated and hormone-related cancers: estrogen deficiency

    Directory of Open Access Journals (Sweden)

    Zsuzsanna Suba

    2011-12-01

    Full Text Available Accumulation of non-smoker, non-drinker elderly postmenopausal female patients among smokingassociated oral cancer cases raised the plausible idea: estrogen deficiency maybe a cancer risk factor. On the other hand, the extremely rare cases of young women with oral cancer regularly exhibited hormonal disorders, such as irregular menstrual cycles and infertility. Furthermore, in the history of middle-aged female, oral cancer cases a primary ovarian failure or complete hysterectomy was a conspicuously frequent finding suggestive of an estrogen deficient milieu. There were many striking contradictions concerning the associations of female sexual steroids and cancer risk as well. Until now, breast and endometrial cancers were regarded as typically estrogen-induced tumors, particularly in postmenopausal cases. However, unexplained beneficial anti-cancer effects of hormonereplacement therapy were reported against cancers at several sites, even tumors of the highly hormone-responsive organs. Re-evaluation of results of the experimental and epidemiological studies, which endeavored to justify the carcinogenic capacity of estrogen, exhibited many shortcomings and controversies. The new findings both on smoking associated and on hormone related cancers added up to the same conversion; not estrogen but rather its deficiency might provoke cancer initiation. Thorough review of the literary data justified that the exquisite regulatory capacity of estrogen and its surveillance on growth, development, differentiation, and metabolism are indispensable, whereas an estrogen-deficient milieu may induce a breakdown in gene-regulation. Recognition of the anticancer capacity of estrogen may provide new insights into the etiology of malignancies and leads to new strategies for cancer prevention and cure.

  11. Protein turnover in lactating mink (Mustela vison) is not affected by dietary protein supply

    DEFF Research Database (Denmark)

    Tauson, Anne-Helene; Fink, Rikke; Chwalibog, André

    2006-01-01

    The mink is a strict carnivore and may therefore serve as a model for the cat. Current recommendations for protein supply for lactating mink are based on production experiments with preweaning kit growth as a measure of dietary adequacy (1,2). Recently, nitrogen balance and substrate oxidation ha...

  12. Glucagon-Like Peptide 2 Stimulates Postresection Intestinal Adaptation in Preterm Pigs by Affecting Proteins Related to Protein, Carbohydrate, and Sulphur Metabolism

    DEFF Research Database (Denmark)

    Jiang, Pingping; Vegge, Andreas; Thymann, Thomas

    2017-01-01

    BACKGROUND: Exogenous glucagon-like peptide 2 (GLP-2) stimulates intestinal adaptation after resection in animal models of pediatric short bowel syndrome (SBS). It is unknown whether the molecular mechanisms of such GLP-2 effects are similar to those of postresection spontaneous adaptation. Using...... cellular structural proteins, while the added GLP-2 treatment affected proteins involved in protein processing and the metabolism of protein, carbohydrate, and sulphur. CONCLUSION: In the first days following resection, proteins affected by resection plus GLP-2 treatment differed markedly from those...... affected by the spontaneous intestinal adaptation following resection alone. Whether more long-term GLP-2 treatment may affect the intestinal proteome following intestinal resection remains unknown....

  13. Effects of abaloparatide, a human parathyroid hormone-related peptide analog, on bone mineral density in postmenopausal women with osteoporosis.

    Science.gov (United States)

    Leder, Benjamin Z; O'Dea, Louis St L; Zanchetta, José R; Kumar, Prasana; Banks, Kathleen; McKay, Kathleen; Lyttle, C Richard; Hattersley, Gary

    2015-02-01

    Abaloparatide is a novel synthetic peptide analog of parathyroid hormone-related protein (PTHrP) that is currently being developed as a potential anabolic agent in the treatment of postmenopausal osteoporosis. This study sought to assess the effects of abaloparatide on bone mineral density (BMD) at the lumbar spine, total hip, and femoral neck in postmenopausal women with osteoporosis. Multi-center, multi-national, double-blind placebo controlled trial in which postmenopausal women were randomly assigned to receive 24 weeks of treatment with daily sc injections of placebo, abaloparatide, 20, 40, or 80 μg, or teriparatide, 20 μg. A 24-week extension was also performed in a subset of subjects. Postmenopausal women with osteoporosis (n = 222). BMD by dual-x-ray absorptiometry and biochemical markers of bone turnover. At 24 weeks, lumbar spine BMD increased by 2.9, 5.2, and 6.7% in the abaloparatide, 20-, 40-, and 80-μg groups, respectively, and 5.5% in the teriparatide group. The increases in the 40- and 80-μg abaloparatide groups and the teriparatide group were significantly greater than placebo (1.6%). Femoral neck BMD increased by 2.7, 2.2, and 3.1% in abaloparatide, 20-, 40-, and 80-μg groups, respectively, and 1.1% in the teriparatide group. The increase in femoral neck BMD with abaloparatide, 80 μg was significantly greater than placebo (0.8%). Total hip BMD increased by 1.4, 2.0, and 2.6% in the abaloparatide, 20-, 40-, and 80-μg groups, respectively. The total hip increases in the 40- and 80-μg abaloparatide groups were greater than both placebo (0.4%) and teriparatide (0.5%). Compared with placebo, 24 weeks of daily sc abaloparatide increases BMD of the lumbar spine, femoral neck, and total hip in a dose-dependent fashion. Moreover, the abaloparatide-induced BMD increases at the total hip are greater than with the marketed dose of teriparatide. These results support the further investigation of abaloparatide as an anabolic therapy in postmenopausal

  14. REEPs are membrane shaping adapter proteins that modulate specific g protein-coupled receptor trafficking by affecting ER cargo capacity.

    Directory of Open Access Journals (Sweden)

    Susann Björk

    Full Text Available Receptor expression enhancing proteins (REEPs were identified by their ability to enhance cell surface expression of a subset of G protein-coupled receptors (GPCRs, specifically GPCRs that have proven difficult to express in heterologous cell systems. Further analysis revealed that they belong to the Yip (Ypt-interacting protein family and that some REEP subtypes affect ER structure. Yip family comparisons have established other potential roles for REEPs, including regulation of ER-Golgi transport and processing/neuronal localization of cargo proteins. However, these other potential REEP functions and the mechanism by which they selectively enhance GPCR cell surface expression have not been clarified. By utilizing several REEP family members (REEP1, REEP2, and REEP6 and model GPCRs (α2A and α2C adrenergic receptors, we examined REEP regulation of GPCR plasma membrane expression, intracellular processing, and trafficking. Using a combination of immunolocalization and biochemical methods, we demonstrated that this REEP subset is localized primarily to ER, but not plasma membranes. Single cell analysis demonstrated that these REEPs do not specifically enhance surface expression of all GPCRs, but affect ER cargo capacity of specific GPCRs and thus their surface expression. REEP co-expression with α2 adrenergic receptors (ARs revealed that this REEP subset interacts with and alter glycosidic processing of α2C, but not α2A ARs, demonstrating selective interaction with cargo proteins. Specifically, these REEPs enhanced expression of and interacted with minimally/non-glycosylated forms of α2C ARs. Most importantly, expression of a mutant REEP1 allele (hereditary spastic paraplegia SPG31 lacking the carboxyl terminus led to loss of this interaction. Thus specific REEP isoforms have additional intracellular functions besides altering ER structure, such as enhancing ER cargo capacity, regulating ER-Golgi processing, and interacting with select cargo

  15. REEPs are membrane shaping adapter proteins that modulate specific g protein-coupled receptor trafficking by affecting ER cargo capacity.

    Science.gov (United States)

    Björk, Susann; Hurt, Carl M; Ho, Vincent K; Angelotti, Timothy

    2013-01-01

    Receptor expression enhancing proteins (REEPs) were identified by their ability to enhance cell surface expression of a subset of G protein-coupled receptors (GPCRs), specifically GPCRs that have proven difficult to express in heterologous cell systems. Further analysis revealed that they belong to the Yip (Ypt-interacting protein) family and that some REEP subtypes affect ER structure. Yip family comparisons have established other potential roles for REEPs, including regulation of ER-Golgi transport and processing/neuronal localization of cargo proteins. However, these other potential REEP functions and the mechanism by which they selectively enhance GPCR cell surface expression have not been clarified. By utilizing several REEP family members (REEP1, REEP2, and REEP6) and model GPCRs (α2A and α2C adrenergic receptors), we examined REEP regulation of GPCR plasma membrane expression, intracellular processing, and trafficking. Using a combination of immunolocalization and biochemical methods, we demonstrated that this REEP subset is localized primarily to ER, but not plasma membranes. Single cell analysis demonstrated that these REEPs do not specifically enhance surface expression of all GPCRs, but affect ER cargo capacity of specific GPCRs and thus their surface expression. REEP co-expression with α2 adrenergic receptors (ARs) revealed that this REEP subset interacts with and alter glycosidic processing of α2C, but not α2A ARs, demonstrating selective interaction with cargo proteins. Specifically, these REEPs enhanced expression of and interacted with minimally/non-glycosylated forms of α2C ARs. Most importantly, expression of a mutant REEP1 allele (hereditary spastic paraplegia SPG31) lacking the carboxyl terminus led to loss of this interaction. Thus specific REEP isoforms have additional intracellular functions besides altering ER structure, such as enhancing ER cargo capacity, regulating ER-Golgi processing, and interacting with select cargo proteins

  16. Timing and pattern of postexercise protein ingestion affects whole-body protein balance in healthy children: a randomized trial.

    Science.gov (United States)

    Volterman, Kimberly A; Moore, Daniel R; Breithaupt, Peter; Grathwohl, Dominik; Offord, Elizabeth A; Karagounis, Leonidas G; Timmons, Brian W

    2017-11-01

    The dose and timing of postexercise protein ingestion can influence whole-body protein balance (WBPB) in adults, although comparable data from children are scarce. This study investigated how protein intake (both amount and distribution) postexercise can affect WBPB in physically active children. Thirty-five children (26 males; 9-13 years old) underwent a 5-day adaptation diet, maintaining a protein intake of 0.95 g·kg-1·day-1. Participants consumed [15N]glycine (2 mg·kg-1) before performing 3 × 20 min of variable-intensity cycling, and whole-body protein kinetics were assessed over 6 and 24 h of recovery. Fifteen grams of protein was distributed across 2 isoenergetic carbohydrate-containing beverages (15 and 240 min postexercise) containing reciprocal amounts of protein (i.e., 0 + 15 g, 5 + 10 g, 10 + 5 g, and 15 + 0 g for Groups A-D, respectively). Over the 6 h that included the exercise bout and consumption of the first beverage at 15 min postexercise, WBPB (i.e., synthesis - breakdown) demonstrated a linear increase of 0.647 g·kg-1·day-1 per 1 g protein intake (P < 0.001). Over 24 h, robust regression revealed that WBPB was best modeled by a parabola (P < 0.05), suggesting that a maximum in WBPB was achieved between groups B and C. In conclusion, despite a dose response early in recovery, a periodized protein intake with multiple smaller doses after physical activity may be more beneficial than a single bolus dose in promoting daily WBPB in healthy active children.

  17. LDL receptor-related protein 5 (LRP5) affects bone accrual and eye development

    NARCIS (Netherlands)

    Gong, Y.; Slee, R. B.; Fukai, N.; Rawadi, G.; Roman-Roman, S.; Reginato, A. M.; Wang, H.; Cundy, T.; Glorieux, F. H.; Lev, D.; Zacharin, M.; Oexle, K.; Marcelino, J.; Suwairi, W.; Heeger, S.; Sabatakos, G.; Apte, S.; Adkins, W. N.; Allgrove, J.; Arslan-Kirchner, M.; Batch, J. A.; Beighton, P.; Black, G. C.; Boles, R. G.; Boon, L. M.; Borrone, C.; Brunner, H. G.; Carle, G. F.; Dallapiccola, B.; de Paepe, A.; Floege, B.; Halfhide, M. L.; Hall, B.; Hennekam, R. C.; Hirose, T.; Jans, A.; Jüppner, H.; Kim, C. A.; Keppler-Noreuil, K.; Kohlschuetter, A.; Lacombe, D.; Lambert, M.; Lemyre, E.; Letteboer, T.; Peltonen, L.; Ramesar, R. S.; Romanengo, M.; Somer, H.; Steichen-Gersdorf, E.; Steinmann, B.; Sullivan, B.; Superti-Furga, A.; Swoboda, W.; van den Boogaard, M. J.; van Hul, W.; Vikkula, M.; Votruba, M.; Zabel, B.; Garcia, T.; Baron, R.; Olsen, B. R.; Warman, M. L.

    2001-01-01

    In humans, low peak bone mass is a significant risk factor for osteoporosis. We report that LRP5, encoding the low-density lipoprotein receptor-related protein 5, affects bone mass accrual during growth. Mutations in LRP5 cause the autosomal recessive disorder osteoporosis-pseudoglioma syndrome

  18. LDL receptor-related protein 5 (LRP5) affects bone accrual and eye development.

    NARCIS (Netherlands)

    Gong, Y.; Slee, R.B.; Fukai, N.; Rawadi, G.; Roman-Roman, S.; Reginato, A.M.; Wang, H.; Cundy, T.; Glorieux, F.H.; Lev, D.; Zacharin, M.; Oexle, K.; Marcelino, J.; Suwairi, W.; Heeger, S.; Sabatakos, G.; Apte, S.; Adkins, W.; Allgrove, J.; Arslan-Kirchner, M.; Batch, J.A.; Beighton, P.; Black, G.C.M.; Boles, R.G.; Boon, L.; Borrone, C.; Brunner, H.G.; Carle, G.F.; Dallapicola, B.; Paepe, A. de; Floege, B.; Halfhide, M.L.; Hall, B.D.; Hennekam, R.C.M.; Hirose, T.; Jans, A.; Juppner, H.; Kim, C.; Keppler-Noreuil, K.; Kohlschuetter, A.; Lacombe, D.; Lambert, M.; Lemyre, E.; Letteboer, T.; Peltonen, L.; Ramesar, R.S.; Romanengo, M.; Somer, H.; Steichen-Gersdorf, E.; Steinmann, B.; Sullivan, B.; Superti-Furga, A.; Swoboda, W.; Boogaard, M.J. van den; Hul, W. van; Vikkula, M.; Votruba, M.; Zabel, B.; Garcia, T.; Baron, R.; Olsen, B.R.; Warman, M.L.

    2001-01-01

    In humans, low peak bone mass is a significant risk factor for osteoporosis. We report that LRP5, encoding the low-density lipoprotein receptor-related protein 5, affects bone mass accrual during growth. Mutations in LRP5 cause the autosomal recessive disorder osteoporosis-pseudoglioma syndrome

  19. Dietary Protein Affects Gene Expression and Prevents Lipid Accumulation in the Liver in Mice

    NARCIS (Netherlands)

    Schwarz, J.; Tome, D.G.; Baars, A.; Hooiveld, G.J.E.J.; Müller, M.R.

    2012-01-01

    Background and Aims: High protein (HP) diets are suggested to positively modulate obesity and associated increased prevalence of non-alcoholic fatty liver (NAFLD) disease in humans and rodents. The aim of our study was to detect mechanisms by which a HP diet affects hepatic lipid accumulation.

  20. Alternative splicing affects the targeting sequence of peroxisome proteins in Arabidopsis.

    Science.gov (United States)

    An, Chuanjing; Gao, Yuefang; Li, Jinyu; Liu, Xiaomin; Gao, Fuli; Gao, Hongbo

    2017-07-01

    A systematic analysis of the Arabidopsis genome in combination with localization experiments indicates that alternative splicing affects the peroxisomal targeting sequence of at least 71 genes in Arabidopsis. Peroxisomes are ubiquitous eukaryotic cellular organelles that play a key role in diverse metabolic functions. All peroxisome proteins are encoded by nuclear genes and target to peroxisomes mainly through two types of targeting signals: peroxisomal targeting signal type 1 (PTS1) and PTS2. Alternative splicing (AS) is a process occurring in all eukaryotes by which a single pre-mRNA can generate multiple mRNA variants, often encoding proteins with functional differences. However, the effects of AS on the PTS1 or PTS2 and the targeting of the protein were rarely studied, especially in plants. Here, we systematically analyzed the genome of Arabidopsis, and found that the C-terminal targeting sequence PTS1 of 66 genes and the N-terminal targeting sequence PTS2 of 5 genes are affected by AS. Experimental determination of the targeting of selected protein isoforms further demonstrated that AS at both the 5' and 3' region of a gene can affect the inclusion of PTS2 and PTS1, respectively. This work underscores the importance of AS on the global regulation of peroxisome protein targeting.

  1. How does fluorescent labeling affect the binding kinetics of proteins with intact cells?

    Science.gov (United States)

    Yin, Linliang; Wang, Wei; Wang, Shaopeng; Zhang, Fenni; Zhang, Shengtao; Tao, Nongjian

    2016-01-01

    Fluorescent labeling is a mainstream technology for detecting molecular binding. Despite the importance, few studies have been devoted to quantitatively examine the effect of labeling on the molecular binding processes. Here we present a quantitative study on the binding kinetics of fluorescent-labeled and un-labeled molecules (lectin proteins) with glycoproteins on the membrane of cells using surface plasmon resonance imaging (SPRi) technique. The study shows that fluorescent labeling has a significant influence on the binding behaviors of proteins, especially the association processes, and the influence depends sensitively on the charge of fluorescent labels. It further shows that the labels also affect the local distribution of probe proteins, due to the inhomogeneous surface charge distribution of the cell membrane. Our work indicates that fluorescent labeling in general affects the binding behaviors, but proper design of the label will help to minimize its effect. PMID:25486538

  2. The Regulatory Protein RosR Affects Rhizobium leguminosarum bv. trifolii Protein Profiles, Cell Surface Properties, and Symbiosis with Clover.

    Science.gov (United States)

    Rachwał, Kamila; Boguszewska, Aleksandra; Kopcińska, Joanna; Karaś, Magdalena; Tchórzewski, Marek; Janczarek, Monika

    2016-01-01

    Rhizobium leguminosarum bv. trifolii is capable of establishing a symbiotic relationship with plants from the genus Trifolium. Previously, a regulatory protein encoded by rosR was identified and characterized in this bacterium. RosR possesses a Cys2-His2-type zinc finger motif and belongs to Ros/MucR family of rhizobial transcriptional regulators. Transcriptome profiling of the rosR mutant revealed a role of this protein in several cellular processes, including the synthesis of cell-surface components and polysaccharides, motility, and bacterial metabolism. Here, we show that a mutation in rosR resulted in considerable changes in R. leguminosarum bv. trifolii protein profiles. Extracellular, membrane, and periplasmic protein profiles of R. leguminosarum bv. trifolii wild type and the rosR mutant were examined, and proteins with substantially different abundances between these strains were identified. Compared with the wild type, extracellular fraction of the rosR mutant contained greater amounts of several proteins, including Ca(2+)-binding cadherin-like proteins, a RTX-like protein, autoaggregation protein RapA1, and flagellins FlaA and FlaB. In contrast, several proteins involved in the uptake of various substrates were less abundant in the mutant strain (DppA, BraC, and SfuA). In addition, differences were observed in membrane proteins of the mutant and wild-type strains, which mainly concerned various transport system components. Using atomic force microscopy (AFM) imaging, we characterized the topography and surface properties of the rosR mutant and wild-type cells. We found that the mutation in rosR gene also affected surface properties of R. leguminosarum bv. trifolii. The mutant cells were significantly more hydrophobic than the wild-type cells, and their outer membrane was three times more permeable to the hydrophobic dye N-phenyl-1-naphthylamine. The mutation of rosR also caused defects in bacterial symbiotic interaction with clover plants. Compared with

  3. The regulatory protein RosR affects Rhizobium leguminosarum bv. trifolii protein profiles, cell surface properties, and symbiosis with clover

    Directory of Open Access Journals (Sweden)

    Kamila Rachwał

    2016-08-01

    Full Text Available Rhizobium leguminosarum bv. trifolii is capable of establishing a symbiotic relationship with plants from the genus Trifolium. Previously, a regulatory protein encoded by rosR was identified and characterized in this bacterium. RosR possesses a Cys2-His2-type zinc finger motif and belongs to Ros/MucR family of rhizobial transcriptional regulators. Transcriptome profiling of the rosR mutant revealed a role of this protein in several cellular processes, including the synthesis of cell-surface components and polysaccharides, motility, and bacterial metabolism. Here, we show that a mutation in rosR resulted in considerable changes in R. leguminosarum bv. trifolii protein profiles. Extracellular, membrane, and periplasmic protein profiles of R. leguminosarum bv. trifolii wild type and the rosR mutant were examined, and proteins with substantially different abundances between these strains were identified. Compared with the wild type, extracellular fraction of the rosR mutant contained greater amounts of several proteins, including Ca2+-binding cadherin-like proteins, a RTX-like protein, autoaggregation protein RapA1, and flagellins FlaA and FlaB. In contrast, several proteins involved in the uptake of various substrates were less abundant in the mutant strain (DppA, BraC, and SfuA. In addition, differences were observed in membrane proteins of the mutant and wild-type strains, which mainly concerned various transport system components. Using atomic force microscopy imaging, we characterized the topography and surface properties of the rosR mutant and wild-type cells. We found that the mutation in rosR gene also affected surface properties of R. leguminosarum bv. trifolii. The mutant cells were significantly more hydrophobic than the wild-type cells, and their outer membrane was three times more permeable to the hydrophobic dye N-phenyl-1-naphthylamine. The mutation of rosR also caused defects in bacterial symbiotic interaction with clover plants

  4. Contraction intensity and feeding affect collagen and myofibrillar protein synthesis rates differently in human skeletal muscle

    DEFF Research Database (Denmark)

    Holm, Lars; van Hall, Gerrit; Rose, Adam

    2010-01-01

    with the fasting condition. The Rp-s6k-4E-binding protein-1 (BP1) and the mitogen-activated protein kinase (MAPk) pathways were activated by the HL intensity and were suggested to be responsible for regulating myofibrillar FSR in response to adequate contractile activity. Feeding predominantly affected Rp-s6k...... to contractile activity, whereas elongation mainly was found to respond to feeding. Furthermore, although functionally linked, the contractile and the supportive matrix structures upregulate their protein synthesis rate quite differently in response to feeding and contractile activity and intensity.......Exercise stimulates muscle protein fractional synthesis rate (FSR), but the importance of contractile intensity and whether it interplays with feeding is not understood. This was investigated following two distinct resistance exercise (RE) contraction intensities using an intrasubject design...

  5. Contaminant loading in remote Arctic lakes affects cellular stress-related proteins expression in feral charr.

    Science.gov (United States)

    Wiseman, Steve; Jorgensen, Even H.; Maule, Alec G.; Vijayan, Mathilakath M.

    2011-01-01

    The remote Arctic lakes on Bjornoya Island, Norway, offer a unique opportunity to study possible affect of lifelong contaminant exposure in wild populations of landlocked Arctic charr (Salvelinus alpinus). This is because Lake Ellasjoen has persistent organic pollutant (POP) levels that are significantly greater than in the nearby Lake Oyangen. We examined whether this differential contaminant loading was reflected in the expression of protein markers of exposure and effect in the native fish. We assessed the expressions of cellular stress markers, including cytochrome P4501A (Cyp1A), heat shock protein 70 (hsp70), and glucocorticoid receptor (GR) in feral charr from the two lakes. The average polychlorinated biphenyl (PCB) load in the charr liver from Ellasjoen was approximately 25-fold higher than in individuals from Oyangen. Liver Cyp1A protein expression was significantly higher in individuals from Ellasjoen compared with Oyangen, confirming differential PCB exposure. There was no significant difference in hsp70 protein expression in charr liver between the two lakes. However, brain hsp70 protein expression was significantly elevated in charr from Ellasjoen compared with Oyangen. Also, liver GR protein expression was significantly higher in the Ellasjoen charr compared with Oyangen charr. Taken together, our results suggest changes to cellular stress-related protein expression as a possible adaptation to chronic-contaminant exposure in feral charr in the Norwegian high-Arctic.

  6. Mycobacterium tuberculosis Complex Exhibits Lineage-Specific Variations Affecting Protein Ductility and Epitope Recognition

    Science.gov (United States)

    Yruela, Inmaculada; Contreras-Moreira, Bruno; Magalhães, Carlos; Osório, Nuno S.

    2016-01-01

    Abstract The advent of whole-genome sequencing has provided an unprecedented detail about the evolution and genetic significance of species-specific variations across the whole Mycobacterium tuberculosis Complex. However, little attention has been focused on understanding the functional roles of these variations in the protein coding sequences. In this work, we compare the coding sequences from 74 sequenced mycobacterial species including M. africanum, M. bovis, M. canettii, M. caprae, M. orygis, and M. tuberculosis. Results show that albeit protein variations affect all functional classes, those proteins involved in lipid and intermediary metabolism and respiration have accumulated mutations during evolution. To understand the impact of these mutations on protein functionality, we explored their implications on protein ductility/disorder, a yet unexplored feature of mycobacterial proteomes. In agreement with previous studies, we found that a Gly71Ile substitution in the PhoPR virulence system severely affects the ductility of its nearby region in M. africanum and animal-adapted species. In the same line of evidence, the SmtB transcriptional regulator shows amino acid variations specific to the Beijing lineage, which affects the flexibility of the N-terminal trans-activation domain. Furthermore, despite the fact that MTBC epitopes are evolutionary hyperconserved, we identify strain- and lineage-specific amino acid mutations affecting previously known T-cell epitopes such as EsxH and FbpA (Ag85A). Interestingly, in silico studies reveal that these variations result in differential interaction of epitopes with the main HLA haplogroups. PMID:28062754

  7. Protein corona composition of gold nanoparticles/nanorods affects amyloid beta fibrillation process

    Science.gov (United States)

    Mirsadeghi, Somayeh; Dinarvand, Rassoul; Ghahremani, Mohammad Hossein; Hormozi-Nezhad, Mohammad Reza; Mahmoudi, Zohreh; Hajipour, Mohammad Javad; Atyabi, Fatemeh; Ghavami, Mahdi; Mahmoudi, Morteza

    2015-03-01

    Protein fibrillation process (e.g., from amyloid beta (Aβ) and α-synuclein) is the main cause of several catastrophic neurodegenerative diseases such as Alzheimer's and Parkinson diseases. During the past few decades, nanoparticles (NPs) were recognized as one of the most promising tools for inhibiting the progress of the disease by controlling the fibrillation kinetic process; for instance, gold NPs have a strong capability to inhibit Aβ fibrillations. It is now well understood that a layer of biomolecules would cover the surface of NPs (so called ``protein corona'') upon the interaction of NPs with protein sources. Due to the fact that the biological species (e.g., cells and amyloidal proteins) ``see'' the protein corona coated NPs rather than the pristine coated particles, one should monitor the fibrillation process of amyloidal proteins in the presence of corona coated NPs (and not pristine coated ones). Therefore, the previously obtained data on NPs effects on the fibrillation process should be modified to achieve a more reliable and predictable in vivo results. Herein, we probed the effects of various gold NPs (with different sizes and shapes) on the fibrillation process of Aβ in the presence and absence of protein sources (i.e., serum and plasma). We found that the protein corona formed a shell at the surface of gold NPs, regardless of their size and shape, reducing the access of Aβ to the gold inhibitory surface and, therefore, affecting the rate of Aβ fibril formation. More specifically, the anti-fibrillation potencies of various corona coated gold NPs were strongly dependent on the protein source and their concentrations (10% serum/plasma (simulation of an in vitro milieu) and 100% serum/plasma (simulation of an in vivo milieu)).Protein fibrillation process (e.g., from amyloid beta (Aβ) and α-synuclein) is the main cause of several catastrophic neurodegenerative diseases such as Alzheimer's and Parkinson diseases. During the past few decades

  8. Single Molecule Force Spectroscopy Reveals that Electrostatic Interactions Affect the Mechanical Stability of Proteins

    Science.gov (United States)

    Zheng, Peng; Cao, Yi; Bu, Tianjia; Straus, Suzana K.; Li, Hongbin

    2011-01-01

    It is well known that electrostatic interactions play important roles in determining the thermodynamic stability of proteins. However, the investigation into the role of electrostatic interactions in mechanical unfolding of proteins has just begun. Here we used single molecule atomic force microscopy techniques to directly evaluate the effect of electrostatic interactions on the mechanical stability of a small protein GB1. We engineered a bi-histidine motif into the force-bearing region of GB1. By varying the pH, histidine residues can switch between protonated and deprotonated states, leading to the change of the electrostatic interactions between the two histidine residues. We found that the mechanical unfolding force of the engineered protein decreased by ∼34% (from 115 pN to 76 pN) on changing the pH from 8.5 to 3, due to the increased electrostatic repulsion between the two positively charged histidines at acidic pH. Our results demonstrated that electrostatic interactions can significantly affect the mechanical stability of elastomeric proteins, and modulating the electrostatic interactions of key charged residues can become a promising method for regulating the mechanical stability of elastomeric proteins. PMID:21402036

  9. Assessment of Multifactor Gene-Environment Interactions and Ovarian Cancer Risk: Candidate Genes, Obesity, and Hormone-Related Risk Factors.

    Science.gov (United States)

    Usset, Joseph L; Raghavan, Rama; Tyrer, Jonathan P; McGuire, Valerie; Sieh, Weiva; Webb, Penelope; Chang-Claude, Jenny; Rudolph, Anja; Anton-Culver, Hoda; Berchuck, Andrew; Brinton, Louise; Cunningham, Julie M; DeFazio, Anna; Doherty, Jennifer A; Edwards, Robert P; Gayther, Simon A; Gentry-Maharaj, Aleksandra; Goodman, Marc T; Høgdall, Estrid; Jensen, Allan; Johnatty, Sharon E; Kiemeney, Lambertus A; Kjaer, Susanne K; Larson, Melissa C; Lurie, Galina; Massuger, Leon; Menon, Usha; Modugno, Francesmary; Moysich, Kirsten B; Ness, Roberta B; Pike, Malcolm C; Ramus, Susan J; Rossing, Mary Anne; Rothstein, Joseph; Song, Honglin; Thompson, Pamela J; van den Berg, David J; Vierkant, Robert A; Wang-Gohrke, Shan; Wentzensen, Nicolas; Whittemore, Alice S; Wilkens, Lynne R; Wu, Anna H; Yang, Hannah; Pearce, Celeste Leigh; Schildkraut, Joellen M; Pharoah, Paul; Goode, Ellen L; Fridley, Brooke L

    2016-05-01

    Many epithelial ovarian cancer (EOC) risk factors relate to hormone exposure and elevated estrogen levels are associated with obesity in postmenopausal women. Therefore, we hypothesized that gene-environment interactions related to hormone-related risk factors could differ between obese and non-obese women. We considered interactions between 11,441 SNPs within 80 candidate genes related to hormone biosynthesis and metabolism and insulin-like growth factors with six hormone-related factors (oral contraceptive use, parity, endometriosis, tubal ligation, hormone replacement therapy, and estrogen use) and assessed whether these interactions differed between obese and non-obese women. Interactions were assessed using logistic regression models and data from 14 case-control studies (6,247 cases; 10,379 controls). Histotype-specific analyses were also completed. SNPs in the following candidate genes showed notable interaction: IGF1R (rs41497346, estrogen plus progesterone hormone therapy, histology = all, P = 4.9 × 10(-6)) and ESR1 (rs12661437, endometriosis, histology = all, P = 1.5 × 10(-5)). The most notable obesity-gene-hormone risk factor interaction was within INSR (rs113759408, parity, histology = endometrioid, P = 8.8 × 10(-6)). We have demonstrated the feasibility of assessing multifactor interactions in large genetic epidemiology studies. Follow-up studies are necessary to assess the robustness of our findings for ESR1, CYP11A1, IGF1R, CYP11B1, INSR, and IGFBP2 Future work is needed to develop powerful statistical methods able to detect these complex interactions. Assessment of multifactor interaction is feasible, and, here, suggests that the relationship between genetic variants within candidate genes and hormone-related risk factors may vary EOC susceptibility. Cancer Epidemiol Biomarkers Prev; 25(5); 780-90. ©2016 AACR. ©2016 American Association for Cancer Research.

  10. Transcriptional profile analysis of E3 ligase and hormone-related genes expressed during wheat grain development

    Directory of Open Access Journals (Sweden)

    Capron Delphine

    2012-03-01

    Full Text Available Abstract Background Wheat grains are an important source of food, stock feed and raw materials for industry, but current production levels cannot meet world needs. Elucidation of the molecular mechanisms underlying wheat grain development will contribute valuable information to improving wheat cultivation. One of the most important mechanisms implicated in plant developmental processes is the ubiquitin-proteasome system (UPS. Among the different roles of the UPS, it is clear that it is essential to hormone signaling. In particular, E3 ubiquitin ligases of the UPS have been shown to play critical roles in hormone perception and signal transduction. Results A NimbleGen microarray containing 39,179 UniGenes was used to study the kinetics of gene expression during wheat grain development from the early stages of cell division to the mid-grain filling stage. By comparing 11 consecutive time-points, 9284 differentially expressed genes were identified and annotated during this study. A comparison of the temporal profiles of these genes revealed dynamic transcript accumulation profiles with major reprogramming events that occurred during the time intervals of 80-120 and 220-240°Cdays. The list of the genes expressed differentially during these transitions were identified and annotated. Emphasis was placed on E3 ligase and hormone-related genes. In total, 173 E3 ligase coding genes and 126 hormone-related genes were differentially expressed during the cell division and grain filling stages, with each family displaying a different expression profile. Conclusions The differential expression of genes involved in the UPS and plant hormone pathways suggests that phytohormones and UPS crosstalk might play a critical role in the wheat grain developmental process. Some E3 ligase and hormone-related genes seem to be up- or down-regulated during the early and late stages of the grain development.

  11. Protein denaturation and water-protein interactions as affected by low temperature long time treatment of porcine longissimus dorsi.

    Science.gov (United States)

    Christensen, Line; Bertram, Hanne C; Aaslyng, Margit D; Christensen, Mette

    2011-08-01

    The relationship between water-protein interactions and heat-induced protein denaturation in low temperature long time (LTLT) treated pork Longissimus dorsi was investigated by combining low-field NMR T₂ relaxometry with DSC measurements and measures of shrinkage of porcine Longissimus dorsi heated to 53 °C, 55 °C, 57 °C and 59 °C for either 3 or 20 h. Water within the myofibrils, measured by NMR T₂₁ relaxation times, was affected by both temperature and holding time during LTLT treatment between 53 °C and 59 °C. The changes in NMR T₂₁ relaxation times were associated with decreased fiber diameter and increased cooking loss, revealing a relationship between transverse shrinkage, water-protein interactions and cooking loss. DSC measurements revealed a concomitant decrease in ΔH(68 °C), which suggests impact of collagen denaturation on the retention of water within the meat during LTLT treatment. Furthermore, a decrease in ΔH(75 °C) suggested that prolonged cooking (20 h) resulted in actin denaturation leading to decreased T₂₁ relaxation times and higher cooking loss. Copyright © 2011 Elsevier Ltd. All rights reserved.

  12. Hemoglobin S and C affect protein export in Plasmodium falciparum-infected erythrocytes

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    Nicole Kilian

    2015-02-01

    Full Text Available Malaria is a potentially deadly disease. However, not every infected person develops severe symptoms. Some people are protected by naturally occurring mechanisms that frequently involve inheritable modifications in their hemoglobin. The best studied protective hemoglobins are the sickle cell hemoglobin (HbS and hemoglobin C (HbC which both result from a single amino acid substitution in β-globin: glutamic acid at position 6 is replaced by valine or lysine, respectively. How these hemoglobinopathies protect from severe malaria is only partly understood. Models currently proposed in the literature include reduced disease-mediating cytoadherence of parasitized hemoglobinopathic erythrocytes, impaired intraerythrocytic development of the parasite, dampened inflammatory responses, or a combination thereof. Using a conditional protein export system and tightly synchronized Plasmodium falciparum cultures, we now show that export of parasite-encoded proteins across the parasitophorous vacuolar membrane is delayed, slower, and reduced in amount in hemoglobinopathic erythrocytes as compared to parasitized wild type red blood cells. Impaired protein export affects proteins targeted to the host cell cytoplasm, Maurer's clefts, and the host cell plasma membrane. Impaired protein export into the host cell compartment provides a mechanistic explanation for the reduced cytoadherence phenotype associated with parasitized hemoglobinopathic erythrocytes.

  13. Signaling proteins that influence energy intake may affect unintentional weight loss in elderly persons.

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    Wernette, Catherine M; White, B Douglas; Zizza, Claire A

    2011-06-01

    After age 70 to 75 years, average body weight decreases both in ailing and healthy people because of a loss of appetite that results in reduced energy intake and the loss of body fat and lean muscle tissue. This so-called anorexia of aging predisposes elderly people to continued pathologic weight loss and malnutrition-major causes of morbidity and mortality. Health care professionals must understand the many factors involved in the anorexia of aging to help older adults prevent unintentional weight loss. Psychological, social, and cultural factors are important effectors; however, physiological factors are emphasized here because they are not thoroughly understood and they make it inherently difficult for most people to alter their body weight. Monoamines, steroid hormones (glucocorticoids and mineralocorticoids), endocannabinoids, and proteins all influence body weight. This review is an analysis of proteins from the brain, pancreas, adipose tissue, and gastrointestinal tract that are known to affect energy intake and energy balance, with an attempt to identify those factors that may change with aging. The articles included in this review were obtained by a PubMed database search using the keywords mouse OR rat OR human AND aged OR aging OR older OR elderly AND adult AND anorexia OR "unintentional weight loss," and each of the individual proteins discussed, as well as from the reference lists of those articles. The results reveal that some proteins may be important in the development of unintentional weight loss in elderly persons, whereas others may not have a significant role. However, many of the proteins that could conceivably have a role in unintentional weight loss have not yet been studied with that question in mind. Preventing unintentional weight loss in older adults is an important goal and further research on the role of proteins important for the maintenance of energy balance and the development of unintentional weight loss in elderly persons is

  14. Dietary lipid and gross energy affect protein utilization in the rare minnow Gobiocypris rarus

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    Wu, Benli; Xiong, Xiaoqin; Xie, Shouqi; Wang, Jianwei

    2016-07-01

    An 8-week feeding trial was conducted to detect the optimal dietary protein and energy, as well as the effects of protein to energy ratio on growth, for the rare minnow ( Gobiocypris rarus), which are critical to nutrition standardization for model fish. Twenty-four diets were formulated to contain three gross energy (10, 12.5, 15 kJ/g), four protein (20%, 25%, 30%, 35%), and two lipid levels (3%, 6%). The results showed that optimal dietary E/P was 41.7-50 kJ/g for maximum growth in juvenile rare minnows at 6% dietary crude lipid. At 3% dietary lipid, specific growth rate (SGR) increased markedly when E/P decreased from 62.5 kJ/g to 35.7 kJ/g and gross energy was 12.5 kJ/g, and from 75 kJ/g to 42.9 kJ/g when gross energy was 15.0 kJ/g. The optimal gross energy was estimated at 12.5 kJ/g and excess energy decreased food intake and growth. Dietary lipid exhibited an apparent protein-sparing effect. Optimal protein decreased from 35% to 25%-30% with an increase in dietary lipid from 3% to 6% without adversely effecting growth. Dietary lipid level affects the optimal dietary E/P ratio. In conclusion, recommended dietary protein and energy for rare minnow are 20%-35% and 10-12.5 kJ/g, respectively.

  15. Evaluation of acute phase proteins in cows affected by Traumatic Reticuloperitonitis

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    Ali Rezaei

    2008-11-01

    Full Text Available Traumatic Reticuloperitonitis is one of the most important diseases of the bovine digestive system which if diagnosed in early stage and before severe adhesions develop, more effective treatments leading to better prognosis could be under taken. In the course of accurate and timely diagnosis                            of these diseases, acute phase proteins of the affected animals were evaluated. For this purpose,                       blood samples were taken form jugular vein of 25 Holstein cows in which the presence of disease         clinically and confirmed with laparatomy and their sera separated. Blood sample were also collected from 20 healthy cows as the control group. After preparing serum sample, total protein haptoglobin, serum amyloid A and ceruloplasmin were measured in both groups. Total protein was determined by the Biuret method, haptoglobin and serum amyloid A by ELISA and ceruloplasmin through determination of the oxidative activity. Statistical analysis indicated significant difference in the amounts of total protein, haptoglobin and serum amyloid A between the two groups (p

  16. Deoxygenation Affects Composition of Membrane-Bound Proteins in Human Erythrocytes

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    Oksana G. Luneva

    2016-06-01

    Full Text Available Background/Aims: ATP release from erythrocyte plays a key role in hypoxia-induced elevation of blood flow in systematic circulation. We have previously shown that hemolysis contributes to erythrocyte ATP release triggered by several stimuli, including hypoxia, but the molecular mechanisms of hypoxia-increased membrane fragility remain unknown. Methods: In this study, we compared the action of hypoxia on hemolysis, ATP release and the composition of membrane-bound proteins in human erythrocytes. Results: Twenty minutes incubation of human erythrocytes in the oxygen-free environment increased the content of extracellular hemoglobin by ∼1.5 fold. Paired measurements of hemoglobin and ATP content in the same samples, showed a positive correlation between hemolysis and ATP release. Comparative analysis of SDS-PAGE electrophoresis of erythrocyte ghosts obtained under control and deoxygenated conditions revealed a ∼2-fold elevation of the content of membrane-bound protein with Mr of ∼60 kDa. Conclusion: Deoxygenation of human erythrocytes affects composition of membrane-bound proteins. Additional experiments should be performed to identify the molecular origin of 60 kDa protein and its role in the attenuation of erythrocyte integrity and ATP release in hypoxic conditions.

  17. The Tzs protein and exogenous cytokinin affect virulence gene expression and bacterial growth of Agrobacterium tumefaciens.

    Science.gov (United States)

    Hwang, Hau-Hsuan; Yang, Fong-Jhih; Cheng, Tun-Fang; Chen, Yi-Chun; Lee, Ying-Ling; Tsai, Yun-Long; Lai, Erh-Min

    2013-09-01

    The soil phytopathogen Agrobacterium tumefaciens causes crown gall disease in a wide range of plant species. The neoplastic growth at the infection sites is caused by transferring, integrating, and expressing transfer DNA (T-DNA) from A. tumefaciens into plant cells. A trans-zeatin synthesizing (tzs) gene is located in the nopaline-type tumor-inducing plasmid and causes trans-zeatin production in A. tumefaciens. Similar to known virulence (Vir) proteins that are induced by the vir gene inducer acetosyringone (AS) at acidic pH 5.5, Tzs protein is highly induced by AS under this growth condition but also constitutively expressed and moderately upregulated by AS at neutral pH 7.0. We found that the promoter activities and protein levels of several AS-induced vir genes increased in the tzs deletion mutant, a mutant with decreased tumorigenesis and transient transformation efficiencies, in Arabidopsis roots. During AS induction and infection of Arabidopsis roots, the tzs deletion mutant conferred impaired growth, which could be rescued by genetic complementation and supplementing exogenous cytokinin. Exogenous cytokinin also repressed vir promoter activities and Vir protein accumulation in both the wild-type and tzs mutant bacteria with AS induction. Thus, the tzs gene or its product, cytokinin, may be involved in regulating AS-induced vir gene expression and, therefore, affect bacterial growth and virulence during A. tumefaciens infection.

  18. Protein kinase C affects the internalization and recycling of organic anion transporting polypeptide 1B1.

    Science.gov (United States)

    Hong, Mei; Hong, Weifang; Ni, Chunxu; Huang, Jiujiu; Zhou, Chao

    2015-10-01

    Organic anion-transporting polypeptides are members of the solute carrier (SLC) family and key determinants for the transmembrane transport of a wide variety of compounds. OATP1B1 is predominantly expressed at the basolateral membrane of human hepatocytes and play an important role in drug clearance from the body. It has been demonstrated to be responsible for the hepatic uptake of various drugs. Computer-based hydropathy analysis predicted several putative phosphorylation sites at the amino and carboxyl termini and at intracellular loop 3 of OATP family members. Therefore, their transport functions may be regulated by phosphorylation. Previous studies have demonstrated that uptake function of OATP2B1 and OATP1A2 is regulated by protein kinase C (PKC). In the present study, we treated HEK293 cells stably expressing OATP1B1 with different PKC modulators and measured their transport activity for prototypic substrate estrone-3-sulfate. It was found that OATP1B1 uptake function was reduced upon PKC activation. Further studies indicated that PKC may affect OATP1B1 activity through regulation of the cell surface protein level. Moreover, we found out that PKC activator phorbol 12-myristate 13-acetate (PMA) not only affects the internalization of OATP1B1 but its recycling as well. Immunocytochemistry analysis revealed that internalized OATP1B1 co-localized with early and recycling endosomal markers and the co-localization of OATP1B1 with recycling endosome is dependent on PKC activation. Taken together, our present study demonstrated that PKC regulates the function of OATP1B1 by affecting internalization and recycling of the transporter protein. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Does whey protein supplementation affect blood pressure in hypoalbuminemic peritoneal dialysis patients?

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    Hassan K

    2017-08-01

    Full Text Available Kamal Hassan,1,2 Fadi Hassan3 1Faculty of Medicine in the Galilee, Bar-Ilan University, Safed, 2Department of Nephrology and Hypertension, Peritoneal Dialysis Unit, 3Department of Internal Medicine E, Galilee Medical Center, Nahariya, Israel Objective: Hypertension and hypoalbuminemia are common risk factors for cardiovascular complications in peritoneal dialysis (PD patients. Data are limited regarding the effects of whey protein consumption on blood pressure in this population. The aim of the present study was to examine if whey protein supplementation for 12 weeks to hypoalbuminemic PD patients affects their blood pressure.Patients and methods: This prospective randomized study included 36 stable PD patients with serum albumin levels <3.8 g/dL. During 12 weeks, 18 patients were instructed to consume 1.2 g/kg/day of protein and an additional whey protein supplement at a dose of 25% of the instructed daily protein (whey protein group. Eighteen patients were instructed to consume protein in the amount of 1.2 g/kg/day and an additional 25%, without whey protein supplementation (control group. Results: Compared to the control group, in the whey protein group, serum albumin levels, oncotic pressure, and dialysate ultrafiltration significantly increased (3.55±0.14 to 4.08±0.15 g/dL, P<0.001; 21.81±2.03 to 24.06±1.54 mmHg, P<0.001; 927.8±120.3 to 1,125.0±125.1 mL/day, P<0.001; respectively and were significantly higher after 12 weeks (4.08±0.15 vs 3.41±0.49 g/dL, P<0.001; 24.06±1.54 vs 22.71±1.77 mmHg, P=0.010; 1,125.0±125.1 vs 930.6±352.8 mL/day, P=0.017; respectively in the whey protein group compared to the control group. Fluid overload, the extracellular to intracellular ratio and mean arterial pressure (MAP significantly decreased (2.46±1.08 to 1.52±0.33, P<0.001; 1.080±0.142 to 0.954±0.124, P<0.001; 102.6±3.80 to 99.83±3.85, P=0.018; respectively and were significantly lower in the whey protein group after 12 weeks (1.52±0

  20. Pregnancy-induced Changes in Corneal Biomechanics and Topography Are Thyroid Hormone Related.

    Science.gov (United States)

    Tabibian, David; de Tejada, Begoña M; Gatzioufas, Zisis; Kling, Sabine; Meiss, Vanessa S; Boldi, Marc-Olivier; Othenin-Girard, Véronique; Chilin, Antonina; Lambiel, Julien; Hoogewoud, Florence; Hafezi, Farhad

    2017-12-01

    To identify biomechanical and topographic changes of the cornea during pregnancy and the postpartum period and its association to hormonal changes. Prospective single-center observational cohort study. Participants were 24 pregnant women (48 eyes), monitored throughout pregnancy and after delivery. Biomechanical and topographic corneal properties were measured using the Ocular Response Analyzer (ORA) and a Scheimpflug imaging system (Pentacam HR) each trimester and 1 month after delivery. At the same consultations blood plasma levels of estradiol (E2) and thyroid hormones (TSH, T3t, T4t) were also determined. A factorial MANCOVA was used to detect interactions between hormonal plasma levels and ocular parameters. Significant differences in corneal biomechanical and topographic parameters were found during pregnancy in relation to T3t (p = .01), T4t (p < .001), T3t/T4t (P = .001), and TSH (p = .001) plasma levels. E2 plasma levels (p = .092) and time period of measurement (p = .975) did not significantly affect corneal parameters. TSH levels significantly affected the maximal keratometry reading (p = .036), the vertical keratometry reading (p = .04), and the index of height asymmetry (p = .014). Those results persist after excluding hypothyroidism patients from the statistical analysis. Hormonal changes affecting corneal biomechanics and topography during pregnancy could be thyroid related. Dysthyroidism may directly influence corneal biomechanics and represents a clinically relevant factor that needs further investigation. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Intraepithelial and interstitial deposition of pathological prion protein in kidneys of scrapie-affected sheep.

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    Ciriaco Ligios

    Full Text Available Prions have been documented in extra-neuronal and extra-lymphatic tissues of humans and various ruminants affected by Transmissible Spongiform Encephalopathy (TSE. The presence of prion infectivity detected in cervid and ovine blood tempted us to reason that kidney, the organ filtrating blood derived proteins, may accumulate disease associated PrP(Sc. We collected and screened kidneys of experimentally, naturally scrapie-affected and control sheep for renal deposition of PrP(Sc from distinct, geographically separated flocks. By performing Western blot, PET blot analysis and immunohistochemistry we found intraepithelial (cortex, medulla and papilla and occasional interstitial (papilla deposition of PrP(Sc in kidneys of scrapie-affected sheep. Interestingly, glomerula lacked detectable signals indicative of PrP(Sc. PrP(Sc was also detected in kidneys of subclinical sheep, but to significantly lower degree. Depending on the stage of the disease the incidence of PrP(Sc in kidney varied from approximately 27% (subclinical to 73.6% (clinical in naturally scrapie-affected sheep. Kidneys from flocks without scrapie outbreak were devoid of PrP(Sc. Here we demonstrate unexpectedly frequent deposition of high levels of PrP(Sc in ovine kidneys of various flocks. Renal deposition of PrP(Sc is likely to be a pre-requisite enabling prionuria, a possible co-factor of horizontal prion-transmission in sheep.

  2. Expression of OATP family members in hormone-related cancers: potential markers of progression.

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    Heather Pressler

    Full Text Available The organic anion transporting polypeptide (OATP family of transporters has been implicated in prostate cancer disease progression probably by transporting hormones or drugs. In this study, we aimed to elucidate the expression, frequency, and relevance of OATPs as a biomarker in hormone-dependent cancers. We completed a study examining SLCO1B3, SLCO1B1 and SLCO2B1 mRNA expression in 381 primary, independent patient samples representing 21 cancers and normal tissues. From a separate cohort, protein expression of OATP1B3 was examined in prostate, colon, and bladder tissue. Based on expression frequency, SLCO2B1 was lower in liver cancer (P = 0.04 which also trended lower with decreasing differentiation (P = 0.004 and lower magnitude in pancreatic cancer (P = 0.05. SLCO2B1 also had a higher frequency in thyroid cancer (67% than normal (0% and expression increased with stage (P = 0.04. SLCO1B3 was expressed in 52% of cancerous prostate samples and increased SLCO1B3 expression trended with higher Gleason score (P = 0.03. SLCO1B3 expression was also higher in testicular cancer (P = 0.02. SLCO1B1 expression was lower in liver cancer (P = 0.04 which trended lower with liver cancer grade (P = 0.0004 and higher with colon cancer grade (P = 0.05. Protein expression of OATP1B3 was examined in normal and cancerous prostate, colon, and bladder tissue samples from an independent cohort. The results were similar to the transcription data, but showed distinct localization. OATPs correlate to differentiation in certain hormone-dependent cancers, thus may be useful as biomarkers for assessing clinical treatment and stage of disease.

  3. Polycomb Protein OsFIE2 Affects Plant Height and Grain Yield in Rice.

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    Xianbo Liu

    Full Text Available Polycomb group (PcG proteins have been shown to affect growth and development in plants. To further elucidate their role in these processes in rice, we isolated and characterized a rice mutant which exhibits dwarfism, reduced seed setting rate, defective floral organ, and small grains. Map-based cloning revealed that abnormal phenotypes were attributed to a mutation of the Fertilization Independent Endosperm 2 (OsFIE2 protein, which belongs to the PcG protein family. So we named the mutant as osfie2-1. Histological analysis revealed that the number of longitudinal cells in the internodes decreased in osfie2-1, and that lateral cell layer of the internodes was markedly thinner than wild-type. In addition, compared to wild-type, the number of large and small vascular bundles decreased in osfie2-1, as well as cell number and cell size in spikelet hulls. OsFIE2 is expressed in most tissues and the coded protein localizes in both nucleus and cytoplasm. Yeast two-hybrid and bimolecular fluorescence complementation assays demonstrated that OsFIE2 interacts with OsiEZ1 which encodes an enhancer of zeste protein previously identified as a histone methylation enzyme. RNA sequencing-based transcriptome profiling and qRT-PCR analysis revealed that some homeotic genes and genes involved in endosperm starch synthesis, cell division/expansion and hormone synthesis and signaling are differentially expressed between osfie2-1 and wild-type. In addition, the contents of IAA, GA3, ABA, JA and SA in osfie2-1 are significantly different from those in wild-type. Taken together, these results indicate that OsFIE2 plays an important role in the regulation of plant height and grain yield in rice.

  4. Codon usage regulates protein structure and function by affecting translation elongation speed in Drosophila cells.

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    Zhao, Fangzhou; Yu, Chien-Hung; Liu, Yi

    2017-08-21

    Codon usage biases are found in all eukaryotic and prokaryotic genomes and have been proposed to regulate different aspects of translation process. Codon optimality has been shown to regulate translation elongation speed in fungal systems, but its effect on translation elongation speed in animal systems is not clear. In this study, we used a Drosophila cell-free translation system to directly compare the velocity of mRNA translation elongation. Our results demonstrate that optimal synonymous codons speed up translation elongation while non-optimal codons slow down translation. In addition, codon usage regulates ribosome movement and stalling on mRNA during translation. Finally, we show that codon usage affects protein structure and function in vitro and in Drosophila cells. Together, these results suggest that the effect of codon usage on translation elongation speed is a conserved mechanism from fungi to animals that can affect protein folding in eukaryotic organisms. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  5. Maternal protein restriction affects gene expression and enzyme activity of intestinal disaccharidases in adult rat offspring

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    Pinheiro, D.F.; Pacheco, P.D.G.; Alvarenga, P.V.; Buratini, J. Jr; Castilho, A.C.S.; Lima, P.F.; Sartori, D.R.S.; Vicentini-Paulino, M.L.M. [Departamento de Fisiologia, Instituto de Biociências, Universidade Estadual Paulista, Botucatu, SP (Brazil)

    2013-03-15

    This study investigated the consequences of intrauterine protein restriction on the gastrointestinal tract and particularly on the gene expression and activity of intestinal disaccharidases in the adult offspring. Wistar rat dams were fed isocaloric diets containing 6% protein (restricted, n = 8) or 17% protein (control, n = 8) throughout gestation. Male offspring (n = 5-8 in each group) were evaluated at 3 or 16 weeks of age. Maternal protein restriction during pregnancy produced offspring with growth restriction from birth (5.7 ± 0.1 vs 6.3 ± 0.1 g; mean ± SE) to weaning (42.4 ± 1.3 vs 49.1 ± 1.6 g), although at 16 weeks of age their body weight was similar to control (421.7 ± 8.9 and 428.5 ± 8.5 g). Maternal protein restriction also increased lactase activity in the proximal (0.23 ± 0.02 vs 0.15 ± 0.02), medial (0.30 ± 0.06 vs 0.14 ± 0.01) and distal (0.43 ± 0.07 vs 0.07 ± 0.02 U·g{sup -1}·min{sup -1}) small intestine, and mRNA lactase abundance in the proximal intestine (7.96 ± 1.11 vs 2.38 ± 0.47 relative units) of 3-week-old offspring rats. In addition, maternal protein restriction increased sucrase activity (1.20 ± 0.02 vs 0.91 ± 0.02 U·g{sup -1}·min{sup -1}) and sucrase mRNA abundance (4.48 ± 0.51 vs 1.95 ± 0.17 relative units) in the duodenum of 16-week-old rats. In conclusion, the present study shows for the first time that intrauterine protein restriction affects gene expression of intestinal enzymes in offspring.

  6. Quality of buffalo milk as affected by dietary protein level and flaxseed supplementation.

    Science.gov (United States)

    Santillo, A; Caroprese, M; Marino, R; Sevi, A; Albenzio, M

    2016-10-01

    The aim of the present research was to evaluate the effects of protein level and flaxseed supplementation on the yield and quality of buffalo milk. In particular, the fatty acid profile of milk from buffalo cows subjected to different diets has been investigated. A 2×3 factorial design was tested with buffalo cows receiving 2 dietary crude protein (CP) and 3 flaxseed (FS) supplementation levels. Treatments were (1) low dietary CP level [12% of dry matter (DM)] and no flaxseed supplementation (LP); (2) low dietary CP level (12% of DM) and low flaxseed supplementation (500g/d) (LPFS500); (3) low dietary CP level (12% of DM) and moderate flaxseed supplementation (1,000g/d) (LPFS1000); (4) moderate dietary CP level (15% of DM) and no flaxseed supplementation (MP); (5) moderate dietary CP level (15% of DM) and low flaxseed supplementation (500g/d) (MPFS500); and (6) moderate dietary CP level (15% of DM) and moderate flaxseed supplementation (1,000g/d) (MPFS1000). Milk protein and casein were affected by flaxseed supplementation being higher in MP, intermediate in LP, and lower in flaxseed-supplemented diets. However, the results from the present study highlighted that low protein diets sustained milk yield, protein, and casein synthesis in milk when whole flaxseed was administered. Short-chain fatty acids, in particular C8:0 and C10:0, were the lowest in milk from buffalo cows fed the highest level of flaxseed supplementation. Medium-chain fatty acids were the lowest in FS1000, intermediate in FS500, and the highest in the HP and LP groups. Long-chain fatty acids were the highest in FS1000, intermediate in FS500 groups, and the lowest in milk from buffalo receiving no flaxseed supplementation. Protein level of the diet influenced the percentage of C18:0, which was higher in MP than LP groups. Total conjugated linoleic acid content evidenced the same trend of long-chain fatty acids, with an increase of about 7% in FL500 and of 22% in FL1000 than the control. Apart from

  7. A temperature sensitive VSV identifies L protein residues that affect transcription but not replication.

    Science.gov (United States)

    Galloway, Summer E; Wertz, Gail W

    2009-06-05

    To investigate the polymerase components selectively involved in transcription versus replication of vesicular stomatitis virus (VSV), we sequenced the polymerase gene of a conditionally RNA defective, temperature sensitive VSV: ts(G)114, which has a phenotype upon shift from permissive to non-permissive temperature of shut-down of mRNA transcription and unaffected genome replication. Sequence analysis of the ts(G)114 L gene identified three altered amino acid residues in the L protein. These three changes were specifically engineered individually and in combinations into a functional cDNA clone encoding the VSV genome and tested for association with the temperature sensitive and RNA defective phenotypes in the background of recovered engineered viruses. The data presented in this study show a specific amino acid substitution in domain II of the VSV L protein that significantly affects total RNA synthesis, but when in combination with two additional amino acid substitutions identified in the ts(G)114 L protein, leads to a specific reduction in mRNA transcription, but not replication.

  8. Identification of archaeal proteins that affect the exosome function in vitro

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    Palhano Fernando L

    2010-05-01

    Full Text Available Abstract Background The archaeal exosome is formed by a hexameric RNase PH ring and three RNA binding subunits and has been shown to bind and degrade RNA in vitro. Despite extensive studies on the eukaryotic exosome and on the proteins interacting with this complex, little information is yet available on the identification and function of archaeal exosome regulatory factors. Results Here, we show that the proteins PaSBDS and PaNip7, which bind preferentially to poly-A and AU-rich RNAs, respectively, affect the Pyrococcus abyssi exosome activity in vitro. PaSBDS inhibits slightly degradation of a poly-rA substrate, while PaNip7 strongly inhibits the degradation of poly-A and poly-AU by the exosome. The exosome inhibition by PaNip7 appears to depend at least partially on its interaction with RNA, since mutants of PaNip7 that no longer bind RNA, inhibit the exosome less strongly. We also show that FITC-labeled PaNip7 associates with the exosome in the absence of substrate RNA. Conclusions Given the high structural homology between the archaeal and eukaryotic proteins, the effect of archaeal Nip7 and SBDS on the exosome provides a model for an evolutionarily conserved exosome control mechanism.

  9. Sucrose Sensitivity of Honey Bees Is Differently Affected by Dietary Protein and a Neonicotinoid Pesticide.

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    Fabien J Démares

    Full Text Available Over a decade, declines in honey bee colonies have raised worldwide concerns. Several potentially contributing factors have been investigated, e.g. parasites, diseases, and pesticides. Neonicotinoid pesticides have received much attention due to their intensive use in crop protection, and their adverse effects on many levels of honey bee physiology led the European Union to ban these compounds. Due to their neuronal target, a receptor expressed throughout the insect nervous system, studies have focused mainly on neuroscience and behaviour. Through the Geometric Framework of nutrition, we investigated effects of the neonicotinoid thiamethoxam on survival, food consumption and sucrose sensitivity of honey bees (Apis mellifera. Thiamethoxam did not affect protein and carbohydrate intake, but decreased responses to high concentrations of sucrose. Interestingly, when bees ate fixed unbalanced diets, dietary protein facilitated better sucrose detection. Both thiamethoxam and dietary protein influenced survival. These findings suggest that, in the presence of a pesticide and unbalanced food, honey bee health may be severely challenged. Consequences for foraging efficiency and colony activity, cornerstones of honey bee health, are also discussed.

  10. Sucrose Sensitivity of Honey Bees Is Differently Affected by Dietary Protein and a Neonicotinoid Pesticide

    Science.gov (United States)

    Démares, Fabien J.; Crous, Kendall L.; Pirk, Christian W. W.; Nicolson, Susan W.; Human, Hannelie

    2016-01-01

    Over a decade, declines in honey bee colonies have raised worldwide concerns. Several potentially contributing factors have been investigated, e.g. parasites, diseases, and pesticides. Neonicotinoid pesticides have received much attention due to their intensive use in crop protection, and their adverse effects on many levels of honey bee physiology led the European Union to ban these compounds. Due to their neuronal target, a receptor expressed throughout the insect nervous system, studies have focused mainly on neuroscience and behaviour. Through the Geometric Framework of nutrition, we investigated effects of the neonicotinoid thiamethoxam on survival, food consumption and sucrose sensitivity of honey bees (Apis mellifera). Thiamethoxam did not affect protein and carbohydrate intake, but decreased responses to high concentrations of sucrose. Interestingly, when bees ate fixed unbalanced diets, dietary protein facilitated better sucrose detection. Both thiamethoxam and dietary protein influenced survival. These findings suggest that, in the presence of a pesticide and unbalanced food, honey bee health may be severely challenged. Consequences for foraging efficiency and colony activity, cornerstones of honey bee health, are also discussed. PMID:27272274

  11. Dietary protein during gestation affects circulating indicators of placental function and fetal development in heifers.

    Science.gov (United States)

    Sullivan, T M; Micke, G C; Magalhaes, R S; Martin, G B; Wallace, C R; Green, J A; Perry, V E A

    2009-04-01

    The influences of nutritional protein during the first and second trimesters of pregnancy on placental hormones and fetal growth were determined in composite beef heifers. At artificial insemination, heifers were stratified by weight within each composite genotype into 4 treatment groups: High High (HH=1.4kg crude protein (CP)/day for first and second trimesters of gestation; n=16), High Low (HL=1.4kg CP/day for first trimester and 0.4kg CP/day for second trimester; n=19), Low High (LH=0.4kg CP/day for first trimester and 1.4kg CP/day for second trimester; n=17) or Low Low (LL=0.4kg CP/day for first and second trimesters; n=19). Maternal plasma bovine pregnancy associated glycoprotein (bPAG) and progesterone (P4) were determined at gestation day (gd) 28, 82, 179 and 271 (mean gestation length 286 days) in addition to P4 at term. Estrone sulphate (ES) and bovine placental lactogen (bPL) concentrations were measured at gd 124, 179, 236 and 271 and at term in addition to ES at gd 82. Low dietary protein increased placental function as indicated by increased bPAG (P<0.001) and ES (P=0.02) concentrations in first trimester and increased bPL concentrations (P=0.01) in the second trimester of gestation. In the third trimester, when dietary treatment had ceased, placental function was no longer associated with previous dietary treatments. Dam genotype affected placental function as measured by bPL (P<0.001) and ES concentrations (P=0.02). Calf gender, heifer age and maternal insulin-like growth factor (IGF)-I, -II and leptin did not affect hormonal indicators or circulating markers of placental function. Enhanced placental function during the third trimester, as measured by ES, was associated with increased calf birth weight (P=0.003).

  12. In vitro antioxidant activity of rice protein affected by alkaline degree and gastrointestinal protease digestion.

    Science.gov (United States)

    Liu, Ye; Wang, Zhengxuan; Li, Hui; Liang, Mingcai; Yang, Lin

    2016-12-01

    To elucidate whether and how alkali treatment, which is a common process for rice protein (RP) extraction, affects antioxidant activity of RP, the different degree of alkali (from 0.1% to 0.4% of NaOH) was used to extract RP (RP-1, RP-2, RP-3, RP-4). The antioxidant capacities of scavenging free radicals [2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid] diammonium salt, ABTS; 1,1-diphenyl-2-picrylhydrazyl, DPPH), chelating metals (iron, copper) and reducing power investigated in the hydrolysates of RPs (RP-1, RP-2, RP-3, RP-4) during in vitro pepsin-pancreatin digestion were effectively affected by alkali treatment. The present study demonstrated that the weakest antioxidant responses to ABTS radical-scavenging activity, DPPH radical-scavenging activity, iron chelating activity, copper chelating activity and reducing power were produced by RP-4 extracted by the highest alkali proportion (0.4% NaOH). The present study indicates that antioxidant capacity of RP could be more readily depressed by strict alkali degree and affected by gastrointestinal proteases. Results suggest that alkali extraction is a vital process to regulate the antioxidant activity of RP through modifying the compositions of amino acids, which are dependent on alkali magnitude. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  13. Amyloid beta (A4) precursor protein expression in human periodontitis-affected gingival tissues.

    Science.gov (United States)

    Kubota, T; Maruyama, S; Abe, D; Tomita, T; Morozumi, T; Nakasone, N; Saku, T; Yoshie, H

    2014-06-01

    Periodontitis involves periodontal tissue destruction and is associated with chronic inflammation and ageing. Periodontitis has recently been recognised as a risk factor for Alzheimer's disease (AD). We showed upregulation of molecules in the AD pathway including amyloid beta (A4) precursor protein (APP), a key gene in AD, interleukin-1 beta (IL-1β), and complement component 1 (q subcomponent, A chain) (C1QA) in periodontitis compared to healthy tissues. Here, we quantitatively analysed the expression levels of APP, IL-1β, and C1QA and determined the localisation of APP in gingival tissues. Fourteen chronic periodontitis patients and 14 healthy participants were enrolled. Six samples of total RNA from two distinct sites of healthy and periodontitis-affected gingival tissues from three randomly selected patients were used for microarray analyses, and significant biological pathways in periodontitis were identified. Differential gene expression of APP, IL-1β, and C1QA, which belong to the AD pathway, were analysed with quantitative reverse transcription real-time polymerase chain reaction (qRT-PCR) using samples from these 14 chronic periodontitis patients and 14 healthy controls. APP localisation was analysed with immunohistochemistry. APP, IL-1β, and C1QA mRNA levels were significantly upregulated in periodontitis-affected gingival tissues. APP was mainly localised in macrophages in gingival connective tissues underneath the epithelial layers. An association between AD and periodontitis was detected with microarray and computer-aided data mining analyses. qRT-PCR identified differential gene expression in periodontitis-affected gingival tissue that may be related to AD pathogenesis. Elevated APP, IL-1β, and C1QA transcripts and APP-expressing macrophages in periodontitis-affected gingival tissues were observed, suggesting a relationship between periodontitis and AD pathogenesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Cholesterol Promotes Protein Binding by Affecting Membrane Electrostatics and Solvation Properties.

    Science.gov (United States)

    Doktorova, Milka; Heberle, Frederick A; Kingston, Richard L; Khelashvili, George; Cuendet, Michel A; Wen, Yi; Katsaras, John; Feigenson, Gerald W; Vogt, Volker M; Dick, Robert A

    2017-11-07

    Binding of the retroviral structural protein Gag to the cellular plasma membrane is mediated by the protein's matrix (MA) domain. Prominent among MA-PM interactions is electrostatic attraction between the positively charged MA domain and the negatively charged plasma membrane inner leaflet. Previously, we reported that membrane association of HIV-1 Gag, as well as purified Rous sarcoma virus (RSV) MA and Gag, depends strongly on the presence of acidic lipids and is enhanced by cholesterol (Chol). The mechanism underlying this enhancement was unclear. Here, using a broad set of in vitro and in silico techniques we addressed molecular mechanisms of association between RSV MA and model membranes, and investigated how Chol enhances this association. In neutron scattering experiments with liposomes in the presence or absence of Chol, MA preferentially interacted with preexisting POPS-rich clusters formed by nonideal lipid mixing, binding peripherally to the lipid headgroups with minimal perturbation to the bilayer structure. Molecular dynamics simulations showed a stronger MA-bilayer interaction in the presence of Chol, and a large Chol-driven increase in lipid packing and membrane surface charge density. Although in vitro MA-liposome association is influenced by disparate variables, including ionic strength and concentrations of Chol and charged lipids, continuum electrostatic theory revealed an underlying dependence on membrane surface potential. Together, these results conclusively show that Chol affects RSV MA-membrane association by making the electrostatic potential at the membrane surface more negative, while decreasing the penalty for lipid headgroup desolvation. The presented approach can be applied to other viral and nonviral proteins. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  15. Pathophysiological changes that affect drug disposition in protein-energy malnourished children

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    Oshikoya Kazeem A

    2009-12-01

    Full Text Available Abstract Protein-energy malnutrition (PEM is a major public health problem affecting a high proportion of infants and older children world-wide and accounts for a high childhood morbidity and mortality in the developing countries. The epidemiology of PEM has been extensively studied globally and management guidelines formulated by the World Health Organization (WHO. A wide spectrum of infections such as measles, malaria, acute respiratory tract infection, intestinal parasitosis, tuberculosis and HIV/AIDS may complicate PEM with two or more infections co-existing. Thus, numerous drugs may be required to treat the patients. In-spite of abundant literature on the epidemiology and management of PEM, focus on metabolism and therapeutic drug monitoring is lacking. A sound knowledge of pathophysiology of PEM and pharmacology of the drugs frequently used for their treatment is required for safe and rational treatment. In this review, we discuss the pathophysiological changes in children with PEM that may affect the disposition of drugs frequently used for their treatment. This review has established abnormal disposition of drugs in children with PEM that may require dosage modification. However, the relevance of these abnormalities to the clinical management of PEM remains inconclusive. At present, there are no good indications for drug dosage modification in PEM; but for drug safety purposes, further studies are required to accurately determine dosages of drugs frequently used for children with PEM.

  16. Daytime pattern of post-exercise protein intake affects whole-body protein turnover in resistance-trained males

    Directory of Open Access Journals (Sweden)

    Moore Daniel R

    2012-10-01

    Full Text Available Abstract Background The pattern of protein intake following exercise may impact whole-body protein turnover and net protein retention. We determined the effects of different protein feeding strategies on protein metabolism in resistance-trained young men. Methods Participants were randomly assigned to ingest either 80g of whey protein as 8x10g every 1.5h (PULSE; n=8, 4x20g every 3h (intermediate, INT; n=7, or 2x40g every 6h (BOLUS; n=8 after an acute bout of bilateral knee extension exercise (4x10 repetitions at 80% maximal strength. Whole-body protein turnover (Q, synthesis (S, breakdown (B, and net balance (NB were measured throughout 12h of recovery by a bolus ingestion of [15N]glycine with urinary [15N]ammonia enrichment as the collected end-product. Results PULSE Q rates were greater than BOLUS (~19%, P Conclusion We conclude that the pattern of ingested protein, and not only the total daily amount, can impact whole-body protein metabolism. Individuals aiming to maximize NB would likely benefit from repeated ingestion of moderate amounts of protein (~20g at regular intervals (~3h throughout the day.

  17. Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair

    Energy Technology Data Exchange (ETDEWEB)

    Gustafsson, Ann-Sofie, E-mail: ann-sofie.gustafsson@bms.uu.se; Abramenkovs, Andris; Stenerlöw, Bo

    2014-11-15

    Highlights: • We reduced the level of DNA-PKcs with siRNA and examined cells after γ-irradiation. • Low DNA-PKcs levels lead to radiosensitivity but did not affect repair of DSB. • Low DNA-PKcs levels may block progression of mitosis. • DNA-PKcs role in mitotic progression is independent of its role in DSB repair. • We suggest different mechanisms by which loss of DNA-PKcs function sensitize cells. - Abstract: Efficient and correct repair of DNA double-strand break (DSB) is critical for cell survival. Defects in the DNA repair may lead to cell death, genomic instability and development of cancer. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of the non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. In the present study, by using siRNA against DNA-PKcs in four human cell lines, we examined how low levels of DNA-PKcs affected cellular response to ionizing radiation. Decrease of DNA-PKcs levels by 80–95%, induced by siRNA treatment, lead to extreme radiosensitivity, similar to that seen in cells completely lacking DNA-PKcs and low levels of DNA-PKcs promoted cell accumulation in G2/M phase after irradiation and blocked progression of mitosis. Surprisingly, low levels of DNA-PKcs did not affect the repair capacity and the removal of 53BP1 or γ-H2AX foci and rejoining of DSB appeared normal. This was in strong contrast to cells completely lacking DNA-PKcs and cells treated with the DNA-PKcs inhibitor NU7441, in which DSB repair were severely compromised. This suggests that there are different mechanisms by which loss of DNA-PKcs functions can sensitize cells to ionizing radiation. Further, foci of phosphorylated DNA-PKcs (T2609 and S2056) co-localized with DSB and this was independent of the amount of DNA-PKcs but foci of DNA-PKcs was only seen in siRNA-treated cells. Our study emphasizes on the critical role of DNA-PKcs for maintaining survival after radiation exposure

  18. Humoral Hypercalcemia of Malignancy with a Parathyroid Hormone-Related Peptide-Secreting Intrahepatic Cholangiocarcinoma Accompanied by a Gastric Cancer

    Directory of Open Access Journals (Sweden)

    Katsushi Takeda

    2017-01-01

    Full Text Available Humoral hypercalcemia of malignancy (HHM is caused by the oversecretion of parathyroid hormone-related peptide (PTHrP from malignant tumors. Although any tumor may cause HHM, that induced by intrahepatic cholangiocarcinoma (ICC or gastric cancer (GC is rare. We report here a 74-year-old male who displayed HHM with both ICC and GC and showed an elevated serum PTHrP level. Treatment of the hypercalcemia with saline, furosemide, elcatonin, and zoledronic acid corrected his serum calcium level and improved symptoms. Because treatment of ICC should precede that of GC, we chose chemotherapy with cisplatin (CDDP and gemcitabine (GEM. Chemotherapy reduced the size of the ICC and decreased the serum PTHrP level. One year after diagnosis, the patient was alive in the face of a poor prognosis for an ICC that produced PTHrP. Immunohistochemical staining for PTHrP was positive for the ICC and negative for the GC, leading us to believe that the cause of the HHM was a PTHrP-secreting ICC. In conclusion, immunohistochemical staining for PTHrP may be useful in discovering the cause of HHM in the case of two cancers accompanied by an elevated serum PHTrP level. Chemotherapy with CDDP and GEM may be the most appropriate treatment for a PTHrP-secreting ICC.

  19. The Phenotype of Hormone-Related Allergic and Autoimmune Diseases in the Skin: Annular Lesions That Lateralize

    Directory of Open Access Journals (Sweden)

    Ramya Kollipara

    2012-01-01

    Full Text Available Introduction. Sexual dimorphism with an increased prevalence in women has long been observed in various autoimmune, allergic, and skin diseases. Recent research has attempted to correlate this female predilection to physiologic changes seen in the menstrual cycle in order to more effectively diagnose and treat these diseases. Cases. We present five cases of cutaneous diseases in women with annular morphology and distributive features that favor one side over the other. In all cases, skin disease improved with ovarian suppression. Conclusion. Sexual dimorphism in the innate and adaptive immune systems has long been observed, with females demonstrating a more vigorous immune response compared to males. Female sex hormones promote T and B lymphocyte autoreactivity and favor the humoral arm of adaptive immunity. In addition to ovarian steroidogenesis and immunity, intricate pathways coexist in order to engage a single oocyte in each cycle, while simultaneously sustaining the ovarian reserve. Vigorous proinflammatory, vasoactive, and pigment-related cytokines emerge during the demise of the corpus luteum, influencing peripherical sex hormone metabolism\tof the level of the macrophage and fibroblast. We propose that annular and lateralizing lesions are important manifestations of hormone-related inflammation and recognition of this linkage can lead to improved immune and reproductive health.

  20. Glycosaminoglycan sulphation affects the seeded misfolding of a mutant prion protein.

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    Victoria A Lawson

    Full Text Available BACKGROUND: The accumulation of protease resistant conformers of the prion protein (PrP(res is a key pathological feature of prion diseases. Polyanions, including RNA and glycosaminoglycans have been identified as factors that contribute to the propagation, transmission and pathogenesis of prion disease. Recent studies have suggested that the contribution of these cofactors to prion propagation may be species specific. METHODOLOGY/PRINCIPAL FINDING: In this study a cell-free assay was used to investigate the molecular basis of polyanion stimulated PrP(res formation using brain tissue or cell line derived murine PrP. Enzymatic depletion of endogenous nucleic acids or heparan sulphate (HS from the PrP(C substrate was found to specifically prevent PrP(res formation seeded by mouse derived PrP(Sc. Modification of the negative charge afforded by the sulphation of glycosaminoglycans increased the ability of a familial PrP mutant to act as a substrate for PrP(res formation, while having no effect on PrP(res formed by wildtype PrP. This difference may be due to the observed differences in the binding of wild type and mutant PrP for glycosaminoglycans. CONCLUSIONS/SIGNIFICANCE: Cofactor requirements for PrP(res formation are host species and prion strain specific and affected by disease associated mutations of the prion protein. This may explain both species and strain dependent propagation characteristics and provide insights into the underlying mechanisms of familial prion disease. It further highlights the challenge of designing effective therapeutics against a disease which effects a range of mammalian species, caused by range of aetiologies and prion strains.

  1. Reproductive and hormone-related risk factors for epithelial ovarian cancer by histologic pathways, invasiveness and histologic subtypes : Results from the EPIC cohort

    NARCIS (Netherlands)

    Fortner, Renée T.; Ose, Jennifer; Merritt, Melissa A.; Schock, Helena; Tjønneland, Anne; Hansen, Louise; Overvad, Kim; Dossus, Laure; Clavel-Chapelon, Françoise; Baglietto, Laura; Boeing, Heiner; Trichopoulou, Antonia; Benetou, Vassiliki; Lagiou, Pagona; Agnoli, Claudia; Mattiello, Amalia; Masala, Giovanna; Tumino, Rosario; Sacerdote, Carlotta; Bueno-De-Mesquita, H. B.|info:eu-repo/dai/nl/06929528X; Onland-Moret, N. Charlotte|info:eu-repo/dai/nl/26504362X; Peeters, Petra H.|info:eu-repo/dai/nl/074099655; Weiderpass, Elisabete; Torhild Gram, Inger; Duell, Eric J.; Larrañaga, Nerea; Ardanaz, Eva; Sánchez, María José; Chirlaque, M. D.; Brändstedt, Jenny; Idahl, Annika; Lundin, Eva; Khaw, Kay Tee; Wareham, Nick; Travis, Ruth C.; Rinaldi, Sabina; Romieu, Isabelle; Gunter, Marc J.; Riboli, Elio; Kaaks, Rudolf

    2015-01-01

    Whether risk factors for epithelial ovarian cancer (EOC) differ by subtype (i.e., dualistic pathway of carcinogenesis, histologic subtype) is not well understood; however, data to date suggest risk factor differences. We examined associations between reproductive and hormone-related risk factors for

  2. Changes in gravity affect gene expression, protein modulation and metabolite pools of arabidopsis

    Science.gov (United States)

    Hampp, R.; Martzivanou, M.; Maier, R. M.; Magel, E.

    Callus cultures of Arabidopsis thaliana (cv. Columbia) in Petri dishes / suspension cultures were exposed to altered g-forces by centrifugation (1 to 10 g), klinorotation, and μ g (sounding rocket flights). Using semi-quantitative RT-PCR, transcripts of genes coding for metabolic key enzymes (ADP-glucose pyrophosphorylase, ADPG-PP; ß-amylase, fructose-1,6-bisphosphatase, FBPase; glyceraldehyde-P dehydrogenase, GAPDH; hydroxymethylglutaryl-CoA reductase, HMG; phenylalanine-ammonium-lyase, PAL; PEP carboxylase, PEPC) were used to monitor threshold conditions for g-number (all) and time of exposure (ß-amylase) which led to altered amounts of the gene product. Exposure to approx. 5 g and higher for 1h resulted in altered transcript levels: transcripts of ß-amylase, PAL, and PEPC were increased, those of ADPG-PP decreased, while those of FBPase, GAPDH, and HMG were not affected. This probably indicates a shift from starch synthesis to starch degradation and increased rates of anaplerosis (PEPC: supply of ketoacids for amino acid synthesis). In order to get more information about g-related effects on gene expression, we used a 1h-exposure to 7 g for a microarray analysis. Transcripts of more than 200 genes were significantly increased in amount (ratio 7g / 1g control; 21.6 and larger). They fall into several categories. Transcripts coding for enzymes of major pathways form the largest group (25%), followed by gene products involved in cellular organisation and cell wall formation / rearrangement (17%), signalling, phosphorylation/dephosphorylation (12%), proteolysis and transport (10% each), hormone synthesis plus related events (8%), defense (4%), stress-response (2%), and gravisensing (2%). Many of the alterations are part of a general stress response, but some changes related to the synthesis / rearrangement of cell wall components could be more hyper-g-specific. Using macroarrays with selected genes according to our hypergravity study (metabolism / signalling

  3. Protein Phosphorylation and Mineral Binding Affect the Secondary Structure of the Leucine-Rich Amelogenin Peptide

    Directory of Open Access Journals (Sweden)

    Hajime Yamazaki

    2017-06-01

    Full Text Available Previously, we have shown that serine-16 phosphorylation in native full-length porcine amelogenin (P173 and the Leucine-Rich Amelogenin Peptide (LRAP(+P, an alternative amelogenin splice product, affects protein assembly and mineralization in vitro. Notably, P173 and LRAP(+P stabilize amorphous calcium phosphate (ACP and inhibit hydroxyapatite (HA formation, while non-phosphorylated counterparts (rP172, LRAP(−P guide the growth of ordered bundles of HA crystals. Based on these findings, we hypothesize that the phosphorylation of full-length amelogenin and LRAP induces conformational changes that critically affect its capacity to interact with forming calcium phosphate mineral phases. To test this hypothesis, we have utilized Fourier transform infrared spectroscopy (FTIR to determine the secondary structure of LRAP(−P and LRAP(+P in the absence/presence of calcium and selected mineral phases relevant to amelogenesis; i.e., hydroxyapatite (HA: an enamel crystal prototype and (ACP: an enamel crystal precursor phase. Aqueous solutions of LRAP(−P or LRAP(+P were prepared with or without 7.5 mM of CaCl2 at pH 7.4. FTIR spectra of each solution were obtained using attenuated total reflectance, and amide-I peaks were analyzed to provide secondary structure information. Secondary structures of LRAP(+P and LRAP(−P were similarly assessed following incubation with suspensions of HA and pyrophosphate-stabilized ACP. Amide I spectra of LRAP(−P and LRAP(+P were found to be distinct from each other in all cases. Spectra analyses showed that LRAP(−P is comprised mostly of random coil and β-sheet, while LRAP(+P exhibits more β-sheet and α-helix with little random coil. With added Ca, the random coil content increased in LRAP(−P, while LRAP(+P exhibited a decrease in α-helix components. Incubation of LRAP(−P with HA or ACP resulted in comparable increases in β-sheet structure. Notably, however, LRAP(+P secondary structure was more affected by

  4. Bone morphogenetic protein Smads signaling in mesenchymal stem cells affected by osteoinductive calcium phosphate ceramics.

    Science.gov (United States)

    Tang, Zhurong; Wang, Zhe; Qing, Fangzhu; Ni, Yilu; Fan, Yujiang; Tan, Yanfei; Zhang, Xingdong

    2015-03-01

    Porous calcium phosphate ceramics (CaP ceramics) could induce ectopic bone formation which was regulated by various signal molecules. In this work, bone marrow mesenchymal stem cells (MSCs) were cultured on the surface of osteoinductive hydroxyapatite (HA) and biphasic calcium phosphate (BCP) ceramics in comparison with control (culture plate) for up to 14 days to detect the signal molecules which might be affected by the CaP ceramics. Without adding osteogenic factors, MSCs cultured on HA and BCP both expressed higher Runx2, Osterix, collagen type I, osteopontin, bone sialoprotein, and osteocalcin at various stages compared with control, thus confirmed the osteoblastic differentiation of MSCs. Later study demonstrated the messenger RNA level of bone morphogenetic protein 2 (BMP2) and BMP4 were also significantly enhanced by HA and BCP. Furthermore, Smad1, 4, 5, and Dlx5, the main molecules in the BMP/Smads signaling pathway, were upregulated by HA and BCP. Moreover, the higher expression of Smads and BMP2, 4 in BCP over HA, corresponded to the better performance of BCP in stimulating in vitro osteoblastic differentiation of MSCs. This was in accordance with the better osteoinductivity of BCP over HA in vivo. Altogether, these results implied that the CaP ceramics may initiate the osteoblastic differentiation of MSCs by influencing the expression of molecules in BMP/Smads pathway. © 2014 Wiley Periodicals, Inc.

  5. RNA editing differently affects protein-coding genes in D. melanogaster and H. sapiens.

    Science.gov (United States)

    Grassi, Luigi; Leoni, Guido; Tramontano, Anna

    2015-07-14

    When an RNA editing event occurs within a coding sequence it can lead to a different encoded amino acid. The biological significance of these events remains an open question: they can modulate protein functionality, increase the complexity of transcriptomes or arise from a loose specificity of the involved enzymes. We analysed the editing events in coding regions that produce or not a change in the encoded amino acid (nonsynonymous and synonymous events, respectively) in D. melanogaster and in H. sapiens and compared them with the appropriate random models. Interestingly, our results show that the phenomenon has rather different characteristics in the two organisms. For example, we confirm the observation that editing events occur more frequently in non-coding than in coding regions, and report that this effect is much more evident in H. sapiens. Additionally, in this latter organism, editing events tend to affect less conserved residues. The less frequently occurring editing events in Drosophila tend to avoid drastic amino acid changes. Interestingly, we find that, in Drosophila, changes from less frequently used codons to more frequently used ones are favoured, while this is not the case in H. sapiens.

  6. Hydrogen sulfide emissions from a swine building affected by dietary crude protein.

    Science.gov (United States)

    Liu, Shule; Ni, Ji-Qin; Radcliffe, John S; Vonderohe, Caitlin

    2017-12-15

    Hydrogen sulfide (H2S) is a toxic air pollutant at animal facilities; but the understanding of its generation and emission processes has been limited. This paper studied H2S emissions during a complete cycle of wean-finish pigs from a research building, where 12 pig rooms were divided into three groups that were fed with standard feed (control), and 2.1-3.8% (T1) and 4.4-7.8% (T2) reduced dietary crude protein (CP) feed. The group cycle mean H2S emission rates were 4.0 ± 2.9, 4.3 ± 3.2, and 5.4 ± 4.0 g d(-1) AU(-1) (Animal Unit = 500 kg live mass), respectively, for the control, T1, and T2 groups. Emissions of H2S were promoted by 10.0 and 36.7%, respectively, for the T1 and T2 groups (p < 0.001), although large variabilities existed in the emissions from different rooms within the same groups. The enhanced H2S emissions from the T1 and T2 groups were related to the reduced manure pH and were possibly affected through a number of pathways, which could involve volatile fatty acids and nitrogen concentrations, and microbial activities in manure. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Borna disease virus P protein affects neural transmission through interactions with gamma-aminobutyric acid receptor-associated protein.

    Science.gov (United States)

    Peng, Guiqing; Yan, Yan; Zhu, Chengliang; Wang, Shiqun; Yan, Xiaohong; Lu, Lili; Li, Wei; Hu, Jing; Wei, Wei; Mu, Yongxin; Chen, Yanni; Feng, Yong; Gong, Rui; Wu, Kailang; Zhang, Fengmin; Zhang, Xiaolian; Zhu, Ying; Wu, Jianguo

    2008-12-01

    Borna disease virus (BDV) is one of the infectious agents that causes diseases of the central nervous system in a wide range of vertebrate species and, perhaps, in humans. The phosphoprotein (P) of BDV, an essential cofactor of virus RNA-dependent RNA polymerase, is required for virus replication. In this study, we identified the gamma-aminobutyric acid receptor-associated protein (GABARAP) with functions in neurobiology as one of the viral P protein-interacting cellular factors by using an approach of phage display-based protein-protein interaction analysis. Direct binding between GABARAP and P protein was confirmed by coimmunoprecipitation, protein pull-down, and mammalian two-hybrid analyses. GABARAP originally was identified as a linker between the gamma-aminobutyric acid receptor (GABAR) and the microtubule to regulate receptor trafficking and plays important roles in the regulation of the inhibitory neural transmitter gamma-aminobutyric acid (GABA). We showed that GABARAP colocalizes with P protein in the cells infected with BDV or transfected with the P gene, which resulted in shifting the localization of GABARAP from the cytosol to the nucleus. We further demonstrated that P protein blocks the trafficking of GABAR, a principal GABA-gated ion channel that plays important roles in neural transmission, to the surface of cells infected with BDV or transfected with the P gene. We proposed that during BDV infection, P protein binds to GABARAP, shifts the distribution of GABARAP from the cytoplasm to the nucleus, and disrupts the trafficking of GABARs to the cell membranes, which may result in the inhibition of GABA-induced currents and in the enhancement of hyperactivity and anxiety.

  8. Minor milk constituents are affected by protein concentration and forage digestibility in the feed ration.

    Science.gov (United States)

    Larsen, Torben; Alstrup, Lene; Weisbjerg, Martin Riis

    2016-02-01

    The present study was conducted in order to investigate if selected minor milk components would be indicative for the nutritional situation of the cow. Forty-eight dairy cows were offered a high digestible ration vs. a lower digestible ration combined with 2 protein levels in a 4 × 4 Latin square design. Milk glucose, glucose-6-phosphate, cholesterol, triacylglycerides (TAG), uric acid and β-hydroxybutyrate (BHBA) were measured and correlated mutually and towards other milking parameters (yield, h since last milking, days in milk (DIM), urea, etc). The variation range of the suggested variables were broad, a fact that may support their utilisation as predictive parameters. The content of milk metabolites was significantly affected by the change in rations as milk glucose, glucose-6-phosphate, uric acid, and the ratio cholesterol: triacylglycerides increased with higher energy intake while BHBA and TAG decreased. The content of some of the milk metabolites changed during 24 h day/night periods: BHBA, cholesterol, uric acid and TAG increased whereas free glucose decreased in the night period. Certain associations between milk metabolites and calculated energy parameters like ECM, body condition score (BCS), and body weight gain were found, however, these associations were to some extent explained by an interaction with DIM, just as changes in milk metabolites during a 24 h period seems to interfere. It is concluded that the practical use of the suggested milk variables should be based on more than one metabolite and that stage of lactation and possibly time of the day where the milk is collected should be incorporated in predictive models.

  9. Quantity and functionality of protein fractions in chicken breast fillets affected by white striping.

    Science.gov (United States)

    Mudalal, S; Babini, E; Cavani, C; Petracci, M

    2014-08-01

    Recently, white striations parallel to muscle fibers direction have been observed on the surface of chicken breast, which could be ascribed to intensive growth selection. The aim of this study was to evaluate the effect of white striping on chemical composition with special emphasis on myofibrillar and sarcoplasmic protein fractions that are relevant to the processing features of chicken breast meat. During this study, a total of 12 pectoralis major muscles from both normal and white striped fillets were used to evaluate chemical composition, protein solubility (sarcoplasmic, myofibrillar, and total protein solubility), protein quantity (sarcoplasmic, myofibrillar, and stromal proteins), water holding capacity, and protein profile by SDS-PAGE analysis. White-striped fillets exhibited a higher percentage of moisture (75.4 vs. 73.8%; P content of protein (18.7 vs. 22.8%; P content and solubility as well as an increase in cooking loss (33.7 vs. 27.4%; P proteins corresponding to actin (42 kDa); LC1, slow-twitch light chain myosin (27.5 kDa); and LC3, fast-twitch light chain myosin (16 kDa), and almost all sarcoplasmic proteins were lower than normal. In conclusion, the findings of this study revealed that chicken breast meat with white striping defect had different chemical composition (more fat and less protein) and protein quality and quantity (low content of myofibrillar proteins and high content of stromal proteins) with respect to normal meat. Furthermore, white striped fillets had lower protein functionality (higher cooking loss). All the former changes indicate that white striping has great impact on quality characteristics of chicken breast meat. © Poultry Science Association Inc.

  10. Energy and Protein Supplementation Does Not Affect Protein and Amino Acid Kinetics or Pregnancy Outcomes in Underweight Indian Women.

    Science.gov (United States)

    Dwarkanath, Pratibha; Hsu, Jean W; Tang, Grace J; Anand, Pauline; Thomas, Tinku; Thomas, Annamma; Sheela, C N; Kurpad, Anura V; Jahoor, Farook

    2016-02-01

    In India, the prevalence of low birth weight is high in women with a low body mass index (BMI), suggesting that underweight women are not capable of providing adequate energy and protein for fetal growth. Furthermore, as pregnancy progresses, there is increased need to provide methyl groups for methylation reactions associated with the synthesis of new proteins and, unlike normal-BMI American women, low-BMI Indian women are unable to increase methionine transmethylation and remethylation rates as pregnancy progresses from trimester 1 to 3. This also negatively influences birth weight. The aim was to determine the effect of dietary supplementation with energy and protein from 12 ± 1 wk of gestation to time of delivery compared with no supplement on pregnancy outcomes, protein kinetics, and the fluxes of the methyl group donors serine and glycine. Protein kinetics and serine and glycine fluxes were measured by using standard stable isotope tracer methods in the fasting and postprandial states in 24 pregnant women aged 22.9 ± 0.7 y with low BMIs [BMI (in kg/m(2)) ≤18.5] at 12 ± 1 wk (trimester 1) and 30 ± 1 wk (trimester 3) of gestation. After the first measurement, subjects were randomly assigned to either receive the supplement (300 kcal/d, 15 g protein/d) or no supplement. Supplementation had no significant effect on any variable of pregnancy outcome, and except for fasting state decreases in leucine flux (125 ± 7.14 compared with 113 ± 5.06 μmol ⋅ kg(-1) ⋅ h(-1); P = 0.04) and nonoxidative disposal (110 ± 6.97 compared with 101 ± 3.69 μmol ⋅ kg(-1) ⋅ h(-1); P = 0.02) from trimesters 1 to 3, it had no effect on any other leucine kinetic variable or urea, glycine, and serine fluxes. We conclude that in Indian women with a low BMI, supplementation with energy and protein from week 12 of pregnancy to time of delivery does not improve pregnancy outcome, whole-body protein kinetics, or serine and glycine fluxes. © 2016 American Society for Nutrition.

  11. Acute moderate elevation of TNF-{alpha} does not affect systemic and skeletal muscle protein turnover in healthy humans

    DEFF Research Database (Denmark)

    Petersen, Anne Marie; Plomgaard, Peter; Fischer, Christian P

    2009-01-01

    Context: Skeletal muscle wasting has been associated with elevations in circulating inflammatory cytokines, in particular TNF-alpha. Objective: In this study, we investigated whether TNF-alpha affects human systemic and skeletal muscle protein turnover, via a 4 hours recombinant human TNF...... of either rhTNF-alpha (700 ng.m(-2).h(-1)) or 20% human albumin (Control) which was the vehicle of rhTNF-alpha. Systemic and skeletal muscle protein turnover were estimated by a combination of tracer dilution methodology (primed continuous infusion of L-[ring-(2)H5]phenylalanine and L-[(15)N...... with the phenylalanine 3-compartment model showed similar muscle synthesis, breakdown and net muscle degradation after 2 hours basal and after 4 hours Control or rhTNF-alpha infusion. Conclusion: This study is the first to show in humans that TNF-alpha does not affect systemic and skeletal muscle protein turnover, when...

  12. Factors Affecting the Binding of a Recombinant Heavy Metal-Binding Domain (CXXC motif Protein to Heavy Metals

    Directory of Open Access Journals (Sweden)

    Kamala Boonyodying

    2012-06-01

    Full Text Available A number of heavy metal-binding proteins have been used to study bioremediation. CXXC motif, a metal binding domain containing Cys-X-X-Cys motif, has been identified in various organisms. These proteins are capable of binding various types of heavy metals. In this study, heavy metal binding domain (CXXC motif recombinant protein encoded from mcsA gene of S. aureus were cloned and overexpressed in Escherichia coli. The factors involved in the metal-binding activity were determined in order to analyze the potential of recombinant protein for bioremediation. A recombinant protein can be bound to Cd2+, Co2+, Cu2+ and Zn2+. The thermal stability of a recombinant protein was tested, and the results showed that the metal binding activity to Cu2+ and Zn2+ still exist after treating the protein at 85ºC for 30 min. The temperature and pH that affected the metal binding activity was tested and the results showed that recombinant protein was still bound to Cu2+ at 65ºC, whereas a pH of 3-7 did not affect the metal binding E. coli harboring a pRset with a heavy metal-binding domain CXXC motif increased the resistance of heavy metals against CuCl2 and CdCl2. This study shows that metal binding domain (CXXC motif recombinant protein can be effectively bound to various types of heavy metals and may be used as a potential tool for studying bioremediation.

  13. Quantity of dietary protein intake, but not pattern of intake, affects net protein balance primarily through differences in protein synthesis in older adults.

    Science.gov (United States)

    Kim, Il-Young; Schutzler, Scott; Schrader, Amy; Spencer, Horace; Kortebein, Patrick; Deutz, Nicolaas E P; Wolfe, Robert R; Ferrando, Arny A

    2015-01-01

    To examine whole body protein turnover and muscle protein fractional synthesis rate (MPS) following ingestions of protein in mixed meals at two doses of protein and two intake patterns, 20 healthy older adult subjects (52-75 yr) participated in one of four groups in a randomized clinical trial: a level of protein intake of 0.8 g (1RDA) or 1.5 g·kg(-1)·day(-1) (∼2RDA) with uneven (U: 15/20/65%) or even distribution (E: 33/33/33%) patterns of intake for breakfast, lunch, and dinner over the day (1RDA-U, 1RDA-E, 2RDA-U, or 2RDA-E). Subjects were studied with primed continuous infusions of L-[(2)H5]phenylalanine and L-[(2)H2]tyrosine on day 4 following 3 days of diet habituation. Whole body protein kinetics [protein synthesis (PS), breakdown, and net balance (NB)] were expressed as changes from the fasted to the fed states. Positive NB was achieved at both protein levels, but NB was greater in 2RDA vs. 1RDA (94.8 ± 6.0 vs. 58.9 ± 4.9 g protein/750 min; P = 0.0001), without effects of distribution on NB. The greater NB was due to the higher PS with 2RDA vs. 1RDA (15.4 ± 4.8 vs. -18.0 ± 8.4 g protein/750 min; P = 0.0018). Consistent with PS, MPS was greater with 2RDA vs. 1RDA, regardless of distribution patterns. In conclusion, whole body net protein balance was greater with protein intake above recommended dietary allowance (0.8 g protein·kg(-1)·day(-1)) in the context of mixed meals, without demonstrated effects of protein intake pattern, primarily through higher rates of protein synthesis at whole body and muscle levels. Copyright © 2015 the American Physiological Society.

  14. Narciclasine, a potential allelochemical, affects subcellular trafficking of auxin transporter proteins and actin cytoskeleton dynamics in Arabidopsis roots.

    Science.gov (United States)

    Hu, Yanfeng; Na, Xiaofan; Li, Jiaolong; Yang, Lijing; You, Jia; Liang, Xiaolei; Wang, Jianfeng; Peng, Liang; Bi, Yurong

    2015-12-01

    The present study documented the action of a potential allelochemical, narciclasine, on auxin transport in Arabidopsis by mainly affecting subcellular trafficking of PIN and AUX1 proteins and through interfering actin cytoskeletal organization. Narciclasine (NCS), an Amaryllidaceae alkaloid isolated from Narcissus tazetta bulbs, has potential allelopathic activity and affects auxin transport. However, little is known about the cellular mechanism of this inhibitory effect of NCS on auxin transport. The present study characterizes the effects of NCS at the cellular level using transgenic Arabidopsis plants harboring the promoters of PIN, in combination with PIN-GFP proteins or AUX1-YFP fusions. NCS treatment caused significant reduction in the abundance of PIN and AUX1 proteins at the plasma membrane (PM). Analysis of the subcellular distribution of PIN and AUX1 proteins in roots revealed that NCS induced the intracellular accumulation of auxin transporters, including PIN2, PIN3, PIN4, PIN7 and AUX1. However, other PM proteins, such as PIP2, BRI1, and low temperature inducible protein 6b (LTI6b), were insensitive to NCS treatment. NCS-induced PIN2 compartments were further defined using endocytic tracer FM 4-64 labeled early endosomes and suggested that this compound affects the endocytosis trafficking of PIN proteins. Furthermore, pharmacological analysis indicated that the brefeldin A (BFA)-insensitive pathway is employed in the cellular effects of NCS on PIN2 trafficking. Although NCS did not alter actin dynamics in vitro, it resulted in the depolymerization of the actin cytoskeleton in vivo. This disruption of actin filaments by NCS subsequently influences the actin-based vesicle motility. Hence, the elucidation of the specific role of NCS is useful for further understanding the mechanisms of allelopathy at the phytohormone levels.

  15. The oral immunogenicity of BioProtein, a bacterial single-cell protein, is affected by its particulate nature.

    Science.gov (United States)

    Christensen, Hanne R; Larsen, Linea C; Frøkiaer, Hanne

    2003-07-01

    The bacterial single-cell protein BioProtein (BP; Norferm Danmark, Odense, Denmark), produced by fermentation of natural gas with methanotrophic bacteria, is a potential protein source for man and animals. For human consumption, removal of the nucleic acid is necessary. Preliminary studies have shown that ingested BP induces a specific immune response. The objective of the present study was to characterize the type of response, its development over time and product-related causative factors. Mice were fed with diets containing 60 g nucleic acid-reduced BP/kg, 240 g nucleic acid-reduced BP/kg, 240 g untreated BP (basic BP)/kg or 240 g casein/kg (control). In another study, mice were fed 240 g basic BP/kg, whole cell-free BP-culture homogenate or control diet. The immune response was monitored using an ELISA for BP-specific immunoglobulin in blood and BP-specific immunoglobulin A in blood and saliva. Ingested BP induced a steady specific mucosal and systemic immune response, characterized by a dose-dependent production of immunoglobulin and immunoglobulin A in blood and immunoglobulin A in saliva. Basic BP and nucleic acid-reduced BP induced identical responses. However, feeding mice BP-culture homogenate induced immunoglobulin A in saliva but there was no systemic response. The antibodies from BP-fed mice cross-reacted with BP-culture homogenate revealing the presence of the same antigenic components in the two products despite the different oral immunogenicity. Thus, ingestion of BP induces a persistent mucosal and systemic immune response of which the systemic response can be avoided by ingesting a BP preparation free of whole cells. This indicates the importance of the non-particulate constitution of single-cell protein products intended for human or animal consumption.

  16. Denaturation and Oxidative Stability of Hemp Seed (Cannabis sativa L.) Protein Isolate as Affected by Heat Treatment.

    Science.gov (United States)

    Raikos, Vassilios; Duthie, Garry; Ranawana, Viren

    2015-09-01

    The present study investigated the impact of heat treatments on the denaturation and oxidative stability of hemp seed protein during simulated gastrointestinal digestion (GID). Heat-denatured hemp protein isolate (HPI) solutions were prepared by heating HPI (2 mg/ml, pH 6.8) to 40, 60, 80 and 100 °C for 10 min. Heat-induced denaturation of the protein isolates was monitored by polyacrylamide gel electrophoresis. Heating HPI at temperatures above 80 °C significantly reduced solubility and led to the formation of large protein aggregates. The isolates were then subjected to in vitro GID and the oxidative stability of the generated peptides was investigated. Heating did not significantly affect the formation of oxidation products during GID. The results suggest that heat treatments should ideally remain below 80 °C if heat stability and solubility of HPI are to be preserved.

  17. Local NSAID infusion does not affect protein synthesis and gene expression in human muscle after eccentric exercise

    DEFF Research Database (Denmark)

    Mikkelsen, U R; Schjerling, P.; Langberg, Henning

    2011-01-01

    models, and inhibit the exercise-induced satellite cell proliferation and protein synthesis in humans. However, the cellular mechanisms eliciting these responses remain unknown. Eight healthy male volunteers performed 200 maximal eccentric contractions with each leg. To block prostaglandin synthesis......Unaccustomed exercise leads to satellite cell proliferation and increased skeletal muscle protein turnover. Several growth factors and cytokines may be involved in the adaptive responses. Non-steroidal anti-inflammatory drugs (NSAIDs) negatively affect muscle regeneration and adaptation in animal...... locally in the skeletal muscle, indomethacin (NSAID) was infused for 7.5 h via microdialysis catheters into m. vastus lateralis of one leg. Protein synthesis was determined by the incorporation of 1,2-(13) C(2) leucine into muscle protein from 24 to 28 h post-exercise. Furthermore, mRNA expression...

  18. How does domain replacement affect fibril formation of the rabbit/human prion proteins.

    Directory of Open Access Journals (Sweden)

    Xu Yan

    Full Text Available It is known that in vivo human prion protein (PrP have the tendency to form fibril deposits and are associated with infectious fatal prion diseases, while the rabbit PrP does not readily form fibrils and is unlikely to cause prion diseases. Although we have previously demonstrated that amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and macromolecular crowding has different effects on fibril formation of the rabbit/human PrPs, we do not know which domains of PrPs cause such differences. In this study, we have constructed two PrP chimeras, rabbit chimera and human chimera, and investigated how domain replacement affects fibril formation of the rabbit/human PrPs.As revealed by thioflavin T binding assays and Sarkosyl-soluble SDS-PAGE, the presence of a strong crowding agent dramatically promotes fibril formation of both chimeras. As evidenced by circular dichroism, Fourier transform infrared spectroscopy, and proteinase K digestion assays, amyloid fibrils formed by human chimera have secondary structures and proteinase K-resistant features similar to those formed by the human PrP. However, amyloid fibrils formed by rabbit chimera have proteinase K-resistant features and secondary structures in crowded physiological environments different from those formed by the rabbit PrP, and secondary structures in dilute solutions similar to the rabbit PrP. The results from transmission electron microscopy show that macromolecular crowding caused human chimera but not rabbit chimera to form short fibrils and non-fibrillar particles.We demonstrate for the first time that the domains beyond PrP-H2H3 (β-strand 1, α-helix 1, and β-strand 2 have a remarkable effect on fibrillization of the rabbit PrP but almost no effect on the human PrP. Our findings can help to explain why amyloid fibrils formed by the rabbit PrP and the human PrP have different secondary structures and why macromolecular crowding has different

  19. The interaction between endogenous 30S ribosomal subunit protein S11 and Cucumber mosaic virus LS2b protein affects viral replication, infection and gene silencing suppressor activity.

    Directory of Open Access Journals (Sweden)

    Ruilin Wang

    Full Text Available Cucumber mosaic virus (CMV is a model virus for plant-virus protein interaction and mechanism research because of its wide distribution, high-level of replication and simple genome structure. The 2b protein is a multifunctional protein encoded by CMV that suppresses RNA silencing-based antiviral defense and contributes to CMV virulence in host plants. In this report, 12 host proteins were identified as CMV LS2b binding partners using the yeast two-hybrid screen system from the Arabidopsis thaliana cDNA library. Among the host proteins, 30S ribosomal subunit protein S11 (RPS11 was selected for further studies. The interaction between LS2b and full-length RPS11 was confirmed using the yeast two-hybrid system. Bimolecular fluorescence complementation (BIFC assays observed by confocal laser microscopy and Glutathione S-transferase (GST pull-down assays were used to verify the interaction between endogenous NbRPS11 and viral CMVLS2b both in vivo and in vitro. TRV-based gene silencing vector was used to knockdown NbRPS11 transcription, and immunoblot analysis revealed a decline in infectious viral RNA replication and a decrease in CMV infection in RPS11 down-regulated Nicotiana benthamiana plants. Thus, the knockdown of RPS11 likely inhibited CMV replication and accumulation. The gene silencing suppressor activity of CMV2b protein was reduced by the RPS11 knockdown. This study demonstrated that the function of viral LS2b protein was remarkably affected by the interaction with host RPS11 protein.

  20. Components of the E. coli envelope are affected by and can react to protein over-production in the cytoplasm

    Directory of Open Access Journals (Sweden)

    Villa Riccardo

    2009-06-01

    of LamB, OmpF, OmpA and TolC. Under all tested conditions, the lipopolysaccharide was not affected, suggesting that a specific rather than a generalized rearrangement of the envelope was induced. Conclusion Taking together physiological and biochemical evidence, our work indicates that the E. coli envelope can sense protein over-expression in the cytoplasm and react by modulating the abundance of some membrane proteins, with possible consequences on the membrane traffic of small solutes, i.e. nutrients, drugs and metabolites. Such a response seems to be independent on the nature of the protein being over-expressed. On the other hand both our data reported herein and previous results indicate that membrane lipids may act as a second stress sensor responsive to the aggregation state of the recombinant protein and further contribute to changes in cellular exchanges with the environment.

  1. Sensory aroma characteristics of alcalase hydrolyzed rice bran protein concentrate as affected by spray drying and sugar addition.

    Science.gov (United States)

    Arsa, Supeeraya; Theerakulkait, Chockchai

    2015-08-01

    The sensory aroma characteristics of alcalase hydrolyzed rice bran protein concentrate as affected by spray drying and sugar addition were investigated. Rice bran protein concentrate (RBPC) was hydrolyzed by alcalase. Sucrose, glucose or fructose was added to the liquid rice bran protein hydrolysate (LRBPH) and subsequently spray dried. The sensory aroma intensities of the hydrolysates were evaluated. Results showed that after spray drying, the rice bran protein concentrate powder (RBPC-P) had higher sweet and cocoa-like aroma intensities than RBPC (p ≤ 0.05) and hydrolyzed rice bran protein powder (HRBPP) had higher milk powder-like aroma intensities than LRBPH (p ≤ 0.05). The sweet, cocoa-like and milk powder-like aroma intensities in hydrolyzed rice bran protein powder with fructose addition (HRBPP-F) were significantly higher (p ≤ 0.05) than those of hydrolyzed rice bran protein powder with sucrose or glucose addition (HRBPP-S or HRBPP-G). HRBPP-F had the highest overall aroma liking score. These results also indicate that spray drying and sugar addition could improve the sensory aroma characteristics of alcalase hydrolyzed RBPC.

  2. Three regulators of G protein signaling differentially affect mating, morphology and virulence in the smut fungus Ustilago maydis.

    Science.gov (United States)

    Moretti, Marino; Wang, Lei; Grognet, Pierre; Lanver, Daniel; Link, Hannes; Kahmann, Regine

    2017-09-01

    Regulators of G protein signaling (RGS) proteins modulate heterotrimeric G protein signaling negatively. To broaden an understanding of the roles of RGS proteins in fungal pathogens, we functionally characterized the three RGS protein-encoding genes (rgs1, rgs2 and rgs3) in the phytopathogenic fungus Ustilago maydis. It was found that RGS proteins played distinct roles in the regulation of development and virulence. rgs1 had a minor role in virulence when deleted in a solopathogenic strain. In crosses, rgs1 was dispensable for mating and filamentation, but was required for teliospore production. Haploid rgs2 mutants were affected in cell morphology, growth, mating and were unable to cause disease symptoms in crosses. However, virulence was unaffected when rgs2 was deleted in a solopathogenic strain, suggesting an exclusive involvement in pre-fusion events. These rgs2 phenotypes are likely connected to elevated intracellular cAMP levels. rgs3 mutants were severely attenuated in mating, in their response to pheromone, virulence and formation of mature teliospores. The mating defect could be traced back to reduced expression of the transcription factor rop1. It was speculated that the distinct roles of the three U. maydis RGS proteins were achieved by direct modulation of the Gα subunit-activated signaling pathways as well as through Gα-independent functions. © 2017 John Wiley & Sons Ltd.

  3. Emulsifying and gelling properties of weakfish myofibrillar proteins as affected by squid mantle myofibrillar proteins in a model system

    Directory of Open Access Journals (Sweden)

    Daniela Mariel Suarez

    2014-03-01

    Full Text Available The aim of the present work was to investigate the physicochemical, biochemical and functional characteristics of both the myofibrils (MF and actomyosin (AM of squid mantle (Illex argentinus and weakfish (Cynoscion guatucupa muscles, and evaluate the influence of the addition of myofibrilar proteins from the squid mantle on the physicochemical and functional properties of those of the weakfish. After extraction, purification and characterization of the MF and AM of both species, emulsions of each protein fraction from each muscle were formulated. Mixtures of the MF or AM of both species were also analyzed. The emulsifying properties were monitoring using the Emulsifying Activity Index (EAI and Emulsion Stability (ES. In addition, gel pastes were formulated from the squid mantle, weakfish muscle and the mixture of both species, and the following functional properties of the gels assessed: water holding capacity, colour, textural profile analysis (TPA (hardness, elasticity, cohesiveness, gumminess and gel strength. The EAI values of emulsions formulated with the MF of the mantle were significantly (p<0.05 higher than those formulated from those of weakfish. The incorporation of squid MF in the mixture increased the EAI values. Conversely, the highest ES values were obtained with weakfish MF, and the incorporation of MF weakfish in the mixture increased the ES values. Similar EAI and ES behaviours were observed for the AM of the corresponding species. Irrespective of the thermal treatment, the gel strength of the gelled paste of squid muscle was significantly (p<0.05 lower than that of weakfish muscle and of those obtained with the different mixtures. The behaviours of the expressible moisture (EM from the gelled pastes were similar to those of gel strength. Irrespective of the thermal treatment, the pastes formulated with a high weakfish: mantle ratio showed less water loss. The gelled pastes of squid mantle showed the highest values for whiteness

  4. An addition of sourdough and whey proteins affects the nutritional quality of wholemeal wheat bread.

    Science.gov (United States)

    Kopeć, Aneta; Borczak, Barbara; Pysz, Mirosław; Sikora, Elżbieta; Sikora, Marek; Curic, Duska; Novotni, Dubravka

    2014-01-01

    Bread can be a good source of nutrients as well as non-nutrient compounds. This study was designed to assess the effect of adding of sourdough and whey proteins to wholemeal (WM) bread produced by bake-off technology on chemical composition and bioavailability of proteins, calcium, phosphorus, magnesium and iron content in Wistar rats. Wholemeal breads were baked with using conventional or bake off technology. In breads chemical composition, selected minerals content, amino acid composition were measured. Five week-old Wistar rats (n = 30, male), were randomly divided into fi ve groups and fed with modified AIN-93G diets containing experimental breads. In animal study the nutritional value of breads' proteins and concentration of selected minerals in serum, liver and femoral bone, were measured. The body weight gain, biological value (BV) and net protein utilization (NPU) were significantly higher in rats fed with partially baked frozen wholemeal (PBF WM) bread with sourdough and whey proteins. The level of magnesium was significantly lower in serum of animals fed with the diet containing PBF WM bread with sourdough and whey proteins in comparison to rodents fed with conventional WM bread with sourdough. The content of iron was significantly higher in liver of rats fed with PBF WM with sourdough bread in comparison to the groups fed with conventional WM and conventional WM with sourdough breads. Sourdough addition can be recommended in a production of whole wheat partially baked frozen bread but its use is further more beneficial if it is fermented with whey proteins.

  5. Mining the structural genomics pipeline: identification of protein properties that affect high-throughput experimental analysis.

    Science.gov (United States)

    Goh, Chern-Sing; Lan, Ning; Douglas, Shawn M; Wu, Baolin; Echols, Nathaniel; Smith, Andrew; Milburn, Duncan; Montelione, Gaetano T; Zhao, Hongyu; Gerstein, Mark

    2004-02-06

    Structural genomics projects represent major undertakings that will change our understanding of proteins. They generate unique datasets that, for the first time, present a standardized view of proteins in terms of their physical and chemical properties. By analyzing these datasets here, we are able to discover correlations between a protein's characteristics and its progress through each stage of the structural genomics pipeline, from cloning, expression, purification, and ultimately to structural determination. First, we use tree-based analyses (decision trees and random forest algorithms) to discover the most significant protein features that influence a protein's amenability to high-throughput experimentation. Based on this, we identify potential bottlenecks in various stages of the structural genomics process through specialized "pipeline schematics". We find that the properties of a protein that are most significant are: (i.) whether it is conserved across many organisms; (ii). the percentage composition of charged residues; (iii). the occurrence of hydrophobic patches; (iv). the number of binding partners it has; and (v). its length. Conversely, a number of other properties that might have been thought to be important, such as nuclear localization signals, are not significant. Thus, using our tree-based analyses, we are able to identify combinations of features that best differentiate the small group of proteins for which a structure has been determined from all the currently selected targets. This information may prove useful in optimizing high-throughput experimentation. Further information is available from http://mining.nesg.org/.

  6. Phosphatidylcholine affects the role of the sorting and assembly machinery in the biogenesis of mitochondrial β-barrel proteins.

    Science.gov (United States)

    Schuler, Max-Hinderk; Di Bartolomeo, Francesca; Böttinger, Lena; Horvath, Susanne E; Wenz, Lena-Sophie; Daum, Günther; Becker, Thomas

    2015-10-30

    Two protein translocases drive the import of β-barrel precursor proteins into the mitochondrial outer membrane: The translocase of the outer membrane (TOM complex) promotes transport of the precursor to the intermembrane space, whereas the sorting and assembly machinery (SAM complex) mediates subsequent folding of the β-barrel and its integration into the target membrane. The non-bilayer-forming phospholipids phosphatidylethanolamine (PE) and cardiolipin (CL) are required for the biogenesis of β-barrel proteins. Whether bilayer-forming phospholipids such as phosphatidylcholine (PC), the most abundant phospholipid of the mitochondrial outer membrane, play a role in the import of β-barrel precursors is unclear. In this study, we show that PC is required for stability and function of the SAM complex during the biogenesis of β-barrel proteins. PC further promotes the SAM-dependent assembly of the TOM complex, indicating a general role of PC for the function of the SAM complex. In contrast to PE-deficient mitochondria precursor accumulation at the TOM complex is not affected by depletion of PC. We conclude that PC and PE affect the function of distinct protein translocases in mitochondrial β-barrel biogenesis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Contraction mode and whey protein intake affect the synthesis rate of intramuscular connective tissue

    DEFF Research Database (Denmark)

    Holm, Lars; Klejs Rahbek, Stine; Farup, Jean

    2016-01-01

    IntroductionIn this study we investigated the impact of whey protein hydrolysate and maltodextrin (WPH) intake on intramuscular connective tissue (IMCT) protein fractional synthesis rate (FSR) after maximal shortening and lengthening contractions. Methods Twenty young men were randomized to receive...... either WPH or maltodextrin [carbohydrate (CHO)] immediately after completion of unilateral shortening and lengthening knee extensions. Ring-13C6-phenylalanine was infused, and muscle biopsies were obtained. IMCT protein FSR was measured at 1–5, as well as 1–3 and 3–5 hours after contractions and nutrient...

  8. Dietary protein content affects evolution for body size, body fat and viability in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Kristensen, Torsten N; Overgaard, Johannes; Loeschcke, Volker

    2011-01-01

    composition and developmental success in Drosophila melanogaster. We reared flies on either a standard diet or a protein-enriched diet for 17 generations before testing them on both diet types. Flies from lines selected on protein-rich diet produced phenotypes with higher total body mass and relative lipid...... content when compared with those selected on a standard diet, irrespective of which of the two diets they were tested on. However, selection on protein-rich diet incurred a cost as flies reared on this diet had markedly lower developmental success in terms of egg-to-adult viability on both medium types...

  9. Homogenization Pressure and Temperature Affect Protein Partitioning and Oxidative Stability of Emulsions

    DEFF Research Database (Denmark)

    Horn, Anna Frisenfeldt; Barouh, Nathalie; Nielsen, Nina Skall

    2013-01-01

    it decreased the oxidative stability of emulsions with α-lactalbumin and β-lactoglobulin. For both types of emulsions the partitioning of proteins between the interface and the aqueous phase appeared to be important for the oxidative stability. The effect of pre-heating the aqueous phase with the milk proteins......The oxidative stability of 10 % fish oil-in-water emulsions was investigated for emulsions prepared under different homogenization conditions. Homogenization was conducted at two different pressures (5 or 22.5 MPa), and at two different temperatures (22 and 72 °C). Milk proteins were used...

  10. Properties of sweetened Indian yogurt (mishti dohi) as affected by added tryptic whey protein hydrolysate

    National Research Council Canada - National Science Library

    Chatterjee, Alok; Kanawjia, S K; Khetra, Yogesh

    2016-01-01

    Utilization of Indian sweetened yogurt (colloquially termed as Mishti Dohi), as vehicle for ACE inhibition and antioxidant activity, by added tryptic whey protein hydrolysate (TWPH) (@ 1, 2, 3 % v/milk), was attempted. Yogurt with 3...

  11. Quantity and functionality of protein fractions in chicken breast fillets affected by white striping

    National Research Council Canada - National Science Library

    Mudalal, S; Babini, E; Cavani, C; Petracci, M

    2014-01-01

    ... features of chicken breast meat. During this study, a total of 12 pectoralis major muscles from both normal and white striped fillets were used to evaluate chemical composition, protein solubility...

  12. Interactions Affected by Arginine Methylation in the Yeast Protein–Protein Interaction Network*

    Science.gov (United States)

    Erce, Melissa A.; Abeygunawardena, Dhanushi; Low, Jason K. K.; Hart-Smith, Gene; Wilkins, Marc R.

    2013-01-01

    Protein–protein interactions can be modulated by the methylation of arginine residues. As a means of testing this, we recently described a conditional two-hybrid system, based on the bacterial adenylate cyclase (BACTH) system. Here, we have used this conditional two-hybrid system to explore the effect of arginine methylation in modulating protein–protein interactions in a subset of the Saccharomyces cerevisiae arginine methylproteome network. Interactions between the yeast hub protein Npl3 and yeast proteins Air2, Ded1, Gbp2, Snp1, and Yra1 were first validated in the absence of methylation. The major yeast arginine methyltransferase Hmt1 was subsequently included in the conditional two-hybrid assay, initially to determine the degree of methylation that occurs. Proteins Snp1 and Yra1 were confirmed as Hmt1 substrates, with five and two novel arginine methylation sites mapped by ETD LC-MS/MS on these proteins, respectively. Proteins Ded1 and Gbp2, previously predicted but not confirmed as substrates of Hmt1, were also found to be methylated with five and seven sites mapped respectively. Air2 was found to be a novel substrate of Hmt1 with two sites mapped. Finally, we investigated the interactions of Npl3 with the five interaction partners in the presence of active Hmt1 and in the presence of Hmt1 with a G68R inactivation mutation. We found that the interaction between Npl3 and Air2, and Npl3 and Ded1, were significantly increased in the presence of active Hmt1; the interaction of Npl3 and Snp1 showed a similar degree of increase in interaction but this was not statistically significant. The interactions of Npl3 and Gbp2, along with Npl3 and Yra1, were not significantly increased or decreased by methylation. We conclude that methylarginine may be a widespread means by which the interactions of proteins are modulated. PMID:23918811

  13. Overexpression of a stress-responsive U-box protein gene VaPUB affects the accumulation of resistance related proteins in Vitis vinifera 'Thompson Seedless'.

    Science.gov (United States)

    Jiao, Li; Zhang, Yali; Lu, Jiang

    2017-03-01

    Many U-box proteins have been identified and characterized as important factors against environmental stresses such as chilling, heat, salinity and pathogen attack in plant. Our previous research reported the cloning of a novel U-box protein gene VaPUB from Vitis amurensis 'Zuoshanyi' grape and suggested a function of it in related to cold stress in the model plant Arabidopsis system. In this study, the role of VaPUB in response to biotic and abiotic stress was further analyzed in the homologous grapevine system by studying the transcript regulation and the protein accumulation in VaPUB transgenic vines. The expression analysis assay shown that VaPUB was significantly up-regulated 6 h after cold treatment and as early as 2 h post inoculation with Plasmopara viticola, a pathogen causing downy mildew disease in grapevine. Over-expressing VaPUB in V. Vinifera 'Thompson Seedless' affected the microstructure of leaves. The proteome assay shown that the accumulation of pathogenesis-related protein PR10 and many proteins involved in carbon and energy metabolism, oxidation reaction and protein metabolism were significantly altered in transgenic vines. In comparison with wild type plants, the expression level of PR10 family genes was significantly decreased in VaPUB transgenic vines under P. viticola treatment or cold stress. Results from this study showed that the U-box protein gene PUB quickly responded to both biotic stress and abiotic stress and significantly influenced the accumulation of resistance related proteins in grapevine. Copyright © 2016. Published by Elsevier Masson SAS.

  14. Human Milk Analyser shows that the lactation period affects protein levels in preterm breastmilk.

    Science.gov (United States)

    Kreissl, Alexandra; Zwiauer, Valentina; Repa, Andreas; Binder, Christoph; Thanhaeuser, Margarita; Jilma, Bernd; Berger, Angelika; Haiden, Nadja

    2016-06-01

    This study measured the composition of preterm human breastmilk, particularly the protein content, with the MIRIS Human Milk Analyser, compared our results with published values and determined the relationship between protein content and lactation period. We analysed 83 samples of 24-hour pooled human milk from 76 mothers who delivered preterm infants weighing under 1500 g at less than 32 weeks of gestational age. The milk's protein, fat and energy were measured by the MIRIS Human Milk Analyser and compared to reference values. The relationship between protein content and lactation period was quantified. On average, the samples contained 1.1 ± 0.37 g (0.2-2.2 g) of protein, 3.2 ± 0.85 g (range 1.1-6.1 g) of fat, 6.6 ± 0.34 g of lactose (5.5-8.0 g) and 60 ± 11 kcal (39-94 kcal) of energy per 100 mL. The wide variations in macronutrient content were not influenced by the gestational age of the infant and the lactation day results from 70 of the mothers correlated inversely with the protein content (p lactation. With adjustments, the MIRIS might provide a helpful tool for individualised fortification. ©2016 Foundation Acta Paediatrica. Published by John Wiley & Sons Ltd.

  15. Implant debris particle size affects serum protein adsorption which may contribute to particle size-based bioreactivity differences

    Science.gov (United States)

    Reddy, Anand; Caicedo, Marco; Samelko, Lauryn; Jacobs, Joshua J; Hallab, Nadim James

    2014-01-01

    Biologic reactivity to orthopedic implant debris mediates long-term clinical performance of total joint arthroplasty implants. However, why some facets of implant debris are more pro-inflammatory remains controversial such as particle size, shape, base material etc. This precludes accurate prediction and optimal design of modern total joint replacements. We hypothesized that debris particle size can influence adsorbed protein film composition and affect subsequent bioreactivity. We measured size-dependent protein film-adsorption, and adsorbed protein film-dependent cytokine release using equal surface areas of different sized cobalt-chromium-alloy (CoCr-alloy) particle and in vitro challenge of human macrophages (THP-1 and human primary). Smaller 5μm vs 70μm sized particles preferentially adsorbed more serum protein in general (p<0.03), where higher molecular weight serum proteins consistent with IgG were identified. Additionally, 5μm CoCr-alloy particles pre-coated with different protein biofilms (IgG vs albumin) resulted in differential cytokine expression where albumin-coated particles induced more TNF-α and IgG-coated particles induced more IL-1β release from human monocyte/macrophages. In these preliminary in vitro studies we demonstrated the capability of equal surface areas of different particle sizes to influence adsorbed protein composition and that adsorbed protein differences on identical particles can translate into complex differences in bioreactivity. Together this suggests adsorbed protein differences on different sized particles of the same material may be a contributing mechanism by which different sized particles induce differences in reactivity. PMID:24941408

  16. Linker length affects expression and bioactivity of the onconase fusion protein in Pichia pastoris.

    Science.gov (United States)

    Yang, G G; Xu, X Y; Ding, Y; Cui, Q Q; Wang, Z; Zhang, Q Y; Shi, S H; Lv, Z Y; Wang, X Y; Zhang, J H; Zhang, R G; Xu, C S

    2015-12-29

    The aim of this study was to analyze the effect of linker length on the expression and biological activity of recombinant protein onconase (ONC) in fusion with human serum albumin (HSA) in Pichia pastoris. Four flexible linkers with different lengths namely Linker L0, L1: (GGGGS)1, L2: (GGGGS)2, and L3:(GGGGS)3 were inserted into the fusion gene and referred to as HSA-n-ONC, where N = 0, 5, 10, or 15. The sequence of the fusion gene HSA-ONC was designed based on the GC content and codon bias in P. pastoris; the signal peptide of albumin was used as the secretion signal. Gene sequences coding for the fusion protein with different linkers were inserted into pPICZα-A to form recombinant plasmids pPICZα-A/HSA-n-ONC, which were then transformed into P. pastoris X-33 for protein expression. Ideal conditions for expression of the fusion proteins were optimized at a small scale, using shake flasks before proceeding to mass production in 10-L fermenters. The recombinant fusion proteins were purified by aqueous two-phase extraction coupled with DEAE anion exchange chromatography, and their cytotoxic effect on the tumor cell was evaluated by the sulforhodamine B assay. The results showed that the expressed amount of fusion proteins had no significant relationship with the length of different linkers and rHSA-0-ONC had no cytotoxic effect on the tumor cells. While rHSA-5-ONC and rHSA-10-ONC had a weak cytotoxic effect, rHSA-15-ONC could kill various tumor cells in vitro. In summary, the biological activity of the fusion protein gradually improved with increasing length of the linker.

  17. Protein acetylation affects acetate metabolism, motility and acid stress response in Escherichia coli.

    Science.gov (United States)

    Castaño-Cerezo, Sara; Bernal, Vicente; Post, Harm; Fuhrer, Tobias; Cappadona, Salvatore; Sánchez-Díaz, Nerea C; Sauer, Uwe; Heck, Albert J R; Altelaar, A F Maarten; Cánovas, Manuel

    2014-11-27

    Although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. To interrogate its functionality, we analyzed the acetylome in Escherichia coli knockout mutants of cobB, the only known sirtuin-like deacetylase, and patZ, the best-known protein acetyltransferase. For four growth conditions, more than 2,000 unique acetylated peptides, belonging to 809 proteins, were identified and differentially quantified. Nearly 65% of these proteins are related to metabolism. The global activity of CobB contributes to the deacetylation of a large number of substrates and has a major impact on physiology. Apart from the regulation of acetyl-CoA synthetase, we found that CobB-controlled acetylation of isocitrate lyase contributes to the fine-tuning of the glyoxylate shunt. Acetylation of the transcription factor RcsB prevents DNA binding, activating flagella biosynthesis and motility, and increases acid stress susceptibility. Surprisingly, deletion of patZ increased acetylation in acetate cultures, which suggests that it regulates the levels of acetylating agents. The results presented offer new insights into functional roles of protein acetylation in metabolic fitness and global cell regulation. © 2014 The Authors. Published under the terms of the CC BY 4.0 license.

  18. The adsorption of α-amylase on barley proteins affects the in vitro digestion of starch in barley flour.

    Science.gov (United States)

    Yu, Wenwen; Zou, Wei; Dhital, Sushil; Wu, Peng; Gidley, Michael J; Fox, Glen P; Gilbert, Robert G

    2018-02-15

    The conversion of barley starch to sugars is a complex enzymic process. Most previous work concerned the biotechnical aspect of in situ barley enzymes. However, the interactions among the macromolecular substrates and their effects on enzymic catalysis has been little examined. Here, we explore the mechanisms whereby interactions of protein and starch in barley flour affect the kinetics of enzymatic hydrolysis of starch in an in vitro system, using digestion rate data and structural analysis by confocal microscopy. The degradation kinetics of both uncooked barley flour and of purified starches are found to be two-step sequential processes. Barley proteins, especially the water-soluble component, are found to retard the digestion of starch degraded by α-amylase: the enzyme binds with water-insoluble protein and with starch granules, leading to reduced starch hydrolysis. These findings are of potential industrial value in both the brewing and food industries. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Disruption of the serine/threonine protein kinase H affects phthiocerol dimycocerosates synthesis in Mycobacterium tuberculosis

    Science.gov (United States)

    Gómez-Velasco, Anaximandro; Bach, Horacio; Rana, Amrita K.; Cox, Liam R.; Bhatt, Apoorva; Besra, Gurdyal S.

    2013-01-01

    Mycobacterium tuberculosis possesses a complex cell wall that is unique and essential for interaction of the pathogen with its human host. Emerging evidence suggests that the biosynthesis of complex cell-wall lipids is mediated by serine/threonine protein kinases (STPKs). Herein, we show, using in vivo radiolabelling, MS and immunostaining analyses, that targeted deletion of one of the STPKs, pknH, attenuates the production of phthiocerol dimycocerosates (PDIMs), a major M. tuberculosis virulence lipid. Comparative protein expression analysis revealed that proteins in the PDIM biosynthetic pathway are differentially expressed in a deleted pknH strain. Furthermore, we analysed the composition of the major lipoglycans, lipoarabinomannan (LAM) and lipomannan (LM), and found a twofold higher LAM/LM ratio in the mutant strain. Thus, we provide experimental evidence that PknH contributes to the production and synthesis of M. tuberculosis cell-wall components. PMID:23412844

  20. Factors affecting the oxidative stability of omega-3 emulsions prepared with milk proteins

    DEFF Research Database (Denmark)

    Horn, Anna Frisenfeldt; Nielsen, Nina Skall; Jacobsen, Charlotte

    Omega-3 fatty acids are prone to lipid oxidation due to their unsaturated nature. In oil-in-water emulsions, lipid oxidation is expected to be initiated at the oil-water interface. The properties of the emulsifier used and the structure at the interface are therefore expected to be of great...... importance for the resulting oxidation. This presentation will give an overview of parameters that are expected to change the properties and structure of milk protein components at the interface of 10% fish oil-in-water emulsions. Results from three different studies will be included. The first study...... compared the effect of two different high pressure homogenizers on oxidation in caseinate and whey protein isolate emulsions. The second study evaluated the effect of homogenization pressure and temperature on emulsions prepared either with whey proteins or a combination of caseinate and β...

  1. Rare codon content affects the solubility of recombinant proteins in a codon bias-adjusted Escherichia coli strain

    Directory of Open Access Journals (Sweden)

    Ceccarelli Eduardo A

    2009-07-01

    Full Text Available Abstract Background The expression of heterologous proteins in Escherichia coli is strongly affected by codon bias. This phenomenon occurs when the codon usage of the mRNA coding for the foreign protein differs from that of the bacterium. The ribosome pauses upon encountering a rare codon and may detach from the mRNA, thereby the yield of protein expression is reduced. Several bacterial strains have been engineered to overcome this effect. However, the increased rate of translation may lead to protein misfolding and insolubilization. In order to prove this assumption, the solubility of several recombinant proteins from plants was studied in a codon bias-adjusted E. coli strain. Results The expression of eight plant proteins in Escherichia coli BL21(DE3-pLysS and BL21(DE3-CodonPlus-pRIL was systematically studied. The CodonPlus strain contains extra copies of the argU, ileY, and leuW tRNA genes, which encode tRNAs that recognize the codons AGA/AGG, AUA and CUA, respectively (RIL codons. The level of expression and solubility of the recombinant proteins were analyzed by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blotting. We found that for all proteins the solubility was at least 25% in the BL21(DE3-pLysS strain. However, when expressed in the BL21(DE3-CodonPlus-pRIL strain, proteins having more than 5% of amino acids coded by RIL codons were localized mainly in the insoluble fraction. Also, their expression caused retarded growth and low cell yield in the codon bias-adjusted strain at all temperatures tested. On the contrary, the solubility of proteins containing less than 5% of amino acids coded by RIL codons remained unchanged in both strains and their expression caused no effect on cell growth. Conclusion Our results show that the expression of heterologous proteins coded by high RIL codon content coding sequences in a codon bias-adjusted strain is detrimental for their solubility. Our data support the

  2. Level of dietary protein intake affects glucose turnover in endurance-trained men

    Directory of Open Access Journals (Sweden)

    Pasiakos Stefan M

    2011-11-01

    Full Text Available Abstract Background To examine the effects of higher-protein diets on endogenous glucose metabolism in healthy, physically active adults, glucose turnover was assessed in five endurance-trained men (age 21.3 ± 0.3 y, VO2peak 70.6 ± 0.1 mL kg-1 min-1 who consumed dietary protein intakes spanning the current dietary reference intakes. Findings Using a randomized, crossover design, volunteers consumed 4 week eucaloric diets providing either a low (0.8 g kg-1 d-1; LP, moderate (1.8 g kg-1 d-1; MP, or high (3.6 g kg-1 d-1; HP level of dietary protein. Glucose turnover (Ra, glucose rate of appearance; and Rd glucose rate of disappearance was assessed under fasted, resting conditions using primed, constant infusions of [6,6-2H2] glucose. Glucose Ra and Rd (mg kg-1 min-1 were higher for MP (2.8 ± 0.1 and 2.7 ± 0.1 compared to HP (2.4 ± 0.1 and 2.3 ± 0.2, P P P > 0.05 between LP (4.6 ± 0.1, MP (4.8 ± 0.1, and HP (4.7 ± 0.1 diets. Conclusions Level of protein consumption influenced resting glucose turnover in endurance athletes in a state of energy balance with a higher rate of turnover noted for a protein intake of 1.8 g kg-1 d-1. Findings suggest that consumption of protein in excess of the recommended dietary allowance but within the current acceptable macronutrient distribution range may contribute to the regulation of blood glucose when carbohydrate intake is reduced by serving as a gluconeogenic substrate in endurance-trained men.

  3. Alcohol binding to the odorant binding protein LUSH: multiple factors affecting binding affinities.

    Science.gov (United States)

    Ader, Lauren; Jones, David N M; Lin, Hai

    2010-07-27

    Density function theory (DFT) calculations have been carried out to investigate the binding of alcohols to the odorant binding protein LUSH from Drosophila melanogaster. LUSH is one of the few proteins known to bind to ethanol at physiologically relevant concentrations and where high-resolution structural information is available for the protein bound to alcohol at these concentrations. The structures of the LUSH-alcohol complexes identify a set of specific hydrogen-bonding interactions as critical for optimal binding of ethanol. A set of truncated models based on the structure of the LUSH-butanol complex were constructed for the wild-type and mutant (T57S, S52A, and T57A) proteins in complexes with a series of n-alcohols and for the apoprotein bound to water and for the ligand-free protein. Using both gas-phase calculations and continuum solvation model calculations, we found that the widely used DFT model, B3LYP, failed to reproduce the experimentally observed trend of increasing binding affinity with the increasing length of the alkyl chain in the alcohol. In contrast, the recently developed M05-2X DFT model successfully reproduced this subtle trend. Analysis of the results indicated that multiple factors contribute to the differences in alcohol binding affinity: the H-bonding with Thr57 and Ser52 (4-5 kcal/mol per H-bond), the desolvation contribution (4-6 kcal/mol for alcohols and 8-10 kcal/mol for water), and the other noncovalent interaction (1.2 kcal/mol per CH(2) group of the alcohol alkyl chain). These results reveal the outstanding potential for using the M05-2X model in calculations of protein-substrate complexes where noncovalent interactions are important.

  4. Consuming a mixed diet enriched with lupin protein beneficially affects plasma lipids in hypercholesterolemic subjects: a randomized controlled trial.

    Science.gov (United States)

    Bähr, Melanie; Fechner, Anita; Kiehntopf, Michael; Jahreis, Gerhard

    2015-02-01

    The objectives of this study were to assess whether 25 g/d lupin protein, integrated into a mixed diet, might affect cardiovascular risk factors and whether l-arginine was responsible for these effects. Seventy-two hypercholesterolemic subjects participated in the randomized, controlled, double-blind three-phase crossover study. They were assigned to three diets with 25 g/d lupin protein (LP), milk protein (MP) or milk protein plus 1.6 g/d arginine (MPA) each for 28 d in a random order interrupted by 6-week washout periods. Lupin protein and the comparator milk protein were incorporated into complex food products (bread, roll, sausage, and vegetarian spread). Arginine was administered via capsules. Sixty-eight subjects were included in final analyses. Compared with MP, LDL cholesterol was significantly lower after LP. Compared with MP and MPA, homocysteine was significantly lower after LP. Compared with baseline, concentrations of total, LDL, and HDL cholesterol significantly decreased after LP and MPA. Triacylglycerols and uric acid significantly decreased after LP. The relative changes in total and LDL cholesterol were significantly greater for subjects with severe hypercholesterolemia (>6.6 mmol/L) than those with moderate hypercholesterolemia (5.2-6.6 mmol/L). The present study showed for the first time that incorporation of 25 g/d of lupin protein into a variety of complex food products lowers total and LDL cholesterol, triacylglycerols, homocysteine, and uric acid in hypercholesterolemic subjects. The hypocholesterolemic effect is stronger in subjects with severe hypercholesterolemia. Arginine might be responsible for some, but not all of the beneficial effects of lupin protein. This trial was registered at http://clinicaltrials.gov (study ID number NCT01598649). Copyright © 2014 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

  5. Dietary heme adversely affects experimental colitis in rats, despite heat-shock protein induction

    NARCIS (Netherlands)

    Schepens, Marloes A. A.; Vink, Carolien; Schonewille, Arjan J.; Dijkstra, Gerard; van der Meer, Roelof; Bovee-Oudenhoven, Ingeborg M. J.

    Objective: Research on dietary modulation of inflammatory bowel disease is in its infancy. Dietary heme, mimicking red meat, is cytotoxic to colonic epithelium and thus may aggravate colitis. Alternatively, heme-induced colonic stress might also result in potential protective heat-shock proteins

  6. Season progression, Ontogenesis and Environment affect Lespedeza cuneata Herbage Condensed Tannin, Fiber and Crude Protein Content

    Science.gov (United States)

    Sericea lespedeza [Lespedeza cuneata (Dumont de Courset) G. Don.; SL] is a perennial, warm-season forage legumes with wide adaptation, freeze tolerance, establishment ease and persistence under grazing. Its condensed tannins (CT) could be useful in ruminant systems, not simply as a protein source, b...

  7. Protein acetylation affects acetate metabolism, motility and acid stress response in Escherichia coli

    NARCIS (Netherlands)

    Castano-Cerezo, Sara|info:eu-repo/dai/nl/371746698; Bernal, Vicente; Post, Harm|info:eu-repo/dai/nl/341667374; Fuhrer, Tobias; Cappadona, Salvatore|info:eu-repo/dai/nl/341539031; Sanchez-Diaz, Nerea C.; Sauer, Uwe; Heck, Albert J. R.|info:eu-repo/dai/nl/105189332; Altelaar, A. F. Maarten|info:eu-repo/dai/nl/304833517; Canovas, Manuel

    2014-01-01

    Although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. To interrogate its functionality, we analyzed the acetylome in Escherichia coli knockout mutants of cobB, the only known sirtuin-like deacetylase, and patZ, the

  8. Protein corona affects the relaxivity and MRI contrast efficiency of magnetic nanoparticles

    NARCIS (Netherlands)

    Amiri, H.; Bordonali, L.; Lascialfari, A.; Wan, S.; Monopoli, M.P.; Lynch, I.; Laurent, S.; Mahmoudi, M.

    2013-01-01

    Magnetic nanoparticles (NPs) are increasingly being considered for use in biomedical applications such as biosensors, imaging contrast agents and drug delivery vehicles. In a biological fluid, proteins associate in a preferential manner with NPs. The small sizes and high curvature angles of NPs

  9. Proteins Play Important Role in Intercellular Adhesion Affecting on Fruit Textural Quality

    DEFF Research Database (Denmark)

    Bahadur Adhikari, Khem; Shomer, Ilan

    2012-01-01

    Fruit textural quality is becoming a major quality parameter for export, postharvest preservation, handling and processing. The main determinant of textural quality is intercellular adhesion (ICA) as attributed by the cell wall (CW) and its components. The importance of CW protein in ICA strength...

  10. Alterations in expression levels of deafness dystonia protein 1 affect mitochondrial morphology

    DEFF Research Database (Denmark)

    Engl, Gertraud; Florian, Stefan; Tranebjærg, Lisbeth

    2012-01-01

    of mammalian mitochondria, we investigated the effects of reduced or elevated DDP1 levels on mitochondrial dynamics and function. Our results show a reduction in the import of ß-barrel proteins into mitochondria from cells overexpressing DDP1. This effect was not observed when the DDON-related mutant form DDP1...

  11. Minor milk constituents are affected by protein concentration and forage digestibility in the feed ration

    DEFF Research Database (Denmark)

    Larsen, Torben; Alstrup, Lene; Weisbjerg, Martin Riis

    2016-01-01

    The present study was conducted in order to investigate if selected minor milk components would be indicative for the nutritional situation of the cow. Forty-eight dairy cows were offered a high digestible ration vs. a lower digestible ration combined with 2 protein levels in a 4 × 4 Latin square...

  12. Dietary protein affects nitrogen excretion and ammonia emission from slurry of growing-finishing pigs.

    NARCIS (Netherlands)

    Canh, T.T.; Aarnink, A.J.A.; Schutte, J.B.; Sutton, A.L.; Langhout, D.J.; Verstegen, M.W.A.

    1998-01-01

    The effects of dietary protein on nitrogen excretion and ammonia emission from slurry of growing–finishing pigs were studied both in vitro and in a pig house. The three diets had similar contents of NE, minerals, vitamins and ileal digestible lysine, methionine cystine, threonine and tryptophan, but

  13. Dietary protein affects nitrogen excretion and ammonia emission from slurry of growing-finishing pigs

    NARCIS (Netherlands)

    Canh, T.T.; Aarnink, A.J.A.; Schutte, J.B.; Sutton, A.; Langhout, D.J.; Verstegen, M.W.A.

    1998-01-01

    The effects of dietary protein on nitrogen excretion and ammonia emission from slurry of growing-finishing pigs were studied both in vitro and in a pig house. The three diets had similar contents of NE, minerals, vitamins and ileal digestible lysine, methionine + cystine, threonine and tryptophan,

  14. The Major Capsid Protein of Herpes Simplex Virus-1 Affects its

    African Journals Online (AJOL)

    rate rose to 85 % compared with virus control. Knocking down VP5 expression abrogated the changes to F-actin that were induced by HSV-1 infection. Conclusion: Interfering with UL19 gene expression inhibits HSV-1 replication efficiently in vitro. The results indicate that the major capsid protein VP5 encoding gene UL19 ...

  15. Odour and ammonia emission from pig manure as affected by dietary crude protein level

    NARCIS (Netherlands)

    Le, P.D.; Aarnink, A.J.A.; Jongbloed, A.W.

    2009-01-01

    The objective of this study was to determine the effects of dietary crude protein (CP) level on odour emission, odour intensity, odour hedonic tone, ammonia and greenhouse gaseous emission from pig manure, and on fresh faeces and manure characteristics. An experiment was conducted with finishing

  16. Prion Protein M129V Polymorphism Affects Retrieval-Related Brain Activity

    Science.gov (United States)

    Buchmann, Andreas; Mondadori, Christian R. A.; Hanggi, Jurgen; Aerni, Amanda; Vrticka, Pascal; Luechinger, Roger; Boesiger, Peter; Hock, Christoph; Nitsch, Roger M.; de Quervain, Dominique J.-F.; Papassotiropoulos, Andreas; Henke, Katharina

    2008-01-01

    The prion protein Met129Val polymorphism has recently been related to human long-term memory with carriers of either the 129[superscript MM] or the 129[superscript MV] genotype recalling 17% more words than 129[superscript VV] carriers at 24 h following learning. Here, we sampled genotype differences in retrieval-related brain activity at 30 min…

  17. Calcium affecting protein expression in longan under simulated acid rain stress.

    Science.gov (United States)

    Pan, Tengfei; Li, Yongyu; Ma, Cuilan; Qiu, Dongliang

    2015-08-01

    Longan (Dimocarpus longana Lour. cv. Wulongling) of uniform one-aged seedlings grown in pots were selected to study specific proteins expressed in leaves under simulated acid rain (SiAR) stress and exogenous Ca(2+) regulation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results showed that there was a protein band specifically expressed under SiAR of pH 2.5 stress for 15 days with its molecular weight of about 23 kD. A 17 kD protein band specifically expressed after SiAR stress 5 days. Compared with pH 2.5, the pH 3.5 of SiAR made a less influence to protein expression. Two-dimensional electrophoresis (2-DE) results showed that six new specific proteins including C4 (20.2 kD pI 6.0), F (24 kD pI 6.35), B3 (22.3 kD pI 6.35), B4 (23.5 kD pI 6.5), C5 (21.8 kD pI 5.6), and C6 (20.2 kD pI 5.6) specifically expressed. C4 always expressed during SiAR stress. F expressed under the stress of pH 2.5 for 15 days and expressed in all pH SiAR stress for 20 days. The expression of proteins including B3, C5, and C6 was related to pH value and stress intensity of SiAR. The expression of B4 resulted from synergistic effects of SiAR and Ca. The expression of G1 (Mr 19.3 kD, pI 4.5), G2 (Mr 17.8 kD, pI 4.65), G3 (Mr 16.6 kD, pI 4.6), and G4 (Mr 14.7 kD, pI 4.4) enhanced under the treatment of 5 mM ethylene glycol tetraacetic acid (EGTA) and 2 mM chlorpromazine (CPZ). These proteins showed antagonistic effects and might be relative to the Ca-calmodulin (Ca-CaM) system of longan in response to SiAR stress.

  18. Human endogenous retrovirus-FRD envelope protein (syncytin 2 expression in normal and trisomy 21-affected placenta

    Directory of Open Access Journals (Sweden)

    Handschuh Karen

    2008-01-01

    Full Text Available Abstract Human trophoblast expresses two fusogenic retroviral envelope proteins, the widely studied syncytin 1, encoded by HERV-W and the recently characterized syncytin 2 encoded by HERV-FRD. Here we studied syncytin 2 in normal and Trisomy 21-affected placenta associated with abnormal trophoblast differentiation. Syncytin 2 immunolocalization was restricted throughout normal pregnancy to some villous cytotrophoblastic cells (CT. During the second trimester of pregnancy, syncytin 2 was immunolocalized in some cuboidal CT in T21 placentas, whereas in normal placentas it was observed in flat CT, extending into their cytoplasmic processes. In vitro, CT isolated from normal placenta fuse and differentiate into syncytiotrophoblast. At the same time, syncytin 2 transcript levels decreased significantly with syncytiotrophoblast formation. In contrast, CT isolated from T21-affected placentas fused and differentiated poorly and no variation in syncytin 2 transcript levels was observed. Syncytin 2 expression illustrates the abnormal trophoblast differentiation observed in placenta of fetal T21-affected pregnancies.

  19. Alpha-phellandrene-induced DNA damage and affect DNA repair protein expression in WEHI-3 murine leukemia cells in vitro.

    Science.gov (United States)

    Lin, Jen-Jyh; Wu, Chih-Chung; Hsu, Shu-Chun; Weng, Shu-Wen; Ma, Yi-Shih; Huang, Yi-Ping; Lin, Jaung-Geng; Chung, Jing-Gung

    2015-11-01

    Although there are few reports regarding α-phellandrene (α-PA), a natural compound from Schinus molle L. essential oil, there is no report to show that α-PA induced DNA damage and affected DNA repair associated protein expression. Herein, we investigated the effects of α-PA on DNA damage and repair associated protein expression in murine leukemia cells. Flow cytometric assay was used to measure the effects of α-PA on total cell viability and the results indicated that α-PA induced cell death. Comet assay and 4,6-diamidino-2-phenylindole dihydrochloride staining were used for measuring DNA damage and condensation, respectively, and the results indicated that α-PA induced DNA damage and condensation in a concentration-dependent manner. DNA gel electrophoresis was used to examine the DNA damage and the results showed that α-PA induced DNA damage in WEHI-3 cells. Western blotting assay was used to measure the changes of DNA damage and repair associated protein expression and the results indicated that α-PA increased p-p53, p-H2A.X, 14-3-3-σ, and MDC1 protein expression but inhibited the protein of p53, MGMT, DNA-PK, and BRCA-1. © 2014 Wiley Periodicals, Inc.

  20. Dietary Protein Sources Affect Internal Quality of Raw and Cooked Shell Eggs under Refrigerated Conditions

    Directory of Open Access Journals (Sweden)

    X. C. Wang

    2015-11-01

    Full Text Available This study was conducted to evaluate the effects of various protein sources (soybean meal, SBM; cottonseed protein, CSP; double-zero rapeseed meal, DRM on the internal quality of refrigerated eggs. A total of 360 laying hens (32 wk of age were randomly allotted to six treatment groups (five replicates per treatment and fed diets containing SBM, CSP, or DRM individually or in combination with equal crude protein content (SBM-CSP, SBM-DRM, and CSP-DRM as the protein ingredient(s. A 6×3 factorial arrangement was employed with dietary types and storage time (0 d, 2 wk, and 4 wk as the main effects. After 12 wk of diet feeding, a total of 270 eggs were collected for egg quality determination. The egg Haugh unit (HU in the CSP, SBM-DRM, and DRM groups were significantly lower than those in the SBM and SBM-CSP groups. The hardness and springiness of the cooked yolk in the CSP group were significantly higher than those in the other treatment groups. A lower HU, lower yolk index and higher albumen pH were observed in the DRM group compared to the SBM and SBM-CSP groups when the eggs were stored to 4 wk, and the HU was improved in the CSP-DRM group compared to the DRM group (p<0.05. Higher yolk hardness was observed in the CSP group compared to the other groups during storage (p<0.05, but the hardness of the cooked yolk in the SBM-CSP and CSP-DRM groups showed no difference in comparison to the SBM group. In conclusion, CSP may ameliorate the negative effects of DRM on the HU of refrigerated eggs, and SBM or DRM may alleviate the adverse effects of CSP on yolk hardness.

  1. Cocoa and Whey Protein Differentially Affect Markers of Lipid and Glucose Metabolism and Satiety.

    Science.gov (United States)

    Campbell, Caroline L; Foegeding, E Allen; Harris, G Keith

    2016-03-01

    Food formulation with bioactive ingredients is a potential strategy to promote satiety and weight management. Whey proteins are high in leucine and are shown to decrease hunger ratings and increase satiety hormone levels; cocoa polyphenolics moderate glucose levels and slow digestion. This study examined the effects of cocoa and whey proteins on lipid and glucose metabolism and satiety in vitro and in a clinical trial. In vitro, 3T3-L1 preadipocytes were treated with 0.5-100 μg/mL cocoa polyphenolic extract (CPE) and/or 1-15 mM leucine (Leu) and assayed for lipid accumulation and leptin production. In vivo, a 6-week clinical trial consisted of nine panelists (age: 22.6 ± 1.7; BMI: 22.3 ± 2.1) consuming chocolate-protein beverages once per week, including placebo, whey protein isolate (WPI), low polyphenolic cocoa (LP), high polyphenolic cocoa (HP), LP-WPI, and HP-WPI. Measurements included blood glucose and adiponectin levels, and hunger ratings at baseline and 0.5-4.0 h following beverage consumption. At levels of 50 and 100 μg/mL, CPE significantly inhibited preadipocyte lipid accumulation by 35% and 50%, respectively, and by 22% and 36% when combined with 15 mM Leu. Leu treatment increased adipocyte leptin production by 26-37%. In the clinical trial, all beverages significantly moderated blood glucose levels 30 min postconsumption. WPI beverages elicited lowest peak glucose levels and HP levels were significantly lower than LP. The WPI and HP beverage treatments significantly increased adiponectin levels, but elicited no significant changes in hunger ratings. These trends suggest that combinations of WPI and cocoa polyphenols may improve markers of metabolic syndrome and satiety.

  2. Ice recrystallization inhibition in ice cream as affected by ice structuring proteins from winter wheat grass.

    Science.gov (United States)

    Regand, A; Goff, H D

    2006-01-01

    Ice recrystallization in quiescently frozen sucrose solutions that contained some of the ingredients commonly found in ice cream and in ice cream manufactured under commercial conditions, with or without ice structuring proteins (ISP) from cold-acclimated winter wheat grass extract (AWWE), was assessed by bright field microscopy. In sucrose solutions, critical differences in moisture content, viscosity, ionic strength, and other properties derived from the presence of other ingredients (skim milk powder, corn syrup solids, locust bean gum) caused a reduction in ice crystal growth. Significant ISP activity in retarding ice crystal growth was observed in all solutions (44% for the most complex mix) containing 0.13% total protein from AWWE. In heat-shocked ice cream, ice recrystallization rates were significantly reduced 40 and 46% with the addition of 0.0025 and 0.0037% total protein from AWWE. The ISP activity in ice cream was not hindered by its inclusion in mix prior to pasteurization. A synergistic effect between ISP and stabilizer was observed, as ISP activity was reduced in the absence of stabilizer in ice cream formulations. A remarkably smoother texture for ice creams containing ISP after heat-shock storage was evident by sensory evaluation. The efficiency of ISP from AWWE in controlling ice crystal growth in ice cream has been demonstrated.

  3. Screening of mutations affecting protein stability and dynamics of FGFR1—A simulation analysis

    Directory of Open Access Journals (Sweden)

    C. George Priya Doss

    2012-12-01

    Full Text Available Single amino acid substitutions in Fibroblast Growth Factor Receptor 1 (FGFR1 destabilize protein and have been implicated in several genetic disorders like various forms of cancer, Kallamann syndrome, Pfeiffer syndrome, Jackson Weiss syndrome, etc. In order to gain functional insight into mutation caused by amino acid substitution to protein function and expression, special emphasis was laid on molecular dynamics simulation techniques in combination with in silico tools such as SIFT, PolyPhen 2.0, I-Mutant 3.0 and SNAP. It has been estimated that 68% nsSNPs were predicted to be deleterious by I-Mutant, slightly higher than SIFT (37%, PolyPhen 2.0 (61% and SNAP (58%. From the observed results, P722S mutation was found to be most deleterious by comparing results of all in silico tools. By molecular dynamics approach, we have shown that P722S mutation leads to increase in flexibility, and deviated more from the native structure which was supported by the decrease in the number of hydrogen bonds. In addition, biophysical analysis revealed a clear insight of stability loss due to P722S mutation in FGFR1 protein. Majority of mutations predicted by these in silico tools were in good concordance with the experimental results.

  4. Dietary protein level and source differentially affect bone metabolism, strength, and intestinal calcium transporter expression during ad libitum and food-restricted conditions in male rats

    Science.gov (United States)

    High protein diets may attenuate bone loss during energy restriction (ER). The objective of the current study was to determine whether high protein diets suppress bone turnover and improve bone quality in rats during ER and whether dietary protein source affects this relationship. Eighty 12-week o...

  5. Protein corona affects the relaxivity and MRI contrast efficiency of magnetic nanoparticles.

    Science.gov (United States)

    Amiri, Houshang; Bordonali, Lorenzo; Lascialfari, Alessandro; Wan, Sha; Monopoli, Marco P; Lynch, Iseult; Laurent, Sophie; Mahmoudi, Morteza

    2013-09-21

    Magnetic nanoparticles (NPs) are increasingly being considered for use in biomedical applications such as biosensors, imaging contrast agents and drug delivery vehicles. In a biological fluid, proteins associate in a preferential manner with NPs. The small sizes and high curvature angles of NPs influence the types and amounts of proteins present on their surfaces. This differential display of proteins bound to the surface of NPs can influence the tissue distribution, cellular uptake and biological effects of NPs. To date, the effects of adsorption of a protein corona (PC) on the magnetic properties of NPs have not been considered, despite the fact that some of their potential applications require their use in human blood. Here, to investigate the effects of a PC (using fetal bovine serum) on the MRI contrast efficiency of superparamagnetic iron oxide NPs (SPIONs), we have synthesized two series of SPIONs with variation in the thickness and functional groups (i.e. surface charges) of the dextran surface coating. We have observed that different physico-chemical characteristics of the dextran coatings on the SPIONs lead to the formation of PCs of different compositions. (1)H relaxometry was used to obtain the longitudinal, r1, and transverse, r2, relaxivities of the SPIONs without and with a PC, as a function of the Larmor frequency. The transverse relaxivity, which determines the efficiency of negative contrast agents (CAs), is very much dependent on the functional group and the surface charge of the SPIONs' coating. The presence of the PC did not alter the relaxivity of plain SPIONs, while it slightly increased the relaxivity of the negatively charged SPIONs and dramatically decreased the relaxivity of the positively charged ones, which was coupled with particle agglomeration in the presence of the proteins. To confirm the effect of the PC on the MRI contrast efficiency, in vitro MRI experiments at ν = 8.5 MHz were performed using a low-field MRI scanner. The MRI

  6. Estrogenic botanical supplements, health-related quality of life, fatigue, and hormone-related symptoms in breast cancer survivors: a HEAL study report

    Directory of Open Access Journals (Sweden)

    Ma Huiyan

    2011-11-01

    Full Text Available Abstract Background It remains unclear whether estrogenic botanical supplement (EBS use influences breast cancer survivors' health-related outcomes. Methods We examined the associations of EBS use with health-related quality of life (HRQOL, with fatigue, and with 15 hormone-related symptoms such as hot flashes and night sweats among 767 breast cancer survivors participating in the Health, Eating, Activity, and Lifestyle (HEAL Study. HRQOL was measured by the Medical Outcomes Study short form-36 physical and mental component scale summary score. Fatigue was measured by the Revised-Piper Fatigue Scale score. Results Neither overall EBS use nor the number of EBS types used was associated with HRQOL, fatigue, or hormone-related symptoms. However, comparisons of those using each specific type of EBS with non-EBS users revealed the following associations. Soy supplements users were more likely to have a better physical health summary score (odds ratio [OR] = 1.66, 95% confidence interval [CI] = 1.02-2.70. Flaxseed oil users were more likely to have a better mental health summary score (OR = 1.76, 95% CI = 1.05-2.94. Ginseng users were more likely to report severe fatigue and several hormone-related symptoms (all ORs ≥ 1.7 and all 95% CIs exclude 1. Red clover users were less likely to report weight gain, night sweats, and difficulty concentrating (all OR approximately 0.4 and all 95% CIs exclude 1. Alfalfa users were less likely to experience sleep interruption (OR = 0.28, 95% CI = 0.12-0.68. Dehydroepiandrosterone users were less likely to have hot flashes (OR = 0.33, 95% CI = 0.14-0.82. Conclusions Our findings indicate that several specific types of EBS might have important influences on a woman's various aspects of quality of life, but further verification is necessary.

  7. Postovulatory aging affects dynamics of mRNA, expression and localization of maternal effect proteins, spindle integrity and pericentromeric proteins in mouse oocytes.

    Science.gov (United States)

    Trapphoff, T; Heiligentag, M; Dankert, D; Demond, H; Deutsch, D; Fröhlich, T; Arnold, G J; Grümmer, R; Horsthemke, B; Eichenlaub-Ritter, U

    2016-01-01

    Is the postovulatory aging-dependent differential decrease of mRNAs and polyadenylation of mRNAs coded by maternal effect genes associated with altered abundance and distribution of maternal effect and RNA-binding proteins (MSY2)? Postovulatory aging results in differential reduction in abundance of maternal effect proteins, loss of RNA-binding proteins from specific cytoplasmic domains and critical alterations of pericentromeric proteins without globally affecting protein abundance. Oocyte postovulatory aging is associated with differential alteration in polyadenylation and reduction in abundance of mRNAs coded by selected maternal effect genes. RNA-binding and -processing proteins are involved in storage, polyadenylation and degradation of mRNAs thus regulating stage-specific recruitment of maternal mRNAs, while chromosomal proteins that are stage-specifically expressed at pericentromeres, contribute to control of chromosome segregation and regulation of gene expression in the zygote. Germinal vesicle (GV) and metaphase II (MII) oocytes from sexually mature C57B1/6J female mice were investigated. Denuded in vivo or in vitro matured MII oocytes were postovulatory aged and analyzed by semiquantitative confocal microscopy for abundance and localization of polyadenylated RNAs, proteins of maternal effect genes (transcription activator BRG1 also known as ATP-dependent helicase SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4 (SMARCA4) and NOD-like receptor family pyrin domain containing 5 (NLRP5) also known as MATER), RNA-binding proteins (MSY2 also known as germ cell-specific Y-box-binding protein, YBX2), and post-transcriptionally modified histones (trimethylated histone H3K9 and acetylated histone H4K12), as well as pericentromeric ATRX (alpha thalassemia/mental retardation syndrome X-linked, also termed ATP-dependent helicase ATRX or X-linked nuclear protein (XNP)). For proteome analysis five replicates of 30 mouse

  8. Protein corona affects the relaxivity and MRI contrast efficiency of magnetic nanoparticles

    Science.gov (United States)

    Amiri, Houshang; Bordonali, Lorenzo; Lascialfari, Alessandro; Wan, Sha; Monopoli, Marco P.; Lynch, Iseult; Laurent, Sophie; Mahmoudi, Morteza

    2013-08-01

    Magnetic nanoparticles (NPs) are increasingly being considered for use in biomedical applications such as biosensors, imaging contrast agents and drug delivery vehicles. In a biological fluid, proteins associate in a preferential manner with NPs. The small sizes and high curvature angles of NPs influence the types and amounts of proteins present on their surfaces. This differential display of proteins bound to the surface of NPs can influence the tissue distribution, cellular uptake and biological effects of NPs. To date, the effects of adsorption of a protein corona (PC) on the magnetic properties of NPs have not been considered, despite the fact that some of their potential applications require their use in human blood. Here, to investigate the effects of a PC (using fetal bovine serum) on the MRI contrast efficiency of superparamagnetic iron oxide NPs (SPIONs), we have synthesized two series of SPIONs with variation in the thickness and functional groups (i.e. surface charges) of the dextran surface coating. We have observed that different physico-chemical characteristics of the dextran coatings on the SPIONs lead to the formation of PCs of different compositions. 1H relaxometry was used to obtain the longitudinal, r1, and transverse, r2, relaxivities of the SPIONs without and with a PC, as a function of the Larmor frequency. The transverse relaxivity, which determines the efficiency of negative contrast agents (CAs), is very much dependent on the functional group and the surface charge of the SPIONs' coating. The presence of the PC did not alter the relaxivity of plain SPIONs, while it slightly increased the relaxivity of the negatively charged SPIONs and dramatically decreased the relaxivity of the positively charged ones, which was coupled with particle agglomeration in the presence of the proteins. To confirm the effect of the PC on the MRI contrast efficiency, in vitro MRI experiments at ν = 8.5 MHz were performed using a low-field MRI scanner. The MRI

  9. Identification of Chloride Intracellular Channel Protein 3 as a Novel Gene Affecting Human Bone Formation

    DEFF Research Database (Denmark)

    Brum, A M; Leije, M; J, Schreuders-Koedam

    2017-01-01

    involved in osteoblast differentiation of human bone marrow–derived MSCs (hMSCs). We identified the gene chloride intracellular channel protein 3 (CLIC3) to be strongly upregulated during MSC-derived osteoblast differentiation. Lentiviral overexpression of CLIC3 in hMSCs caused a 60% increase of matrix......-fold increase in bone formation (0.33% versus 5.05% bone area relative to scaffold). Using a Clic3-His-tagged pull-down assay and liquid chromatography–mass spectrometry (LS/MS)-based proteomics analysis in lysates of osteogenically differentiated hMSCs, we showed that CLIC3 interacts with NIMA...

  10. Inhibition of protein kinase C affects on mode of synaptic vesicle exocytosis due to cholesterol depletion.

    Science.gov (United States)

    Petrov, Alexey M; Zakyrjanova, Guzalija F; Yakovleva, Anastasia A; Zefirov, Andrei L

    2015-01-02

    Previous studies demonstrated that depletion of membrane cholesterol by 10mM methyl-beta-cyclodextrin (MCD) results in increased spontaneous exocytosis at both peripheral and central synapses. Here, we investigated the role of protein kinase C in the enhancement of spontaneous exocytosis at frog motor nerve terminals after cholesterol depletion using electrophysiological and optical methods. Inhibition of the protein kinase C by myristoylated peptide and chelerythrine chloride prevented MCD-induced increases in FM1-43 unloading, whereas the frequency of spontaneous postsynaptic events remained enhanced. The increase in FM1-43 unloading still could be observed if sulforhodamine 101 (the water soluble FM1-43 quencher that can pass through the fusion pore) was added to the extracellular solution. This suggests a possibility that exocytosis of synaptic vesicles under these conditions could occur through the kiss-and-run mechanism with the formation of a transient fusion pore. Inhibition of phospholipase C did not lead to similar change in MCD-induced exocytosis. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Identification of QTL Affecting Protein and Amino Acid Contents in Rice

    Directory of Open Access Journals (Sweden)

    Ming ZHONG

    2011-09-01

    Full Text Available The phenotypes of protein and amino acid contents were measured in an F9 recombinant inbred line population derived from a cross between Zhenshan 97B and Delong 208. A total of 48 and 64 QTLs were identified in 2004 and 2005, respectively. The contribution of each QTL to the phenotypic variation ranged from 4.0% to 43.7%. Most QTLs co-localized, forming 29 QTL clusters on the chromosomes with three major ones detected in both years, which were mapped on chromosomes 1, 7 and 9, respectively. The two QTL clusters for amino acid content, qAa1 and qAa7, influenced almost all the traits with the allele from Zhenshan 97B, and the third QTL cluster for amino acid content, qAa9, increased the lysine content with the allele from Delong 208. A wide coincidence was found between the QTL detected under this study and the loci involved in amino acid metabolism pathways in nitrogen assimilation and transport, or protein biosynthesis. The results would facilitate the identification of candidate genes and could be used in marker-assisted selection for the favorable allele in rice quality improvement.

  12. A mutation in protein phosphatase 2A regulatory subunit A affects auxin transport in Arabidopsis

    Science.gov (United States)

    Garbers, C.; DeLong, A.; Deruere, J.; Bernasconi, P.; Soll, D.; Evans, M. L. (Principal Investigator)

    1996-01-01

    The phytohormone auxin controls processes such as cell elongation, root hair development and root branching. Tropisms, growth curvatures triggered by gravity, light and touch, are also auxin-mediated responses. Auxin is synthesized in the shoot apex and transported through the stem, but the molecular mechanism of auxin transport is not well understood. Naphthylphthalamic acid (NPA) and other inhibitors of auxin transport block tropic curvature responses and inhibit root and shoot elongation. We have isolated a novel Arabidopsis thaliana mutant designated roots curl in NPA (rcn1). Mutant seedlings exhibit altered responses to NPA in root curling and hypocotyl elongation. Auxin efflux in mutant seedlings displays increased sensitivity to NPA. The rcn1 mutation was transferred-DNA (T-DNA) tagged and sequences flanking the T-DNA insert were cloned. Analysis of the RCN1 cDNA reveals that the T-DNA insertion disrupts a gene for the regulatory A subunit of protein phosphatase 2A (PP2A-A). The RCN1 gene rescues the rcn1 mutant phenotype and also complements the temperature-sensitive phenotype of the Saccharomyces cerevisiae PP2A-A mutation, tpd3-1. These data implicate protein phosphatase 2A in the regulation of auxin transport in Arabidopsis.

  13. Maternal High Fat Diet Affects Offspring's Vitamin K-Dependent Proteins Expression Levels.

    Directory of Open Access Journals (Sweden)

    Stuart Lanham

    Full Text Available Studies suggest bone growth & development and susceptibility to vascular disease in later life are influenced by maternal nutrition, during intrauterine and early postnatal life. There is evidence for a role of vitamin K-dependent proteins (VKDPs including Osteocalcin, Matrix-gla protein, Periostin, and Gas6, in bone and vascular development. This study extends the analysis of VKDPs previously conducted in 6 week old offspring, into offspring of 30 weeks of age, to assess the longer term effects of a maternal and postnatal high fat (HF diet on VKDP expression. Overall a HF maternal diet and offspring diet exacerbated the bone changes observed. Sex specific and tissue specific differences were observed in VKDP expression for both aorta and femoral tissues. In addition, significant correlations were observed between femoral OCN, Periostin Gas6, and Vkor expression levels and measures of femoral bone structure. Furthermore, MGP, OCN, Ggcx and Vkor expression levels correlated to mass and fat volume, in both sexes. In summary the current study has highlighted the importance of the long-term effects of maternal nutrition on offspring bone development and the correlation of VKDPs to bone structure.

  14. Inhibition of protein kinase C affects on mode of synaptic vesicle exocytosis due to cholesterol depletion

    Energy Technology Data Exchange (ETDEWEB)

    Petrov, Alexey M., E-mail: fysio@rambler.ru; Zakyrjanova, Guzalija F., E-mail: guzik121192@mail.ru; Yakovleva, Anastasia A., E-mail: nastya1234qwer@mail.ru; Zefirov, Andrei L., E-mail: zefiroval@rambler.ru

    2015-01-02

    Highlights: • We examine the involvement of PKC in MCD induced synaptic vesicle exocytosis. • PKC inhibitor does not decrease the effect MCD on MEPP frequency. • PKC inhibitor prevents MCD induced FM1-43 unloading. • PKC activation may switch MCD induced exocytosis from kiss-and-run to a full mode. • Inhibition of phospholipase C does not lead to similar change in exocytosis. - Abstract: Previous studies demonstrated that depletion of membrane cholesterol by 10 mM methyl-beta-cyclodextrin (MCD) results in increased spontaneous exocytosis at both peripheral and central synapses. Here, we investigated the role of protein kinase C in the enhancement of spontaneous exocytosis at frog motor nerve terminals after cholesterol depletion using electrophysiological and optical methods. Inhibition of the protein kinase C by myristoylated peptide and chelerythrine chloride prevented MCD-induced increases in FM1-43 unloading, whereas the frequency of spontaneous postsynaptic events remained enhanced. The increase in FM1-43 unloading still could be observed if sulforhodamine 101 (the water soluble FM1-43 quencher that can pass through the fusion pore) was added to the extracellular solution. This suggests a possibility that exocytosis of synaptic vesicles under these conditions could occur through the kiss-and-run mechanism with the formation of a transient fusion pore. Inhibition of phospholipase C did not lead to similar change in MCD-induced exocytosis.

  15. The FlgT protein is involved in Aeromonas hydrophila polar flagella stability and not affects anchorage of lateral flagella.

    Directory of Open Access Journals (Sweden)

    Susana Merino

    2016-07-01

    Full Text Available Aeromonas hydrophila sodium-driven polar flagellum has a complex stator-motor. Consist of two sets of redundant and non-exchangeable proteins (PomA/PomB and PomA2/PomB2, which are homologous to other sodium-conducting polar flagellum stator motors; and also two essential proteins (MotX and MotY, that they interact with one of those two redundant pairs of proteins and form the T-ring. In this work, we described an essential protein for polar flagellum stability and rotation which is orthologous to Vibrio spp. FlgT and it is encoded outside of the A. hydrophila polar flagellum regions. The flgT was present in all mesophilic Aeromonas strains tested and also in the non-motile Aeromonas salmonicida. The A. hydrophila ∆flgT mutant is able to assemble the polar flagellum but is more unstable and released into the culture supernatant from the cell upon completion assembly. Presence of FlgT in purified polar hook-basal bodies (HBB of wild-type strain was confirmed by Western blotting and electron microscopy observations showed an outer ring of the T-ring (H-ring which is not present in the ∆flgT mutant. Anchoring and motility of proton-driven lateral flagella was not affected in the ∆flgT mutant and specific antibodies did not detect FlgT in purified lateral HBB of wild type strain.

  16. Apple polyphenols affect protein kinase C activity and the onset of apoptosis in human colon carcinoma cells.

    Science.gov (United States)

    Kern, Melanie; Pahlke, Gudrun; Balavenkatraman, Kamal Kumar; Böhmer, Frank D; Marko, Doris

    2007-06-27

    Polyphenol-rich apple extracts have been reported to suppress human colon cancer cell growth in vitro. The protein kinase C (PKC) is among the signaling elements known to play an important role in colon carcinogenesis. In the present study, we investigated whether apple polyphenols affect PKC activity and induce apoptosis in the human colon carcinoma cell line HT29. A polyphenol-rich apple juice extract (AE02) was shown to inhibit cytosolic PKC activity in a cell-free system. In contrast, incubation of HT29 cells for 1 or 3 h with AE02 up to 2 mg/mL did not affect the cytosolic PKC activity. After prolonged incubation (24 h), cytosolic PKC activity was modulated, albeit a u-shaped curve of effectiveness was observed, with an initial inhibitory effect followed by the recurrence and even induction of enzyme activity. Concomitantly, in the cytosol, a significant decrease of the protein levels of PKCalpha, PKCbetaII, and PKCgamma together with a significant increase of a proapoptotic PKCdelta fragment was observed. However, the effects on the protein levels of these PKC isoforms in the cytosol were not associated with translocation between the different cellular compartments but might instead result from the onset of apoptosis. Indeed, the treatment with AE02 was shown to induce apoptosis by the activation of caspase-3, DNA fragmentation, and cleavage of poly(ADP ribose) polymerase. So far, identified and available constituents of the apple extract did not contribute substantially to the observed effects on PKC and apoptosis induction. In summary, apple polyphenols were found to inhibit PKC activity in a cell-free system. However, our results indicate that within intact cells PKC does not represent the primary target of apple polyphenols but appears to be affected in the course of apoptosis induction.

  17. [Pitfalls in measuring urinary proteins: age-related changes in urinary creatinine excretion that affect the urine protein/creatinine ratio].

    Science.gov (United States)

    Yuno, Tomoji; Hisada, Yukimasa; Nishimura, Yasuyuki

    2011-02-01

    Knowing the amount of protein excreted in the urine is important in determining the severity and activity of renal diseases. In general, screening tests have been carried out using the urine dipstick. However, there are limitations in determining the amount of urinary protein excretion using qualitative tests for protein in spot urine samples due to the concentration and dilution of urine. Therefore, when using spot urine samples, it is helpful to calculate the urine protein/creatinine ratio (P/C) by simultaneous measurement of urinary creatinine for determining daily protein excretion. We examined P/C measurements using the dipstick method in 22,718 subjects who visited our hospital for health examinations. The results showed positive rates for qualitative urinary protein (1 + and more) of 4.2% for males and 2.7% for females. Also positive rates for P/C (150 mg/g.cre and more) were found of 7.7% for males and 10.2% for females. The results showed a reversal of positive rates for males and females compared with the results of qualitative urinary protein. In addition, P/C showed a higher positive rate in 70 years old or older both for males and females. The distribution of urinary creatinine levels simultaneously measured by dipstick method showed that the percentage of diluted urine with urinary creatinine level less than 50 mg/dL was 6.8% for males and 18.3% for females overall. Females showed a higher rate and the percentage tended to increase with age both for males and females. From these results, it was suggested that changes in urinary creatinine excretion with age that affect the P/C ratio are large. We then measured the albumin excretion rate in the 24-hour urine as well as examined the correlation between the urinary creatinine concentration and albumin index with regard to age and sex in 1,280 diabetic patients. The results showed that daily urinary creatinine excretion overall in males, overall in females, in males over 80 years old and in females over

  18. A DYW-protein knockout in Physcomitrella affects two closely spaced mitochondrial editing sites and causes a severe developmental phenotype

    DEFF Research Database (Denmark)

    Schallenberg-Rüdinger, Mareike; Kindgren, Peter; Zehrmann, Anja

    2013-01-01

    RNA-binding pentatricopeptide repeat (PPR) proteins carrying a carboxy-terminal DYW domain similar to cytidine deaminases have been characterized as site-specific factors for C-to-U RNA editing in plant organelles. Here we report that knockout of DYW-PPR_65 in Physcomitrella patens causes a severe...... developmental phenotype in the moss and specifically affects two editing sites located 18 nucleotides apart on the mitochondrial ccmFC mRNA. Intriguingly, PPR_71, another DYW-type PPR, had been identified previously as an editing factor specifically affecting only the downstream editing site, ccmFCeU122SF....... The now characterized PPR_65 binds specifically only to the upstream target site, ccmFCeU103PS, in full agreement with a recent RNA-recognition code for PPR arrays. The functional interference between the two editing events may be caused by a combination of three factors: (i) the destabilization of an RNA...

  19. Metallothionein 2A affects the cell respiration by suppressing the expression of mitochondrial protein cytochrome c oxidase subunit II.

    Science.gov (United States)

    Bragina, Olga; Gurjanova, Karina; Krishtal, Jekaterina; Kulp, Maria; Karro, Niina; Tõugu, Vello; Palumaa, Peep

    2015-06-01

    Metallothioneins (MT) are involved in a broad range of cellular processes and play a major role in protection of cells towards various stressors. Two functions of MTs, namely the maintaining of the homeostasis of transition metal ions and the redox balance, are directly linked to the functioning of mitochondria. Dyshomeostasis of MTs is often related with malfunctioning of mitochondria; however, the mechanism by which MTs affect the mitochondrial respiratory chain is still unknown. We demonstrated that overexpression of MT-2A in HEK cell line decreased the oxidative phosphorylation capacity of the cells. HEK cells overexpressing MT-2A demonstrated reduced oxygen consumption and lower cellular ATP levels. MT-2A did not affect the number of mitochondria, but reduced specifically the level of cytochrome c oxidase subunit II protein, which resulted in lower activity of the complex IV.

  20. POPDC1(S201F) causes muscular dystrophy and arrhythmia by affecting protein trafficking.

    Science.gov (United States)

    Schindler, Roland F R; Scotton, Chiara; Zhang, Jianguo; Passarelli, Chiara; Ortiz-Bonnin, Beatriz; Simrick, Subreena; Schwerte, Thorsten; Poon, Kar-Lai; Fang, Mingyan; Rinné, Susanne; Froese, Alexander; Nikolaev, Viacheslav O; Grunert, Christiane; Müller, Thomas; Tasca, Giorgio; Sarathchandra, Padmini; Drago, Fabrizio; Dallapiccola, Bruno; Rapezzi, Claudio; Arbustini, Eloisa; Di Raimo, Francesca Romana; Neri, Marcella; Selvatici, Rita; Gualandi, Francesca; Fattori, Fabiana; Pietrangelo, Antonello; Li, Wenyan; Jiang, Hui; Xu, Xun; Bertini, Enrico; Decher, Niels; Wang, Jun; Brand, Thomas; Ferlini, Alessandra

    2016-01-01

    The Popeye domain-containing 1 (POPDC1) gene encodes a plasma membrane-localized cAMP-binding protein that is abundantly expressed in striated muscle. In animal models, POPDC1 is an essential regulator of structure and function of cardiac and skeletal muscle; however, POPDC1 mutations have not been associated with human cardiac and muscular diseases. Here, we have described a homozygous missense variant (c.602C>T, p.S201F) in POPDC1, identified by whole-exome sequencing, in a family of 4 with cardiac arrhythmia and limb-girdle muscular dystrophy (LGMD). This allele was absent in known databases and segregated with the pathological phenotype in this family. We did not find the allele in a further screen of 104 patients with a similar phenotype, suggesting this mutation to be family specific. Compared with WT protein, POPDC1(S201F) displayed a 50% reduction in cAMP affinity, and in skeletal muscle from patients, both POPDC1(S201F) and WT POPDC2 displayed impaired membrane trafficking. Forced expression of POPDC1(S201F) in a murine cardiac muscle cell line (HL-1) increased hyperpolarization and upstroke velocity of the action potential. In zebrafish, expression of the homologous mutation (popdc1(S191F)) caused heart and skeletal muscle phenotypes that resembled those observed in patients. Our study therefore identifies POPDC1 as a disease gene causing a very rare autosomal recessive cardiac arrhythmia and LGMD, expanding the genetic causes of this heterogeneous group of inherited rare diseases.

  1. HIV-1 TAT protein enhances sensitization to methamphetamine by affecting dopaminergic function.

    Science.gov (United States)

    Kesby, James P; Najera, Julia A; Romoli, Benedetto; Fang, Yiding; Basova, Liana; Birmingham, Amanda; Marcondes, Maria Cecilia G; Dulcis, Davide; Semenova, Svetlana

    2017-10-01

    Methamphetamine abuse is common among humans with immunodeficiency virus (HIV). The HIV-1 regulatory protein TAT induces dysfunction of mesolimbic dopaminergic systems which may result in impaired reward processes and contribute to methamphetamine abuse. These studies investigated the impact of TAT expression on methamphetamine-induced locomotor sensitization, underlying changes in dopamine function and adenosine receptors in mesolimbic brain areas and neuroinflammation (microgliosis). Transgenic mice with doxycycline-induced TAT protein expression in the brain were tested for locomotor activity in response to repeated methamphetamine injections and methamphetamine challenge after a 7-day abstinence period. Dopamine function in the nucleus accumbens (Acb) was determined using high performance liquid chromatography. Expression of dopamine and/or adenosine A receptors (ADORA) in the Acb and caudate putamen (CPu) was assessed using RT-PCR and immunohistochemistry analyses. Microarrays with pathway analyses assessed dopamine and adenosine signaling in the CPu. Activity-dependent neurotransmitter switching of a reserve pool of non-dopaminergic neurons to a dopaminergic phenotype in the ventral tegmental area (VTA) was determined by immunohistochemistry and quantified with stereology. TAT expression enhanced methamphetamine-induced sensitization. TAT expression alone decreased striatal dopamine (D1, D2, D4, D5) and ADORA1A receptor expression, while increasing ADORA2A receptors expression. Moreover, TAT expression combined with methamphetamine exposure was associated with increased adenosine A receptors (ADORA1A) expression and increased recruitment of dopamine neurons in the VTA. TAT expression and methamphetamine exposure induced microglia activation with the largest effect after combined exposure. Our findings suggest that dopamine-adenosine receptor interactions and reserve pool neuronal recruitment may represent potential targets to develop new treatments for

  2. How chain length and charge affect surfactant denaturation of acyl coenzyme a binding protein (ACBP)

    DEFF Research Database (Denmark)

    Andersen, Kell; Otzen, Daniel

    2009-01-01

    maltoside (DDM). The aim has been to determine how surfactant chain length and micellar charge affect the denaturation mechanism. ACBP denatures in two steps irrespective of surfactant chain length, but with increasing chain length, the potency of the denaturant rises more rapidly than the critical micelle...... concentration (cmc) declines. Increasing proportions of DDM, which significantly reduce the amount of monomelic sodium dodecyl sulfate (SDS), make the first denaturation step occur at lower concentrations but weaken and eventually remove the second denaturation step. The logarithm of the unfolding rate...... constants increases linearly with denaturant concentration below the cmc but declines at higher concentrations. Both shortening chain length and decreasing micellar charge reduce the overall kinetics of unfolding and makes the dependence of unfolding rate constants on surfactant concentration more complex...

  3. Dietary protein intake affects expression of genes for lipid metabolism in porcine skeletal muscle in a genotype-dependent manner.

    Science.gov (United States)

    Liu, Yingying; Li, Fengna; He, Lingyun; Tan, Bie; Deng, Jinping; Kong, Xiangfeng; Li, Yinghui; Geng, Meimei; Yin, Yulong; Wu, Guoyao

    2015-04-14

    Skeletal muscle is a major site for the oxidation of fatty acids (FA) in mammals, including humans. Using a swine model, we tested the hypothesis that dietary protein intake regulates the expression of key genes for lipid metabolism in skeletal muscle. A total of ninety-six barrows (forty-eight pure-bred Bama mini-pigs (fatty genotype) and forty-eight Landrace pigs (lean genotype)) were fed from 5 weeks of age to market weight. Pigs of fatty or lean genotype were randomly assigned to one of two dietary treatments (low- or adequate-protein diet), with twenty-four individually fed pigs per treatment. Our data showed that dietary protein levels affected the expression of genes involved in the anabolism and catabolism of lipids in the longissimus dorsi and biceps femoris muscles in a genotype-dependent manner. Specifically, Bama mini-pigs had more intramuscular fat, SFA and MUFA, as well as elevated mRNA expression levels of lipogenic genes, compared with Landrace pigs. In contrast, Bama mini-pigs had lower mRNA expression levels of lipolytic genes than Landrace pigs fed an adequate-protein diet in the growing phase. These data are consistent with higher white-fat deposition in Bama mini-pigs than in Landrace pigs. In conclusion, adequate provision of dietary protein (amino acids) plays an important role in regulating the expression of key lipogenic genes, and the growth of white adipose tissue, in a genotype- and tissue-specific manner. These findings have important implications for developing novel dietary strategies in pig production.

  4. Single Amino Acid Polymorphisms of Pertussis Toxin Subunit S2 (PtxB Affect Protein Function.

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    Scott H Millen

    Full Text Available Whooping cough due to Bordetella pertussis is increasing in incidence, in part due to accumulation of mutations which increase bacterial fitness in highly vaccinated populations. Polymorphisms in the pertussis toxin, ptxA and ptxB genes, and the pertactin, prn genes of clinical isolates of Bordetella pertussis collected in Cincinnati from 1989 through 2005 were examined. While the ptxA and prn genotypes were variable, all 48 strains had the ptxB2 genotype; ptxB1 encodes glycine at amino acid 18 of the S2 subunit of pertussis toxin, while ptxB2 encodes serine. We investigated antigenic and functional differences of PtxB1 and PtxB2. The S2 protein was not very immunogenic. Only a few vaccinated or individuals infected with B. pertussis developed antibody responses to the S2 subunit, and these sera recognized both polymorphic forms equally well. Amino acid 18 of S2 is in a glycan binding domain, and the PtxB forms displayed differences in receptor recognition and toxicity. PtxB1 bound better to the glycoprotein, fetuin, and Jurkat T cells in vitro, but the two forms were equally effective at promoting CHO cell clustering. To investigate in vivo activity of Ptx, one μg of Ptx was administered to DDY mice and blood was collected on 4 days after injection. PtxB2 was more effective at promoting lymphocytosis in mice.

  5. Obesity, sex and pubertal status affect appetite hormone responses to a mixed glucose and whey protein drink in adolescents.

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    Patel, Barkha P; Anderson, G Harvey; Vien, Shirley; Bellissimo, Nick; McCrindle, Brian W; Hamilton, Jill K

    2014-07-01

    Little information is available on how food intake regulatory hormones may be altered during pubertal development and across the weight spectrum in adolescents. Therefore, the effect of obesity, sex and pubertal status on subjective appetite and appetite hormones in response to a mixed glucose and whey protein drink was determined in 8-18 year old adolescents. A cross-sectional cohort study was conducted at the Hospital for Sick Children, Toronto. After a 12 h fast, normal weight (n = 5 female, 4 male) and obese (n = 5 female, 4 male) adolescents (Experiment 1), and pre-early pubertal (n = 10) and mid-late pubertal (n = 10) obese male adolescents (Experiment 2) consumed a 250 ml glucose (30 g) and whey protein (30 g) beverage. Insulin, PYY, ghrelin and subjective appetite were measured over 120 min. Obese adolescents (Experiment 1) have higher insulin, PYY and lower ghrelin (P Puberty (Experiment 2) did not affect insulin (P = 0·305), but the change in PYY in response to the drink was greater (P = 0·032) and ghrelin was lower (P = 0·026) in mid-late pubertal than pre-early pubertal obese males. Average appetite 60 min post-drink was higher in obese and mid-late pubertal adolescents, but not related to hormone changes. Obesity, sex and pubertal status affect macronutrient-stimulated appetite hormone secretion and these factors may alter food intake in obese children during pubertal development. © 2014 John Wiley & Sons Ltd.

  6. Sake Protein Supplementation Affects Exercise Performance and Biochemical Profiles in Power-Exercise-Trained Mice.

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    Chen, Yi-Ming; Lin, Che-Li; Wei, Li; Hsu, Yi-Ju; Chen, Kuan-Neng; Huang, Chi-Chang; Kao, Chin-Hsung

    2016-02-20

    Exercise and fitness training programs have attracted the public's attention in recent years. Sports nutrition supplementation is an important issue in the global sports market. In this study, we designed a power exercise training (PET) program with a mouse model based on a strength and conditional training protocol for humans. We tested the effect of supplementation with functional branched-chain amino acid (BCAA)-rich sake protein (SP) to determine whether the supplement had a synergistic effect during PET and enhanced athletic performance and resistance to fatigue. Male ICR mice were divided into three groups (n = 8 per group) for four-week treatment: sedentary controls with vehicle (SC), and PET and PET groups with SP supplementation (3.8 g/kg, PET + SP). Exercise performance was evaluated by forelimb grip strength and exhaustive swimming time as well as changes in body composition and anti-fatigue activity levels of serum lactate, ammonia, glucose, and creatine kinase (CK) after a 15-min swimming exercise. The biochemical parameters were measured at the end of the experiment. four-week PET significantly increased grip strength and exhaustive swimming time and decreased epididymal fat pad (EFP) weight and area. Levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinine, and uric acid (UA) were significantly increased. PET + SP supplementation significantly decreased serum lactate, ammonia and CK levels after the 15-min swimming exercise. The resting serum levels of AST, ALT, CREA and UA were all significantly decreased with PET + SP. The PET program could increase the exercise performance and modulate the body composition of mice. PET with SP conferred better anti-fatigue activity, improved biochemical profiles, and may be an effective ergogenic aid in strength training.

  7. Sake Protein Supplementation Affects Exercise Performance and Biochemical Profiles in Power-Exercise-Trained Mice

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    Yi-Ming Chen

    2016-02-01

    Full Text Available Exercise and fitness training programs have attracted the public’s attention in recent years. Sports nutrition supplementation is an important issue in the global sports market. Purpose: In this study, we designed a power exercise training (PET program with a mouse model based on a strength and conditional training protocol for humans. We tested the effect of supplementation with functional branched-chain amino acid (BCAA-rich sake protein (SP to determine whether the supplement had a synergistic effect during PET and enhanced athletic performance and resistance to fatigue. Methods: Male ICR mice were divided into three groups (n = 8 per group for four-week treatment: sedentary controls with vehicle (SC, and PET and PET groups with SP supplementation (3.8 g/kg, PET + SP. Exercise performance was evaluated by forelimb grip strength and exhaustive swimming time as well as changes in body composition and anti-fatigue activity levels of serum lactate, ammonia, glucose, and creatine kinase (CK after a 15-min swimming exercise. The biochemical parameters were measured at the end of the experiment. Results: four-week PET significantly increased grip strength and exhaustive swimming time and decreased epididymal fat pad (EFP weight and area. Levels of aspartate aminotransferase (AST, alanine aminotransferase (ALT, creatinine, and uric acid (UA were significantly increased. PET + SP supplementation significantly decreased serum lactate, ammonia and CK levels after the 15-min swimming exercise. The resting serum levels of AST, ALT, CREA and UA were all significantly decreased with PET + SP. Conclusion: The PET program could increase the exercise performance and modulate the body composition of mice. PET with SP conferred better anti-fatigue activity, improved biochemical profiles, and may be an effective ergogenic aid in strength training.

  8. Repression of TERMINAL FLOWER1 primarily mediates floral induction in pear (Pyrus pyrifolia Nakai) concomitant with change in gene expression of plant hormone-related genes and transcription factors.

    Science.gov (United States)

    Bai, Songling; Tuan, Pham Anh; Saito, Takanori; Ito, Akiko; Ubi, Benjamin Ewa; Ban, Yusuke; Moriguchi, Takaya; Wilson, Zoe

    2017-10-13

    Floral induction is an important event in the annual growth cycle of perennial fruit trees. For pear, this event directly affects fruit production in the following year. The flower buds in many species are induced by FLOWERING LOCUS T (FT), whose effect is repressed by the meristem-expressed gene TERMINAL FLOWER1 (TFL1). In this study, we investigated the functions of pear FT and TFL1 genes during floral development. Expression of pear FTs (PpFT1a and PpFT2a) in reproductive meristems was not obviously induced prior to floral initiation, while expression of TFL1s (PpTFL1-1a and PpTFL1-2a) rapidly decreased. The induction of the productive meristem identity MADS-box gene AP1 after repression of PpTFL1s suggested a primary role for PpTFL1 in floral induction. RNA-seq analysis suggested that plant hormone-related genes and several transcription factors that were coexpressed with PpTFL1 were potentially involved in the PpTFL1-mediated floral induction. Our data indicate the essential function of TFL1 in pear floral induction and add another species in the family Rosaceae in addition to strawberry and rose that shows a role for TFL1 in floral induction. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  9. Disruption of Protein Mannosylation Affects Candida guilliermondii Cell Wall, Immune Sensing, and Virulence

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    María J. Navarro-Arias

    2016-12-01

    Full Text Available The fungal cell wall contains glycoproteins that interact with the host immune system. In the prominent pathogenic yeast Candida albicans, Pmr1 acts as a Golgi-resident ion pump that provides cofactors to mannosyltransferases, regulating the synthesis of mannans attached to glycoproteins. To gain insight into a putative conservation of such a crucial process within opportunistic yeasts, we were particularly interested in studying the role of the PMR1 homolog in a low-virulent species that rarely causes candidiasis, Candida guilliermondii. We disrupted C. guilliermondii PMR1 and found that loss of Pmr1 affected cell growth and morphology, biofilm formation, susceptibility to cell wall perturbing agents, mannan levels, and the wall composition and organization. Despite there was a significant increment in the amount of β1,3-glucan exposed at the wall surface, this positively influenced only the ability of the mutant to stimulate IL-10 production by human monocytes, suggesting that recognition of both mannan and β1,3-glucan, is required to stimulate strong levels of pro-inflammatory cytokines. Accordingly, our results indicate C. guilliermondii sensing by monocytes was critically dependent on the recognition of N-linked mannans and β1,3-glucan, as reported in other Candida species. In addition, chemical remotion of cell wall O-linked mannans was found to positively influence the recognition of C. guilliermondii by human monocytes, suggesting that O-linked mannans mask other cell wall components from immune cells. This observation contrasts with that reported in C. albicans. Finally, mice infected with C. guilliermondii pmr1 null mutant cells had significantly lower fungal burdens compared to animals challenged with the parental strain. Accordingly, the null mutant showed inability to kill larvae in the Galleria mellonella infection model. This study thus demonstrates that mannans are relevant for the C. guilliermondii-host interaction, with

  10. The PINK1 p.I368N mutation affects protein stability and ubiquitin kinase activity.

    Science.gov (United States)

    Ando, Maya; Fiesel, Fabienne C; Hudec, Roman; Caulfield, Thomas R; Ogaki, Kotaro; Górka-Skoczylas, Paulina; Koziorowski, Dariusz; Friedman, Andrzej; Chen, Li; Dawson, Valina L; Dawson, Ted M; Bu, Guojun; Ross, Owen A; Wszolek, Zbigniew K; Springer, Wolfdieter

    2017-04-24

    Mutations in PINK1 and PARKIN are the most common causes of recessive early-onset Parkinson's disease (EOPD). Together, the mitochondrial ubiquitin (Ub) kinase PINK1 and the cytosolic E3 Ub ligase PARKIN direct a complex regulated, sequential mitochondrial quality control. Thereby, damaged mitochondria are identified and targeted to degradation in order to prevent their accumulation and eventually cell death. Homozygous or compound heterozygous loss of either gene function disrupts this protective pathway, though at different steps and by distinct mechanisms. While structure and function of PARKIN variants have been well studied, PINK1 mutations remain poorly characterized, in particular under endogenous conditions. A better understanding of the exact molecular pathogenic mechanisms underlying the pathogenicity is crucial for rational drug design in the future. Here, we characterized the pathogenicity of the PINK1 p.I368N mutation on the clinical and genetic as well as on the structural and functional level in patients' fibroblasts and in cell-based, biochemical assays. Under endogenous conditions, PINK1 p.I368N is expressed, imported, and N-terminally processed in healthy mitochondria similar to PINK1 wild type (WT). Upon mitochondrial damage, however, full-length PINK1 p.I368N is not sufficiently stabilized on the outer mitochondrial membrane (OMM) resulting in loss of mitochondrial quality control. We found that binding of PINK1 p.I368N to the co-chaperone complex HSP90/CDC37 is reduced and stress-induced interaction with TOM40 of the mitochondrial protein import machinery is abolished. Analysis of a structural PINK1 p.I368N model additionally suggested impairments of Ub kinase activity as the ATP-binding pocket was found deformed and the substrate Ub was slightly misaligned within the active site of the kinase. Functional assays confirmed the lack of Ub kinase activity. Here we demonstrated that mutant PINK1 p.I368N can not be stabilized on the OMM upon

  11. Nicotine reward and affective nicotine withdrawal signs are attenuated in calcium/calmodulin-dependent protein kinase IV knockout mice.

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    Kia J Jackson

    Full Text Available The influx of Ca(2+ through calcium-permeable nicotinic acetylcholine receptors (nAChRs leads to activation of various downstream processes that may be relevant to nicotine-mediated behaviors. The calcium activated protein, calcium/calmodulin-dependent protein kinase IV (CaMKIV phosphorylates the downstream transcription factor cyclic AMP response element binding protein (CREB, which mediates nicotine responses; however the role of CaMKIV in nicotine dependence is unknown. Given the proposed role of CaMKIV in CREB activation, we hypothesized that CaMKIV might be a crucial molecular component in the development of nicotine dependence. Using male CaMKIV genetically modified mice, we found that nicotine reward is attenuated in CaMKIV knockout (-/- mice, but cocaine reward is enhanced in these mice. CaMKIV protein levels were also increased in the nucleus accumbens of C57Bl/6 mice after nicotine reward. In a nicotine withdrawal assessment, anxiety-related behavior, but not somatic signs or the hyperalgesia response are attenuated in CaMKIV -/- mice. To complement our animal studies, we also conducted a human genetic association analysis and found that variants in the CaMKIV gene are associated with a protective effect against nicotine dependence. Taken together, our results support an important role for CaMKIV in nicotine reward, and suggest that CaMKIV has opposing roles in nicotine and cocaine reward. Further, CaMKIV mediates affective, but not physical nicotine withdrawal signs, and has a protective effect against nicotine dependence in human genetic association studies. These findings further indicate the importance of calcium-dependent mechanisms in mediating behaviors associated with drugs of abuse.

  12. In Vivo pharmacokinetics of hesperidin are affected by treatment with glucosidase-like BglA protein isolated from yeasts

    Science.gov (United States)

    Li, Yong-mei; Li, Xiao-mian; Li, Guang-ming; Du, Wen-cai; Zhang, Jing; Li, Wei-xia; Xu, Jianshe; Hu, Ming; Zhu, Ze

    2009-01-01

    Hesperidin is an abundant flavanone glycoside in citrus fruits and has been reported to possess a wide range of biological activities. However, hesperidin has poor bioavailability. Here, we test the hypothesis that hesperetin found in Chenpi will have a better bioavailability than hesperidin and that treatment of hesperidin with glucosidase-like yeast Bg1A protein will increase its bioavailability. The results indicate that hesperidin in pure or extract form is hydrolyzed by BglA protein extracted from S. singularis or expressed in E coli BL21 (DE3). This biotransformation affected the plasma pharmacokinetics of total hesperetin in rats, in that the plasma Tmax was significantly shorter after administration of BglA protein-treated hesperidin than after administration of hesperidin extract. In addition, the AUC values for total hesperetin after administration of Bg1A-treated hesperidin were approximately 4-fold higher by oral administration and 3-fold higher by intravenous administration, respectively. In contract, the plasma clearance value and volume of distribution after administration of Bg1A-treated hesperidin extract or pure hesperetin was significant smaller than after administration of untreated hesperidin extract or pure hesperidin In conclusion, this is a first study that systemically determines the absolute bioavailability of hesperidin and hesperetin simultaneously, and the study shows clearly that hesperetin is more bioavailable than hesperidin regardless of the route of administration, and that prior transformation of hesperidin to hesperetin via fermentation should significantly increase its bioavailability because of the action of yeast glycosidase-like protein BglA. PMID:18570429

  13. Rumen microorganisms, methane production, and microbial protein synthesis affected by mangosteen peel powder supplement in lactating dairy cows.

    Science.gov (United States)

    Polyorach, Sineenart; Wanapat, Metha; Cherdthong, Anusorn; Kang, Sungchhang

    2016-03-01

    Four crossbred dairy cows (50 % Holstein-Friesian × 50 % Thai native), 404 ± 50.0 kg of body weight (4 years old) and 90 ± 5 day in milk with daily milk production of 9 ± 2.0 kg/day, were randomly assigned according to a 4 × 4 Latin square design to study the effect of mangosteen (Garcinia mangostana) peel powder (MSP) supplementation on rumen microorganisms, methane production, and microbial protein synthesis fed concentrate containing yeast fermented cassava chip protein (YEFECAP). The treatments were different levels of MSP supplementation at 0, 100, 200, and 300 g/head/day. Rice straw was used as a roughage source fed ad libitum, and concentrate containing YEFECAP at 200 g/kg concentrate was offered corresponding to concentrate-to-milk-yield ratio at 1:2. A quantitative real-time PCR approach was used to determine the population densities of ruminal microorganisms. The results revealed that supplementation of MSP did not affect on Fibrobactor succinogenes, Ruminococcus flavefaciens, and Ruminococcus albus (P > 0.05). However, total bacteria was linearly increased (P methane production from 27.5 to 23.7 mmol/100 ml(3). Furthermore, cows that received MSP at 300 g/head/day had the highest microbial crude protein and efficiency of rumen microbial N synthesis (416.8 g/day and 16.2 g/kg organic matter truly digested in the rumen (OMDR), respectively). In conclusion, supplementation of MSP at 300 g/head/day with YEFECAP as a protein source in the concentrate mixture revealed an enhancement of rumen fermentation and methane reduction in lactating dairy cows.

  14. Molecular characterization of a long range haplotype affecting protein yield and mastitis susceptibility in Norwegian Red cattle

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    Hayes Ben J

    2011-08-01

    Full Text Available Abstract Background Previous fine mapping studies in Norwegian Red cattle (NRC in the region 86-90.4 Mb on Bos taurus chromosome 6 (BTA6 has revealed a quantitative trait locus (QTL for protein yield (PY around 88 Mb and a QTL for clinical mastitis (CM around 90 Mb. The close proximity of these QTLs may partly explain the unfavorable genetic correlation between these two traits in NRC. A long range haplotype covering this region was introduced into the NRC population through the importation of a Holstein-Friesian bull (1606 Frasse from Sweden in the 1970s. It has been suggested that this haplotype has a favorable effect on milk protein content but an unfavorable effect on mastitis susceptibility. Selective breeding for milk production traits is likely to have increased the frequency of this haplotype in the NRC population. Results Association mapping for PY and CM in NRC was performed using genotypes from 556 SNPs throughout the region 86-97 Mb on BTA6 and daughter-yield-deviations (DYDs from 2601 bulls made available from the Norwegian dairy herd recording system. Highest test scores for PY were found for single-nucleotide polymorphisms (SNPs within and surrounding the genes CSN2 and CSN1S2, coding for the β-casein and αS2-casein proteins. High coverage re-sequencing by high throughput sequencing technology enabled molecular characterization of a long range haplotype from 1606 Frasse encompassing these two genes. Haplotype analysis of a large number of descendants from this bull indicated that the haplotype was not markedly disrupted by recombination in this region. The haplotype was associated with both increased milk protein content and increased susceptibility to mastitis, which might explain parts of the observed genetic correlation between PY and CM in NRC. Plausible causal polymorphisms affecting PY were detected in the promoter region and in the 5'-flanking UTR of CSN1S2. These polymorphisms could affect transcription or translation of

  15. Molecular characterization of a long range haplotype affecting protein yield and mastitis susceptibility in Norwegian Red cattle.

    Science.gov (United States)

    Sodeland, Marte; Grove, Harald; Kent, Matthew; Taylor, Simon; Svendsen, Morten; Hayes, Ben J; Lien, Sigbjørn

    2011-08-11

    Previous fine mapping studies in Norwegian Red cattle (NRC) in the region 86-90.4 Mb on Bos taurus chromosome 6 (BTA6) has revealed a quantitative trait locus (QTL) for protein yield (PY) around 88 Mb and a QTL for clinical mastitis (CM) around 90 Mb. The close proximity of these QTLs may partly explain the unfavorable genetic correlation between these two traits in NRC. A long range haplotype covering this region was introduced into the NRC population through the importation of a Holstein-Friesian bull (1606 Frasse) from Sweden in the 1970s. It has been suggested that this haplotype has a favorable effect on milk protein content but an unfavorable effect on mastitis susceptibility. Selective breeding for milk production traits is likely to have increased the frequency of this haplotype in the NRC population. Association mapping for PY and CM in NRC was performed using genotypes from 556 SNPs throughout the region 86-97 Mb on BTA6 and daughter-yield-deviations (DYDs) from 2601 bulls made available from the Norwegian dairy herd recording system. Highest test scores for PY were found for single-nucleotide polymorphisms (SNPs) within and surrounding the genes CSN2 and CSN1S2, coding for the β-casein and α(S2)-casein proteins. High coverage re-sequencing by high throughput sequencing technology enabled molecular characterization of a long range haplotype from 1606 Frasse encompassing these two genes. Haplotype analysis of a large number of descendants from this bull indicated that the haplotype was not markedly disrupted by recombination in this region. The haplotype was associated with both increased milk protein content and increased susceptibility to mastitis, which might explain parts of the observed genetic correlation between PY and CM in NRC. Plausible causal polymorphisms affecting PY were detected in the promoter region and in the 5'-flanking UTR of CSN1S2. These polymorphisms could affect transcription or translation of CSN1S2 and thereby affect the amount

  16. Acute-phase proteins, oxidative stress biomarkers, proinflammatory cytokines, and cardiac troponin in Arabian mares affected with pyometra.

    Science.gov (United States)

    El-Bahr, S M; El-Deeb, W M

    2016-09-01

    New biomarkers are essential for diagnosis of pyometra in mares. In this context, 12 subfertile Arabian mares suffered from pyometra were admitted to the Veterinary Teaching Hospital. The basis for diagnosis of pyometra was positive findings of clinical examination and rectal palpation. Blood samples were collected from diseased animals and from five Arabian healthy mares, which were considered as control group. Acute-phase proteins (APP), oxidative stress biomarkers, proinflammatory cytokines, and cardiac troponin I were estimated in the harvested sera of both groups. Clinical examination revealed purulent yellowish fluid discharged from vagina of affected animals and rectal palpation of the reproductive tract revealed uterine distention. The biochemical analysis of the serum revealed significant increase in cardiac troponin I, creatin kinase, alkaline phosphatase, malondialdehyde, tumor necrosis factor α, interleukins 6, prostaglandin F2α, haptoglobin, and serum amyloid A and significant decrease in reduced glutathione, superoxide dismutase (SOD), total antioxidant capacity, and nitric oxide (NO) of mares affected with pyometra compare to control. Cardiac troponin I was positively correlated with aspartate aminotransferase, creatin kinase, malondialdehyde, alkaline phosphatase, tumor necrosis factor α, interleukins 6, prostaglandin F2α, haptoglobin and serum amyloid A and negatively correlated with glutathione, superoxide dismutase, total antioxidant capacity and nitric oxide in serum of mares affected with pyometra. Moreover, there was high positive correlation between proinflammatory cytokines and APP in serum of mares affected with pyometra. The present study suggests cardiac troponin I together with APP, proinflammatory cytokines, and oxidative stress parameters as biomarkers for pyometra in Arabian mares. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. A high protein diet during pregnancy affects hepatic gene expression of energy sensing pathways along ontogenesis in a porcine model.

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    Michael Oster

    Full Text Available In rodent models and in humans the impact of gestational diets on the offspring's phenotype was shown experimentally and epidemiologically. The underlying programming of fetal development was shown to be associated with an increased risk of degenerative diseases in adulthood, including the metabolic syndrome. There are clues that diet-dependent modifications of the metabolism during fetal life can persist until adulthood. This leads to the hypothesis that the offspring's transcriptomes show short-term and long-term changes depending on the maternal diet. To this end pregnant German landrace gilts were fed either a high protein diet (HP, 30% CP or an adequate protein diet (AP, 12% CP throughout pregnancy. Hepatic transcriptome profiles of the offspring were analyzed at prenatal (94 dpc and postnatal stages (1, 28, 188 dpn. Depending on the gestational dietary exposure, mRNA expression levels of genes related to energy metabolism, N-metabolism, growth factor signaling pathways, lipid metabolism, nucleic acid metabolism and stress/immune response were affected either in a short-term or in a long-term manner. Gene expression profiles at fetal stage 94 dpc were almost unchanged between the diets. The gestational HP diet affected the hepatic expression profiles at prenatal and postnatal stages. The effects encompassed a modulation of the genome in terms of an altered responsiveness of energy and nutrient sensing pathways. Differential expression of genes related to energy production and nutrient utilization contribute to the maintenance of development and growth performance within physiological norms, however the modulation of these pathways may be accompanied by a predisposition for metabolic disturbances up to adult stages.

  18. A high protein diet during pregnancy affects hepatic gene expression of energy sensing pathways along ontogenesis in a porcine model.

    Science.gov (United States)

    Oster, Michael; Murani, Eduard; Metges, Cornelia C; Ponsuksili, Siriluck; Wimmers, Klaus

    2011-01-01

    In rodent models and in humans the impact of gestational diets on the offspring's phenotype was shown experimentally and epidemiologically. The underlying programming of fetal development was shown to be associated with an increased risk of degenerative diseases in adulthood, including the metabolic syndrome. There are clues that diet-dependent modifications of the metabolism during fetal life can persist until adulthood. This leads to the hypothesis that the offspring's transcriptomes show short-term and long-term changes depending on the maternal diet. To this end pregnant German landrace gilts were fed either a high protein diet (HP, 30% CP) or an adequate protein diet (AP, 12% CP) throughout pregnancy. Hepatic transcriptome profiles of the offspring were analyzed at prenatal (94 dpc) and postnatal stages (1, 28, 188 dpn). Depending on the gestational dietary exposure, mRNA expression levels of genes related to energy metabolism, N-metabolism, growth factor signaling pathways, lipid metabolism, nucleic acid metabolism and stress/immune response were affected either in a short-term or in a long-term manner. Gene expression profiles at fetal stage 94 dpc were almost unchanged between the diets. The gestational HP diet affected the hepatic expression profiles at prenatal and postnatal stages. The effects encompassed a modulation of the genome in terms of an altered responsiveness of energy and nutrient sensing pathways. Differential expression of genes related to energy production and nutrient utilization contribute to the maintenance of development and growth performance within physiological norms, however the modulation of these pathways may be accompanied by a predisposition for metabolic disturbances up to adult stages.

  19. Polymorphisms in the human inhibitory signal-regulatory protein α do not affect binding to its ligand CD47.

    Science.gov (United States)

    Hatherley, Deborah; Lea, Susan M; Johnson, Steven; Barclay, A Neil

    2014-04-04

    CD47 is a widely distributed membrane protein that interacts with signal-regulatory protein α (SIRPα), an inhibitory receptor on myeloid cells that gives a "don't-eat-me" signal. Manipulation of the interaction is of considerable interest in the immunotherapy of cancer and in xenotransplantation. The amino-terminal ligand binding domain of SIRPα is highly polymorphic in contrast to the single Ig-like domain of CD47. There is confusion as to whether the polymorphisms will affect ligand binding, but this is an important point for this interaction and other paired receptors being considered as targets for therapy. We show by x-ray crystallography that one human SIRPα allele differing in 13 amino acid residues has a very similar binding site and that several different alleles all bind CD47 with similar affinity as expected because the residues are mostly surface-exposed and distant from the binding site. A peptide from the binding site of CD47 has been reported to mimic the CD47 interaction with SIRPα, but we could find no binding. We discuss the possible pitfalls in determining the affinity of weak interactions and also speculate on how SIRPα polymorphisms may have been selected by pathogens and how this may also be true in other paired receptors such as the KIRs.

  20. Selenium-enriched milk proteins and selenium yeast affect selenoprotein activity and expression differently in mouse colon.

    Science.gov (United States)

    Hu, Ying; McIntosh, Graeme H; Le Leu, Richard K; Young, Graeme P

    2010-07-01

    Certain forms of dietary Se may have an advantage in improving Se status and reducing cancer risk. The present study compared the effects of an Se-enriched milk protein product (dairy-Se) with an Se yeast (yeast-Se) on selenoprotein activity and expression in the mouse colon. Mice were fed four diets for 4 weeks: a control milk protein diet (Se at 0.068 parts per million (ppm)), dairy-Se diets with Se at 0.5 and 1 ppm, and a yeast-Se diet with Se at 1 ppm. Cytosolic glutathione peroxidase-1 (GPx-1) activity, mRNA of selenoprotein P (SeP), GPx-1, gastrointestinal glutathione peroxidase-2 (GPx-2) and thioredoxin reductase-1 (TrxR-1) were examined in the mouse colon. Dairy-Se diets did not significantly affect GPx-1 mRNA and GPx-1 activity but produced a dose-dependent increase in SeP and GPx-2 mRNA, with a significantly higher level achieved at 1 ppm Se (P supplement had any effect on TrxR-1. The present study indicates that selenoprotein levels in the mouse colon are regulated differently depending on the Se supplement. As we have previously shown that dairy-Se at 1 ppm was protective against colorectal cancer (CRC) in an azoxymethane-induced CRC mouse model, this up-regulation of colonic GPx-2 and SeP with Se supplementation may be crucial to its chemopreventive action.

  1. Associations between Single-Nucleotide Polymorphisms in Corticotropin-Releasing Hormone-Related Genes and Irritable Bowel Syndrome.

    Directory of Open Access Journals (Sweden)

    Ayaka Sasaki

    Full Text Available Irritable bowel syndrome (IBS is a common functional disorder with distinct features of stress-related pathophysiology. A key mediator of the stress response is corticotropin-releasing hormone (CRH. Although some candidate genes have been identified in stress-related disorders, few studies have examined CRH-related gene polymorphisms. Therefore, we tested our hypothesis that single-nucleotide polymorphisms (SNPs in CRH-related genes influence the features of IBS.In total, 253 individuals (123 men and 130 women participated in this study. They comprised 111 IBS individuals and 142 healthy controls. The SNP genotypes in CRH (rs28364015 and rs6472258 and CRH-binding protein (CRH-BP (rs10474485 were determined by direct sequencing and real-time polymerase chain reaction. The emotional states of the subjects were evaluated using the State-Trait Anxiety Inventory, Perceived Stress Scale, and the Self-rating Depression Scale.Direct sequencing of the rs28364015 SNP of CRH revealed no genetic variation among the study subjects. There was no difference in the genotype distributions and allele frequencies of rs6472258 and rs10474485 between IBS individuals and controls. However, IBS subjects with diarrhea symptoms without the rs10474485 A allele showed a significantly higher emotional state score than carriers.These results suggest that the CRH and CRH-BP genes have no direct effect on IBS status. However, the CRH-BP SNP rs10474485 has some effect on IBS-related emotional abnormalities and resistance to psychosocial stress.

  2. Modulation of endogenous Cysteine Protease Inhibitor (ICP) 1 expression in Entamoeba histolytica affects amoebic adhesion to Extracellular Matrix proteins.

    Science.gov (United States)

    Lee, Young Ah; Saito-Nakano, Yumiko; Kim, Kyeong Ah; Min, Arim; Nozaki, Tomoyoshi; Shin, Myeong Heon

    2015-02-01

    Entamoeba histolytica is an enteric tissue-invading protozoan parasite that causes amoebic colitis and occasionally liver abscess in humans. During tissue invasion, amoebic adhesion to host components is an important event for host cell death leading to successful invasion and infection. Among amoebic virulence factors, Gal/GalNAc lectin is known to be major adhesion factor to host cells. In this study, we investigated the role of amoebic secreted CP (Cysteine Proteases) in amoebic adhesion to extracellular matrix (ECM) protein using CP inhibitor and E. histolytica strains in which the endogenous inhibitor of cysteine protease (ICP) 1 gene was overexpressed (ICP1(+)) or repressed by antisense small RNA-mediated gene silencing (ICP1(-)). We found that pretreatment of wild-type amoebae with CP inhibitor E64, or thiol-group modifiers such as diamide and N-Ethylmaleimide resulted in a significant decrease in adhesion to laminin and collagen ECM proteins. Furthermore, ICP1(+) strain, with a reduction of secreted CP activity, exhibited reduced ability by 40% to adhere to laminin. In contrast, ICP1(-) strain, with a 1.9-fold increase of secreted CP activity, showed a two-fold increase in amoebic adherence to laminin compared to the control strain. In addition, total amount of secreted CP5 was decreased in ICP1(+) amoeba. Conversely, total amount of secreted CP1 and mature-form CP5 were increased in ICP1(-) amoeba. We also found that ICP1 was secreted into extracellular milieu. These results suggest that secreted CP activity by E. histolytica may be an important factor affecting adhesion to host proteins, and regulation of CP secretion by ICP plays a major role in pathogenesis. This study provides insight into the CP-mediated tissue pathogenesis in amoeba-invaded lesions during human amoebiasis. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Dorsal and ventral striatal protein synthesis inhibition affect reinforcer valuation but not the consolidation of instrumental learning.

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    Jonkman, Sietse; Everitt, Barry J

    2011-10-01

    The evidence for a role of the striatum in the acquisition of uncued instrumental responding is ambiguous. It has been shown that post-session infusions of anisomycin into the core of the nucleus accumbens (NAcc) impaired instrumental acquisition, but pre-training lesions of the NAcc suggest that it is not necessary. Recently, we demonstrated that the infusion of anisomycin into the anterior cingulate cortex impaired instrumental acquisition indirectly through a taste aversion. Thus, we hypothesized that post-session anisomycin infusions into the NAcc affected instrumental acquisition through an effect on reinforcer valuation. For the dorsal striatum, both post-session infusions of anisomycin and pre-training lesion studies suggest that neither the dorsolateral nor the dorsomedial striatum is necessary for the acquisition of instrumental responding. However, it has not been attempted to block plasticity in both regions concurrently, and we hypothesized that both regions independently contribute to acquisition through goal-directed and habitual learning. In the current experiments, we first replicated the effect of unprotected post-session anisomycin infusions into the NAcc on instrumental acquisition. Subsequently, we investigated the effect of protein synthesis inhibition in the NAcc and dorsomedial and dorsolateral striatum concurrently on instrumental acquisition, critically controlling for effects on reinforcer valuation. The anisomycin infusions induced an aversive state, but did not affect instrumental acquisition.

  4. Novel Role of Parathyroid Hormone-Related Protein in the Pathophysiology of the Diabetic Kidney: Evidence from Experimental and Human Diabetic Nephropathy

    National Research Council Canada - National Science Library

    Romero, Montserrat; Ortega, Arantxa; Olea, Nuria; Arenas, María Isabel; Izquierdo, Adriana; Bover, Jordi; Esbrit, Pedro; Bosch, Ricardo J

    2013-01-01

    ... to glomerular diseases, such as diabetic nephropathy (DN). In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria...

  5. Cardiac myosin binding protein C phosphorylation affects cross-bridge cycle's elementary steps in a site-specific manner.

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    Li Wang

    Full Text Available Based on our recent finding that cardiac myosin binding protein C (cMyBP-C phosphorylation affects muscle contractility in a site-specific manner, we further studied the force per cross-bridge and the kinetic constants of the elementary steps in the six-state cross-bridge model in cMyBP-C mutated transgenic mice for better understanding of the influence of cMyBP-C phosphorylation on contractile functions. Papillary muscle fibres were dissected from cMyBP-C mutated mice of ADA (Ala273-Asp282-Ala302, DAD (Asp273-Ala282-Asp302, SAS (Ser273-Ala282-Ser302, and t/t (cMyBP-C null genotypes, and the results were compared to transgenic mice expressing wide-type (WT cMyBP-C. Sinusoidal analyses were performed with serial concentrations of ATP, phosphate (Pi, and ADP. Both t/t and DAD mutants significantly reduced active tension, force per cross-bridge, apparent rate constant (2πc, and the rate constant of cross-bridge detachment. In contrast to the weakened ATP binding and enhanced Pi and ADP release steps in t/t mice, DAD mice showed a decreased ADP release without affecting the ATP binding and the Pi release. ADA showed decreased ADP release, and slightly increased ATP binding and cross-bridge detachment steps, whereas SAS diminished the ATP binding step and accelerated the ADP release step. t/t has the broadest effects with changes in most elementary steps of the cross-bridge cycle, DAD mimics t/t to a large extent, and ADA and SAS predominantly affect the nucleotide binding steps. We conclude that the reduced tension production in DAD and t/t is the result of reduced force per cross-bridge, instead of the less number of strongly attached cross-bridges. We further conclude that cMyBP-C is an allosteric activator of myosin to increase cross-bridge force, and its phosphorylation status modulates the force, which is regulated by variety of protein kinases.

  6. Diversification of the AlpB Outer Membrane Protein of Helicobacter pylori Affects Biofilm Formation and Cellular Adhesion.

    Science.gov (United States)

    Yonezawa, Hideo; Osaki, Takako; Fukutomi, Toshiyuki; Hanawa, Tomoko; Kurata, Satoshi; Zaman, Cynthia; Hojo, Fuhito; Kamiya, Shigeru

    2017-03-15

    Helicobacter pylori is one of the most common causes of bacterial infection in humans, and it forms biofilms on human gastric mucosal epithelium as well as on in vitro abiotic surfaces. Bacterial biofilm is critical not only for environmental survival but also for successful infection. We previously demonstrated that strain TK1402, which was isolated from a Japanese patient with duodenal and gastric ulcers, has high biofilm-forming ability in vitro relative to other strains. In addition, we showed that outer membrane vesicles (OMV) play an important role in biofilm formation. The aim of this study was to analyze which protein(s) in the OMV contributes to biofilm formation in TK1402. We obtained a spontaneous mutant strain derived from TK1402 lacking biofilm-forming ability. The protein profiles of the OMV were compared between this mutant strain and the wild type, and it was found that AlpB, an outer membrane protein in the OMV of the mutant strain, was markedly decreased compared to that of the wild type. Restoration of TK1402 alpB to the mutant strain fully recovered the ability to form biofilm. However, restoration with alpB from other strains demonstrated incomplete recovery of biofilm-forming ability. We therefore inferred that the variable region of AlpB (amino acid positions 121 to 146) was involved in TK1402 biofilm formation. In addition, diversification of the AlpB sequence was shown to affect the ability to adhere to AGS cells. These results demonstrate a new insight into the molecular mechanisms of host colonization by H. pyloriIMPORTANCE Bacterial biofilm is critical not only for environmental survival but also for successful infection. The mechanism of Helicobacter pylori adherence to host cells mediated by cell surface adhesins has been the focus of many studies, but little is known regarding factors involved in H. pylori biofilm formation. Our study demonstrated that AlpB plays an important role in biofilm formation and that this property depends

  7. Mixed compared with single-source proteins in high-protein diets affect kidney structure and function differentially in obese fa/fa Zucker rats.

    Science.gov (United States)

    Devassy, Jessay G; Wojcik, Jennifer L; Ibrahim, Naser H M; Zahradka, Peter; Taylor, Carla G; Aukema, Harold M

    2017-02-01

    Questions remain regarding the potential negative effects of dietary high protein (HP) on kidney health, particularly in the context of obesity in which the risk for renal disease is already increased. To examine whether some of the variability in HP effects on kidney health may be due to source of protein, obese fa/fa Zucker rats were given HP (35% of energy from protein) diets containing either casein, soy protein, or a mixed source of animal and plant proteins for 12 weeks. Control lean and obese rats were given diets containing casein at normal protein (15% of energy from protein) levels. Body weight and blood pressure were measured, and markers of renal structural changes, damage, and function were assessed. Obesity alone resulted in mild renal changes, as evidenced by higher kidney weights, proteinuria, and glomerular volumes. In obese rats, increasing the protein level using the single, but not mixed, protein sources resulted in higher renal fibrosis compared with the lean rats. The mixed-protein HP group also had lower levels of serum monocyte chemoattractant protein-1, even though this diet further increased kidney and glomerular size. Soy and mixed-protein HP diets also resulted in a small number of damaged glomeruli, while soy compared with mixed-protein HP diet delayed the increase in blood pressure over time. Since obesity itself confers added risk of renal disease, an HP diet from mixed-protein sources that enables weight loss but has fewer risks to renal health may be advantageous.

  8. Perinatal protein malnutrition affects mitochondrial function in adult and results in a resistance to high fat diet-induced obesity.

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    Céline Jousse

    Full Text Available Epidemiological findings indicate that transient environmental influences during perinatal life, especially nutrition, may have deleterious heritable health effects lasting for the entire life. Indeed, the fetal organism develops specific adaptations that permanently change its physiology/metabolism and that persist even in the absence of the stimulus that initiated them. This process is termed "nutritional programming". We previously demonstrated that mothers fed a Low-Protein-Diet (LPD during gestation and lactation give birth to F1-LPD animals presenting metabolic consequences that are different from those observed when the nutritional stress is applied during gestation only. Compared to control mice, adult F1-LPD animals have a lower body weight and exhibit a higher food intake suggesting that maternal protein under-nutrition during gestation and lactation affects the energy metabolism of F1-LPD offspring. In this study, we investigated the origin of this apparent energy wasting process in F1-LPD and demonstrated that minimal energy expenditure is increased, due to both an increased mitochondrial function in skeletal muscle and an increased mitochondrial density in White Adipose Tissue. Importantly, F1-LPD mice are protected against high-fat-diet-induced obesity. Clearly, different paradigms of exposure to malnutrition may be associated with differences in energy expenditure, food intake, weight and different susceptibilities to various symptoms associated with metabolic syndrome. Taken together these results demonstrate that intra-uterine environment is a major contributor to the future of individuals and disturbance at a critical period of development may compromise their health. Consequently, understanding the molecular mechanisms may give access to useful knowledge regarding the onset of metabolic diseases.

  9. Whole genome sequencing identifies a deletion in protein phosphatase 2A that affects its stability and localization in Chlamydomonas reinhardtii.

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    Huawen Lin

    Full Text Available Whole genome sequencing is a powerful tool in the discovery of single nucleotide polymorphisms (SNPs and small insertions/deletions (indels among mutant strains, which simplifies forward genetics approaches. However, identification of the causative mutation among a large number of non-causative SNPs in a mutant strain remains a big challenge. In the unicellular biflagellate green alga Chlamydomonas reinhardtii, we generated a SNP/indel library that contains over 2 million polymorphisms from four wild-type strains, one highly polymorphic strain that is frequently used in meiotic mapping, ten mutant strains that have flagellar assembly or motility defects, and one mutant strain, imp3, which has a mating defect. A comparison of polymorphisms in the imp3 strain and the other 15 strains allowed us to identify a deletion of the last three amino acids, Y313F314L315, in a protein phosphatase 2A catalytic subunit (PP2A3 in the imp3 strain. Introduction of a wild-type HA-tagged PP2A3 rescues the mutant phenotype, but mutant HA-PP2A3 at Y313 or L315 fail to rescue. Our immunoprecipitation results indicate that the Y313, L315, or YFLΔ mutations do not affect the binding of PP2A3 to the scaffold subunit, PP2A-2r. In contrast, the Y313, L315, or YFLΔ mutations affect both the stability and the localization of PP2A3. The PP2A3 protein is less abundant in these mutants and fails to accumulate in the basal body area as observed in transformants with either wild-type HA-PP2A3 or a HA-PP2A3 with a V310T change. The accumulation of HA-PP2A3 in the basal body region disappears in mated dikaryons, which suggests that the localization of PP2A3 may be essential to the mating process. Overall, our results demonstrate that the terminal YFL tail of PP2A3 is important in the regulation on Chlamydomonas mating.

  10. Iron-Restricted Diet Affects Brain Ferritin Levels, Dopamine Metabolism and Cellular Prion Protein in a Region-Specific Manner

    Science.gov (United States)

    Pino, Jessica M. V.; da Luz, Marcio H. M.; Antunes, Hanna K. M.; Giampá, Sara Q. de Campos; Martins, Vilma R.; Lee, Kil S.

    2017-01-01

    Iron is an essential micronutrient for several physiological functions, including the regulation of dopaminergic neurotransmission. On the other hand, both iron, and dopamine can affect the folding and aggregation of proteins related with neurodegenerative diseases, such as cellular prion protein (PrPC) and α-synuclein, suggesting that deregulation of iron homeostasis and the consequential disturbance of dopamine metabolism can be a risk factor for conformational diseases. These proteins, in turn, are known to participate in the regulation of iron and dopamine metabolism. In this study, we evaluated the effects of dietary iron restriction on brain ferritin levels, dopamine metabolism, and the expression levels of PrPC and α-synuclein. To achieve this goal, C57BL/6 mice were fed with iron restricted diet (IR) or with normal diet (CTL) for 1 month. IR reduced iron and ferritin levels in liver. Ferritin reduction was also observed in the hippocampus. However, in the striatum of IR group, ferritin level was increased, suggesting that under iron-deficient condition, each brain area might acquire distinct capacity to store iron. Increased lipid peroxidation was observed only in hippocampus of IR group, where ferritin level was reduced. IR also generated discrete results regarding dopamine metabolism of distinct brain regions: in striatum, the level of dopamine metabolites (DOPAC and HVA) was reduced; in prefrontal cortex, only HVA was increased along with the enhanced MAO-A activity; in hippocampus, no alterations were observed. PrPC levels were increased only in the striatum of IR group, where ferritin level was also increased. PrPC is known to play roles in iron uptake. Thus, the increase of PrPC in striatum of IR group might be related to the increased ferritin level. α-synuclein was not altered in any regions. Abnormal accumulation of ferritin, increased MAO-A activity or lipid peroxidation are molecular features observed in several neurological disorders. Our

  11. Iron-Restricted Diet Affects Brain Ferritin Levels, Dopamine Metabolism and Cellular Prion Protein in a Region-Specific Manner

    Directory of Open Access Journals (Sweden)

    Jessica M. V. Pino

    2017-05-01

    Full Text Available Iron is an essential micronutrient for several physiological functions, including the regulation of dopaminergic neurotransmission. On the other hand, both iron, and dopamine can affect the folding and aggregation of proteins related with neurodegenerative diseases, such as cellular prion protein (PrPC and α-synuclein, suggesting that deregulation of iron homeostasis and the consequential disturbance of dopamine metabolism can be a risk factor for conformational diseases. These proteins, in turn, are known to participate in the regulation of iron and dopamine metabolism. In this study, we evaluated the effects of dietary iron restriction on brain ferritin levels, dopamine metabolism, and the expression levels of PrPC and α-synuclein. To achieve this goal, C57BL/6 mice were fed with iron restricted diet (IR or with normal diet (CTL for 1 month. IR reduced iron and ferritin levels in liver. Ferritin reduction was also observed in the hippocampus. However, in the striatum of IR group, ferritin level was increased, suggesting that under iron-deficient condition, each brain area might acquire distinct capacity to store iron. Increased lipid peroxidation was observed only in hippocampus of IR group, where ferritin level was reduced. IR also generated discrete results regarding dopamine metabolism of distinct brain regions: in striatum, the level of dopamine metabolites (DOPAC and HVA was reduced; in prefrontal cortex, only HVA was increased along with the enhanced MAO-A activity; in hippocampus, no alterations were observed. PrPC levels were increased only in the striatum of IR group, where ferritin level was also increased. PrPC is known to play roles in iron uptake. Thus, the increase of PrPC in striatum of IR group might be related to the increased ferritin level. α-synuclein was not altered in any regions. Abnormal accumulation of ferritin, increased MAO-A activity or lipid peroxidation are molecular features observed in several neurological

  12. Apolipoprotein E4 (1–272 fragment is associated with mitochondrial proteins and affects mitochondrial function in neuronal cells

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    Michikawa Makoto

    2009-08-01

    Full Text Available Abstract Background Apolipoprotein E allele ε4 (apoE4 is a strong risk factor for developing Alzheimer's disease (AD. Secreted apoE has a critical function in redistributing lipids among central nervous system cells to maintain normal lipid homeostasis. In addition, previous reports have shown that apoE4 is cleaved by a protease in neurons to generate apoE4(1–272 fragment, which is associated with neurofibrillary tanglelike structures and mitochondria, causing mitochondrial dysfunction. However, it still remains unclear how the apoE fragment associates with mitochondria and induces mitochondrial dysfunction. Results To clarify the molecular mechanism, we carried out experiments to identify intracellular apoE-binding molecules and their functions in modulating mitochondria function. Here, we found that apoE4 binds to ubiquinol cytochrome c reductase core protein 2 (UQCRC2 and cytochrome C1, both of which are components of mitochondrial respiratory complex III, and cytochrome c oxidase subunit 4 isoform 1 (COX IV 1, which is a component of complex IV, in Neuro-2a cells. Interestingly, these proteins associated with apoE4(1–272 more strongly than intact apoE4(1–299. Further analysis showed that in Neuro-2a cells expressing apoE4(1–272, the enzymatic activities of mitochondrial respiratory complexes III and IV were significantly lower than those in Neuro-2a cells expressing apoE4(1–299. Conclusion ApoE4(1–272 fragment expressed in Neuro2a cells is associated with mitochondrial proteins, UQCRC2 and cytochrome C1, which are component of respiratory complex III, and with COX IV 1, which is a member of complex IV. Overexpression of apoE4(1–272 fragment impairs activities of complex III and IV. These results suggest that the C-terminal-truncated fragment of apoE4 binds to mitochondrial complexes and affects their activities, and thereby leading to neurodegeneration.

  13. Breast cancer resistance protein (BCRP/ABCG2) transports fluoroquinolone antibiotics and affects their oral availability, pharmacokinetics, and milk secretion.

    Science.gov (United States)

    Merino, Gracia; Alvarez, Ana I; Pulido, Mivis M; Molina, Antonio J; Schinkel, Alfred H; Prieto, Julio G

    2006-04-01

    The breast cancer resistance protein (BCRP/ABCG2) is an ATP-binding cassette drug efflux transporter that extrudes xenotoxins from cells in intestine, liver, mammary gland, and other organs, affecting the pharmacological and toxicological behavior of many compounds, including their secretion into the milk. The purpose of this study was to determine whether three widely used fluoroquinolone antibiotics (ciprofloxacin, ofloxacin, and norfloxacin) are substrates of Bcrp1/BCRP and to investigate the possible role of this transporter in the in vivo pharmacokinetic profile of these compounds and their secretion into the milk. Using polarized cell lines, we found that ciprofloxacin, ofloxacin, and norfloxacin are transported by mouse Bcrp1 and human BCRP. In vivo pharmacokinetic studies showed that the ciprofloxacin plasma concentration was more than 2-fold increased in Bcrp1(-/-) compared with wild-type mice (1.77 +/- 0.73 versus 0.85 +/- 0.39 microg/ml, p milk concentration and milk/plasma ratio of ciprofloxacin were 2-fold higher in wild-type than in Bcrp1(-/-) lactating mice. We conclude that Bcrp1 is one of the determinants for the bioavailability of fluoroquinolones and their secretion into the milk.

  14. Monoclonal antibodies against accumulation-associated protein affect EPS biosynthesis and enhance bacterial accumulation of Staphylococcus epidermidis.

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    Jian Hu

    Full Text Available Because there is no effective antibiotic to eradicate Staphylococcus epidermidis biofilm infections that lead to the failure of medical device implantations, the development of anti-biofilm vaccines is necessary. Biofilm formation by S. epidermidis requires accumulation-associated protein (Aap that contains sequence repeats known as G5 domains, which are responsible for the Zn(2+-dependent dimerization of Aap to mediate intercellular adhesion. Antibodies against Aap have been reported to inhibit biofilm accumulation. In the present study, three monoclonal antibodies (MAbs against the Aap C-terminal single B-repeat construct followed by the 79-aa half repeat (AapBrpt1.5 were generated. MAb(18B6 inhibited biofilm formation by S. epidermidis RP62A to 60% of the maximum, while MAb(25C11 and MAb(20B9 enhanced biofilm accumulation. All three MAbs aggregated the planktonic bacteria to form visible cell clusters. Epitope mapping revealed that the epitope of MAb(18B6, which recognizes an identical area within AapBrpt constructs from S. epidermidis RP62A, was not shared by MAb(25C11 and MAb(20B9. Furthermore, all three MAbs were found to affect both Aap expression and extracellular polymeric substance (EPS, including extracellular DNA and PIA biosynthesis in S. epidermidis and enhance the cell accumulation. These findings contribute to a better understanding of staphylococcal biofilm formation and will help to develop epitope-peptide vaccines against staphylococcal infections.

  15. Protein phosphatase 2A holoenzyme is targeted to peroxisomes by piggybacking and positively affects peroxisomal β-oxidation.

    Science.gov (United States)

    Kataya, Amr R A; Heidari, Behzad; Hagen, Lars; Kommedal, Roald; Slupphaug, Geir; Lillo, Cathrine

    2015-02-01

    The eukaryotic, highly conserved serine (Ser)/threonine-specific protein phosphatase 2A (PP2A) functions as a heterotrimeric complex composed of a catalytic (C), scaffolding (A), and regulatory (B) subunit. In Arabidopsis (Arabidopsis thaliana), five, three, and 17 genes encode different C, A, and B subunits, respectively. We previously found that a B subunit, B'θ, localized to peroxisomes due to its C-terminal targeting signal Ser-Ser-leucine. This work shows that PP2A C2, C5, andA2 subunits interact and colocalize with B'θ in peroxisomes. C and A subunits lack peroxisomal targeting signals, and their peroxisomal import depends on B'θ and appears to occur by piggybacking transport. B'θ knockout mutants were impaired in peroxisomal β-oxidation as shown by developmental arrest of seedlings germinated without sucrose, accumulation of eicosenoic acid, and resistance to protoauxins indole-butyric acid and 2,4-dichlorophenoxybutyric acid. All of these observations strongly substantiate that a full PP2A complex is present in peroxisomes and positively affects β-oxidation of fatty acids and protoauxins. © 2015 American Society of Plant Biologists. All Rights Reserved.

  16. Protein Phosphatase 2A Holoenzyme Is Targeted to Peroxisomes by Piggybacking and Positively Affects Peroxisomal β-Oxidation1[OPEN

    Science.gov (United States)

    Kataya, Amr R.A.; Heidari, Behzad; Hagen, Lars; Kommedal, Roald; Slupphaug, Geir; Lillo, Cathrine

    2015-01-01

    The eukaryotic, highly conserved serine (Ser)/threonine-specific protein phosphatase 2A (PP2A) functions as a heterotrimeric complex composed of a catalytic (C), scaffolding (A), and regulatory (B) subunit. In Arabidopsis (Arabidopsis thaliana), five, three, and 17 genes encode different C, A, and B subunits, respectively. We previously found that a B subunit, B′θ, localized to peroxisomes due to its C-terminal targeting signal Ser-Ser-leucine. This work shows that PP2A C2, C5, andA2 subunits interact and colocalize with B′θ in peroxisomes. C and A subunits lack peroxisomal targeting signals, and their peroxisomal import depends on B′θ and appears to occur by piggybacking transport. B′θ knockout mutants were impaired in peroxisomal β-oxidation as shown by developmental arrest of seedlings germinated without sucrose, accumulation of eicosenoic acid, and resistance to protoauxins indole-butyric acid and 2,4-dichlorophenoxybutyric acid. All of these observations strongly substantiate that a full PP2A complex is present in peroxisomes and positively affects β-oxidation of fatty acids and protoauxins. PMID:25489022

  17. Down-Regulation of Small Rubber Particle Protein Expression Affects Integrity of Rubber Particles and Rubber Content in Taraxacum brevicorniculatum

    Science.gov (United States)

    Hillebrand, Andrea; Post, Janina J.; Wurbs, David; Wahler, Daniela; Lenders, Malte; Krzyzanek, Vladislav; Prüfer, Dirk; Gronover, Christian Schulze

    2012-01-01

    The biosynthesis of rubber is thought to take place on the surface of rubber particles in laticifers, highly specialized cells that are present in more than 40 plant families. The small rubber particle protein (SRPP) has been supposed to be involved in rubber biosynthesis, and recently five SRPPs (TbSRPP1–5) were identified in the rubber-producing dandelion species Taraxacum brevicorniculatum. Here, we demonstrate by immunogold labeling that TbSRPPs are localized to rubber particles, and that rubber particles mainly consist of TbSRPP3, 4 and 5 as shown by high-resolution two-dimensional gel electrophoresis and mass spectrometric analysis. We also carried out an RNA-interference approach in transgenic plants to address the function of TbSRPPs in rubber biosynthesis as well as rubber particle morphology and stability. TbSRPP-RNAi transgenic T. brevicorniculatum plants showed a 40–50% reduction in the dry rubber content, but neither the rubber weight average molecular mass nor the polydispersity of the rubber were affected. Although no phenotypical differences to wild-type particles could be observed in vivo, rubber particles from the TbSRPP-RNAi transgenic lines were less stable and tend to rapidly aggregate in expelling latex after wounding of laticifers. Our results prove that TbSRPPs are very crucial for rubber production in T. brevicorniculatum, probably by contributing to a most favourable and stable rubber particle architecture for efficient rubber biosynthesis and eventually storage. PMID:22911861

  18. Down-regulation of small rubber particle protein expression affects integrity of rubber particles and rubber content in Taraxacum brevicorniculatum.

    Directory of Open Access Journals (Sweden)

    Andrea Hillebrand

    Full Text Available The biosynthesis of rubber is thought to take place on the surface of rubber particles in laticifers, highly specialized cells that are present in more than 40 plant families. The small rubber particle protein (SRPP has been supposed to be involved in rubber biosynthesis, and recently five SRPPs (TbSRPP1-5 were identified in the rubber-producing dandelion species Taraxacum brevicorniculatum. Here, we demonstrate by immunogold labeling that TbSRPPs are localized to rubber particles, and that rubber particles mainly consist of TbSRPP3, 4 and 5 as shown by high-resolution two-dimensional gel electrophoresis and mass spectrometric analysis. We also carried out an RNA-interference approach in transgenic plants to address the function of TbSRPPs in rubber biosynthesis as well as rubber particle morphology and stability. TbSRPP-RNAi transgenic T. brevicorniculatum plants showed a 40-50% reduction in the dry rubber content, but neither the rubber weight average molecular mass nor the polydispersity of the rubber were affected. Although no phenotypical differences to wild-type particles could be observed in vivo, rubber particles from the TbSRPP-RNAi transgenic lines were less stable and tend to rapidly aggregate in expelling latex after wounding of laticifers. Our results prove that TbSRPPs are very crucial for rubber production in T. brevicorniculatum, probably by contributing to a most favourable and stable rubber particle architecture for efficient rubber biosynthesis and eventually storage.

  19. Gum Arabic Reduces C-Reactive Protein in Chronic Kidney Disease Patients without Affecting Urea or Indoxyl Sulfate Levels

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    Sarra Elamin

    2017-01-01

    Full Text Available Introduction. Gum Arabic (GA is a complex polysaccharide with proven prebiotic properties and potentially beneficial systemic effects. Methods. We randomly allocated 36 chronic kidney disease (CKD patients to receive 10, 20, or 40 grams daily of GA for four weeks and studied the systemic effects of this intervention. Results. Thirty participants completed the study with baseline glomerular filtration rate 29.1±9.9 mL/min/1.7 m2. In contrast to previous observations, we found no effect on serum urea or creatinine levels. GA supplementation was associated with a small but statistically significant drop in serum sodium level (138±2 to 136±3 mmol/L, p = 0.002 without affecting other electrolytes, urine volume, or indoxyl sulfate (IS levels. GA supplementation was also associated with a significant drop in C-reactive protein (CRP level (3.5±1.5 to 2.8±1.6 ng/mL, p = 0.02 even in patients who received only 10 g/day (4.4±1.2 to 3.2±1.5 ng/mL, p = 0.03. Conclusions. Supplementing the diet of CKD patients with 10–40 g/day of GA significantly reduced CRP level which could have a positive impact on these patients’ morbidity and mortality. This trial is registered with Saudi Clinical Trial Registry number 15011402.

  20. The Smc5-Smc6 complex regulates recombination at centromeric regions and affects kinetochore protein sumoylation during normal growth.

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    Vladimir Yong-Gonzales

    Full Text Available The Smc5-Smc6 complex in Saccharomyces cerevisiae is both essential for growth and important for coping with genotoxic stress. While it facilitates damage tolerance throughout the genome under genotoxin treatment, its function during unperturbed growth is mainly documented for repetitive DNA sequence maintenance. Here we provide physical and genetic evidence showing that the Smc5-Smc6 complex regulates recombination at non-repetitive loci such as centromeres in the absence of DNA damaging agents. Mutating Smc6 results in the accumulation of recombination intermediates at centromeres and other unique sequences as assayed by 2D gel analysis. In addition, smc6 mutant cells exhibit increased levels of Rad52 foci that co-localize with centromere markers. A rad52 mutation that decreases centromeric, but not overall, levels of Rad52 foci in smc6 mutants suppresses the nocodazole sensitivity of these cells, suggesting that the Smc6-mediated regulation of recombination at centromeric regions impacts centromere-related functions. In addition to influencing recombination, the SUMO ligase subunit of the Smc5-Smc6 complex promotes the sumoylation of two kinetochore proteins and affects mitotic spindles. These results suggest that the Smc5-Smc6 complex regulates both recombination and kinetochore sumoylation to facilitate chromosomal maintenance during growth.

  1. Loss of the Arabidopsis Protein Kinases ANPs Affects Root Cell Wall Composition, and Triggers the Cell Wall Damage Syndrome

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    Nora Gigli Bisceglia

    2018-01-01

    Full Text Available The Arabidopsis NPK1-related Protein kinases ANP1, ANP2 and ANP3 belong to the MAP kinase kinase kinase (MAPKKK superfamily and were previously described to be crucial for cytokinesis, elicitor-induced immunity and development. Here we investigate the basis of their role in development by using conditional β-estradiol-inducible triple mutants to overcome lethality. In seedlings, lack of ANPs causes root cell bulging, with the transition zone being the most sensitive region. We uncover a role of ANPs in the regulation of cell wall composition and suggest that developmental defects of the triple mutants, observed at the cellular level, might be a consequence of the alterations of the pectic and cellulosic cell wall components. Lack of ANPs also induced a typical cell wall damage syndrome (CWDS similar to that observed in plants treated with the cellulose biosynthesis inhibitor isoxaben (ISX. Moreover, anp double mutants and plants overexpressing single ANPs (ANP1 or ANP3 respectively showed increased and reduced accumulation of jasmonic acid and PDF1.2 transcripts upon ISX treatment, suggesting that ANPs are part of the pathway targeted by this inhibitor and play a role in cell wall integrity surveillance.Highlights: The loss of ANP function affects cell wall composition and leads to typical cell wall damage-induced phenotypes, such as ectopic lignification and jasmonic acid accumulation.

  2. ERK1 and ERK2 mitogen-activated protein kinases affect Ras-dependent cell signaling differentially

    Directory of Open Access Journals (Sweden)

    Bonini Chiara

    2006-06-01

    Full Text Available Abstract Background The mitogen-activated protein (MAP kinases p44ERK1 and p42ERK2 are crucial components of the regulatory machinery underlying normal and malignant cell proliferation. A currently accepted model maintains that ERK1 and ERK2 are regulated similarly and contribute to intracellular signaling by phosphorylating a largely common subset of substrates, both in the cytosol and in the nucleus. Results Here, we show that ablation of ERK1 in mouse embryo fibroblasts and NIH 3T3 cells by gene targeting and RNA interference results in an enhancement of ERK2-dependent signaling and in a significant growth advantage. By contrast, knockdown of ERK2 almost completely abolishes normal and Ras-dependent cell proliferation. Ectopic expression of ERK1 but not of ERK2 in NIH 3T3 cells inhibits oncogenic Ras-mediated proliferation and colony formation. These phenotypes are independent of the kinase activity of ERK1, as expression of a catalytically inactive form of ERK1 is equally effective. Finally, ectopic expression of ERK1 but not ERK2 is sufficient to attenuate Ras-dependent tumor formation in nude mice. Conclusion These results reveal an unexpected interplay between ERK1 and ERK2 in transducing Ras-dependent cell signaling and proliferation. Whereas ERK2 seems to have a positive role in controlling normal and Ras-dependent cell proliferation, ERK1 probably affects the overall signaling output of the cell by antagonizing ERK2 activity.

  3. Assembly of presynaptic filaments. Factors affecting the assembly of RecA protein onto single-stranded DNA

    DEFF Research Database (Denmark)

    Thresher, RJ; Christiansen, Gunna; Griffith, JD

    1988-01-01

    We have previously shown that the assembly of RecA protein onto single-stranded DNA (ssDNA) facilitated by SSB protein occurs in three steps: (1) rapid binding of SSB protein to the ssDNA; (2) nucleation of RecA protein onto this template; and (3) co-operative polymerization of additional Rec......A protein to yield presynaptic filaments. Here, electron microscopy has been used to further explore the parameters of this assembly process. The optimal extent of presynaptic filament formation required at least one RecA protein monomer per three nucleotides, high concentrations of ATP (greater than 3 m......M in the presence of 12 mM-Mg2+), and relatively low concentrations of SSB protein (1 monomer per 18 nucleotides). Assembly was depressed threefold when SSB protein was added to one monomer per nine nucleotides. These effects appeared to be exerted at the nucleation step. Following nucleation, RecA protein...

  4. Dietary exposure to polybrominated diphenyl ether 47 (BDE-47) inhibits development and alters thyroid hormone-related gene expression in the brain of Xenopus laevis tadpoles.

    Science.gov (United States)

    Yost, Alexandra T; Thornton, Leah M; Venables, Barney J; Sellin Jeffries, Marlo K

    2016-12-01

    Few studies have investigated the thyroid-disrupting effects of polybrominated diphenyl ethers (PBDEs) across multiple levels of biological organization in anurans, despite their suitability for the screening of thyroid disruptors. Therefore, the present study evaluated the effects of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) on development, thyroid histology and thyroid hormone-related gene expression in Xenopus laevis exposed to 0 (control), 50 (low), 500 (medium) or 5000μg BDE-47/g food (high) for 21days. Only the high dose of BDE-47 hindered growth and development; however, thyroid hormone-associated gene expression was downregulated in the brains of tadpoles regardless of dose. These results show that BDE-47 disrupts thyroid hormone signaling at the molecular and whole-organism levels and suggest that gene expression in the brain is a more sensitive endpoint than metamorphosis. Furthermore, the altered gene expression patterns among BDE-47-exposed tadpoles provide insight into the mechanisms of PBDE-induced thyroid disruption and highlight the potential for PBDEs to act as neurodevelopmental toxicants. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Missense mutations in the NF2 gene result in the quantitative loss of merlin protein and minimally affect protein intrinsic function.

    Science.gov (United States)

    Yang, Chunzhang; Asthagiri, Ashok R; Iyer, Rajiv R; Lu, Jie; Xu, David S; Ksendzovsky, Alexander; Brady, Roscoe O; Zhuang, Zhengping; Lonser, Russell R

    2011-03-22

    Neurofibromatosis type 2 (NF2) is a multiple neoplasia syndrome and is caused by a mutation of the NF2 tumor suppressor gene that encodes for the tumor suppressor protein merlin. Biallelic NF2 gene inactivation results in the development of central nervous system tumors, including schwannomas, meningiomas, ependymomas, and astrocytomas. Although a wide variety of missense germline mutations in the coding sequences of the NF2 gene can cause loss of merlin function, the mechanism of this functional loss is unknown. To gain insight into the mechanisms underlying loss of merlin function in NF2, we investigated mutated merlin homeostasis and function in NF2-associated tumors and cell lines. Quantitative protein and RT-PCR analysis revealed that whereas merlin protein expression was significantly reduced in NF2-associated tumors, mRNA expression levels were unchanged. Transfection of genetic constructs of common NF2 missense mutations into NF2 gene-deficient meningioma cell lines revealed that merlin loss of function is due to a reduction in mutant protein half-life and increased protein degradation. Transfection analysis also demonstrated that recovery of tumor suppressor protein function is possible, indicating that these mutants maintain intrinsic functional capacity. Further, increased expression of mutant protein is possible after treatment with specific proteostasis regulators, implicating protein quality control systems in the degradative fate of mutant tumor suppressor proteins. These findings provide direct insight into protein function and tumorigenesis in NF2 and indicate a unique treatment paradigm for this disorder.

  6. Effect of Protein-Lipid-Salt Interactions on Sodium Availability in the Mouth and Consequent Perception of Saltiness: As Affected by Hydration in Powders.

    Science.gov (United States)

    Yucel, Umut; Peterson, Devin G

    2015-09-02

    There is a broad need to reformulate lower sodium food products without affecting their original taste. The present study focuses on characterizing the role of protein-salt interactions on the salt release in low-moisture systems and saltiness perception during hydration. Sodium release from freeze-dried protein powders and emulsion powders formulated at different protein/lipid ratios (5:0 to 1:4) were characterized using a chromatography column modified with a porcine tongue. Emulsion systems with protein structured at the interface were found to have faster initial sodium release rates and faster hydration and were perceived to have a higher initial salt intensity with a lower salty aftertaste. In summary, exposure of the hydrophilic segments of the interface-structured proteins in emulsions was suggested to facilitate hydration and release of sodium during dissolution of low-moisture powder samples.

  7. Identification of naturally occurring amino acid variations that affect the ability of the measles virus C protein to regulate genome replication and transcription.

    Science.gov (United States)

    Bankamp, Bettina; Wilson, Jenna; Bellini, William J; Rota, Paul A

    2005-05-25

    The C protein of measles virus (MV C) is a basic protein of 186 amino acids (aa) that plays at least two roles in infected cells, interference with the innate immune response and modulation of viral polymerase activity. In this study, Northern blots were used to demonstrate that C proteins from three vaccine strains and three wild-type isolates of MV downregulated both mRNA transcription and genome replication in a plasmid-based mini-genome assay. The effect on transcription always paralleled the effect on replication; however, the six MV C proteins varied considerably in their ability to inhibit polymerase activity. Though the amino-terminal 45 aa of the C protein are more variable among different MV strains than the remaining 75% of the protein, the ability of the MV C proteins to inhibit polymerase activity was not regulated by substitutions in the amino terminus, but rather by the more conserved region containing aa 46-167. Naturally occurring substitutions at positions 147 and 166, but not 88 and 186, were found to regulate MV C protein activity. Deletion of the carboxyl-terminal 19 aa did not affect the polymerase-modulating activity. Though we did not find a link between the aa changes in MV C and attenuation, these data provide new information regarding the functions of this non-structural protein.

  8. Intentional formation of a protein corona on nanoparticles: Serum concentration affects protein corona mass, surface charge, and nanoparticle-cell interaction.

    Science.gov (United States)

    Gräfe, Christine; Weidner, Andreas; Lühe, Moritz V D; Bergemann, Christian; Schacher, Felix H; Clement, Joachim H; Dutz, Silvio

    2016-06-01

    The protein corona, which immediately is formed after contact of nanoparticles and biological systems, plays a crucial role for the biological fate of nanoparticles. In the here presented study we describe a strategy to control the amount of corona proteins which bind on particle surface and the impact of such a protein corona on particle-cell interactions. For corona formation, polyethyleneimine (PEI) coated magnetic nanoparticles (MNP) were incubated in a medium consisting of fetal calf serum (FCS) and cell culture medium. To modulate the amount of proteins bind to particles, the composition of the incubation medium was varied with regard to the FCS content. The protein corona mass was estimated and the size distribution of the participating proteins was determined by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Additionally, the zeta potential of incubated particles was measured. Human blood-brain barrier-representing cell line HBMEC was used for in vitro incubation experiments. To investigate the consequences of the FCS dependent protein corona formation on the interaction of MNP and cells flow cytometry and laser scanning microscopy were used. Zeta potential as well as SDS-PAGE clearly reveal an increase in the amount of corona proteins on MNP with increasing amount of FCS in incubation medium. For MNP incubated with lower FCS concentrations especially medium-sized proteins of molecular weights between 30kDa and 100kDa could be found within the protein corona, whereas for MNP incubated within higher FCS concentrations the fraction of corona proteins of 30kDa and less increased. The presence of the protein corona reduces the interaction of PEI-coated MNP with HBMEC cells within a 30min-incubation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Low protein provision during the first year of life, but not during foetal life, affects metabolic traits, organ mass development and growth in male mink (Neovison vison).

    Science.gov (United States)

    Vesterdorf, K; Blache, D; Harrison, A; Matthiesen, C F; Tauson, A-H

    2014-04-01

    Low protein provision in utero and post-partum may induce metabolic disorders in adulthood. Studies in mink have mainly focused on short-term consequences of low protein provision in utero whereas the long-term responses to low protein (LP) provision in metabolically programmed mink are unknown. We investigated whether low protein provision in utero affects the long-term response to adequate (AP) or LP provision after weaning in male mink. Eighty-six male mink were exposed to low (19% of ME from CP; crude protein) or adequate (31% of ME from CP) protein provision in utero, and to LP (~20% of ME from CP) or AP (30-42% of ME from CP) provision post-weaning. Being metabolically programmed by low protein provision in utero did not affect the response to post-weaning diets. Dietary protein content in the LP feed after weaning was below requirements; evidenced by lower nitrogen retention (p < 0.001) preventing LP mink from attaining their growth potential (p < 0.02). LP mink had a lower liver, pancreas and kidney weight (p < 0.05) as well as lower plasma IGF-1 concentrations at 8 and 25 (p < 0.05) weeks, and a higher incidence of hepatic lipidosis at 25 weeks (p < 0.05). Furthermore, LP mink had a higher body fat (p < 0.05) and lower body CP content (p < 0.05) at 50 weeks of age. It is concluded that some effects of low protein provision in utero can be alleviated by an adequate nutrient supply post-partum. However, long-term exposure to low protein provision in mink reduces their growth potential and induces transient hepatic lipidosis and modified body composition. Journal of Animal Physiology and Animal Nutrition © 2013 Blackwell Verlag GmbH.

  10. Type 2 diabetes mellitus interacts with obesity and common variations in PLTP to affect plasma phospholipid transfer protein activity.

    Science.gov (United States)

    Dullaart, R P F; Vergeer, M; de Vries, R; Kappelle, P J W H; Dallinga-Thie, G M

    2012-05-01

    Phospholipid transfer protein (PLTP) is an emerging cardiometabolic risk marker that is important in high-density lipoprotein (HDL) and triglyceride metabolism. Plasma PLTP activity is elevated in type 2 diabetes mellitus, whereas glucose may regulate PLTP gene transcription in vitro. Of interest, common PLTP variations that predict cardiovascular disease have been identified recently. We investigated whether the diabetic state is able to amplify relationships between obesity and PLTP gene variations with circulating PLTP levels. Plasma PLTP activity (using a phospholipid vesicles-HDL system), PLTP gene score [number of PLTP activity-decreasing alleles based on two tagging polymorphisms (rs378114 and rs60- 65904)] and waist circumference were determined in two Dutch cohorts comprising 237 patients with type 2 diabetes and 78 control subjects. Patients with diabetes were more obese (P diabetic and control subjects (P = 0.40). PLTP activity was highest in patients with diabetes with an enlarged waist and lowest in control subjects with a normal waist circumference (P Multiple linear regression analysis revealed a positive interaction between diabetes status and waist circumference on PLTP activity (β = 0.200, P = 0.005). Furthermore, diabetes status (β = -0.485, P = 0.046) or HbA1c (β = -0.240, P = 0.035) interacted with PLTP gene score to affect PLTP activity. Type 2 diabetes and enlarged waist circumference interact to impact on plasma PLTP activity. Diabetes may also amplify the association between plasma PLTP activity and common PLTP gene variations. Our findings support the hypothesis that diabetes-environment and diabetes-gene interactions govern plasma PLTP activity. © 2011 The Association for the Publication of the Journal of Internal Medicine.

  11. No evidence for involvement of plasma proteins or blood-borne cells in amyloid plaque formation in scrapie-affected mice. An immunohistoperoxidase study

    NARCIS (Netherlands)

    Eikelenboom, P.; Scott, J. R.; McBride, P. A.; Rozemuller, J. M.; Bruce, M. E.; Fraser, H.

    1987-01-01

    The present study was designed to investigate blood-brain permeability and the possible involvement of plasma proteins and blood-borne cells in amyloid plaque formation in scrapie-affected mice. No abnormal extravasation of intravenously injected horseradish peroxidase (HRP) was found and with

  12. Combined vitamin B-12 and balanced protein-energy supplementation affect homocysteine remethylation in the methionine cycle in pregnant south Indian women of low vitamin B-12 status

    Science.gov (United States)

    Low-quality dietary protein intake and vitamin B-12 deficiency could interact to decrease methionine transmethylation and remethylation rates during pregnancy, and may affect epigenetic modifications of the fetal genome. The objective of this randomized, partially open-labeled intervention trial was...

  13. Milk protein and fat play different roles in affecting the bioavailability and the antioxidant activity of jujube juice phenolics in rats.

    Science.gov (United States)

    Zhang, Hao; Zheng, Junyan; Liu, Xianting; Ding, Qingbo; Jiang, Lu; Guo, Huiyuan; Ren, Fazheng

    2012-10-01

    In this study, we intend to clarify the role that milk proteins and fat may play in affecting the bioavailability and the antioxidant activity of jujube juice phenolics. Three milk preparations-skimmed milk, milk fat, and whole milk were produced to represent milk protein, milk fat, and milk protein and fat together, respectively. The bioavailability of phenolics and the rat plasma antioxidant capacity were measured for 8 h after the consumption of jujube juice with and without milk preparations. The addition of skimmed milk to jujube juice resulted in significant changes in the plasma kinetics profile of phenolics, rather than affecting the overall absorption. Milk fat did not interact with jujube juice phenolics. However, when jujube juice was ingested with whole milk, a significant reduction of the bioavailability of phenolics and the maximum increase in plasma antioxidant capacity was observed. Moreover, a consistent increase in the median diameters of the emulsions indicated the formation of complexes of proteins, fat, and phenolics during digestion. The present study suggests that when ingested with jujube juice, milk proteins and fat play different roles in affecting the bioavailability and the antioxidant activity of jujube juice phenolics. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Elevated protein carbonylation and oxidative stress do not affect protein structure and function in the long-living naked-mole rat: a proteomic approach.

    Science.gov (United States)

    De Waal, Eric M; Liang, Hanyu; Pierce, Anson; Hamilton, Ryan T; Buffenstein, Rochelle; Chaudhuri, Asish R

    2013-05-17

    The 'oxidative stress theory of aging' predicts that aging is primarily regulated by progressive accumulation of oxidized macromolecules that cause deleterious effects to cellular homeostasis and induces a decline in physiological function. However, our reports on the detection of higher level of oxidized protein carbonyls in the soluble cellular fractions of long-living rodent naked-mole rats (NMRs, lifespan ~30yrs) compared to short-lived mice (lifespan ~3.5yrs) apparently contradicts a key tenet of the oxidative theory. As oxidation often inactivates enzyme function and induces higher-order soluble oligomers, we performed a comprehensive study to measure global protein carbonyl level in different tissues of age-matched NMRs and mice to determine if the traditional concept of oxidation mediated impairment of function and induction of higher-order structures of proteins are upheld in the NMRs. We made three intriguing observations with NMRs proteins: (1) protein carbonyl is significantly elevated across different tissues despite of its exceptional longevity, (2) enzyme function is restored despite of experiencing higher level of protein carbonylation, and (3) enzymes show lesser sensitivity to form higher-order non-reducible oligomers compared to short-living mouse proteins in response to oxidative stress. These observations were made based on the global analysis of protein carbonyl and identification of two heavily carbonylated proteins in the kidney, triosephosphate isomerase (TPI) and cytosolic peroxiredoxin (Prdx1). These un-expected intriguing observations thus strongly suggest that oxidative modification may not be the only criteria for impairment of protein and enzyme function; cellular environment is likely be the critical determining factor in this process and may be the underlying mechanism for exceptional longevity of NMR. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. G protein-coupled receptor 120 (GPR120) transcription in intestinal epithelial cells is significantly affected by bacteria belonging to the Bacteroides, Proteobacteria, and Firmicutes phyla

    DEFF Research Database (Denmark)

    Fredborg, Marlene; Theil, Peter Kappel; Jensen, Bent Borg

    2012-01-01

    Free fatty acids (FFA) are produced in the intestine by microbial fermentation. Recently, a family of G protein-coupled receptors (GPR) acting as FFA transporters has been reported including GPR120, which is expressed by intestinal epithelial cells. The GPR120 has been reported to affect the expr......Free fatty acids (FFA) are produced in the intestine by microbial fermentation. Recently, a family of G protein-coupled receptors (GPR) acting as FFA transporters has been reported including GPR120, which is expressed by intestinal epithelial cells. The GPR120 has been reported to affect...... the expression of glucagon-like peptide (GLP)-1 as well as function as a control point for anti-inflammatory effects. The aim of the present study was to evaluate whether 12 selected intestinal bacteria, representing the 4 major phyla present in the intestine, affect intestinal epithelial cell GPR120 and GLP-1 m...

  16. Systematic functional comparative analysis of four single-stranded DNA-binding proteins and their affection on viral RNA metabolism.

    Directory of Open Access Journals (Sweden)

    Haiyan Shi

    Full Text Available The accumulation of single-stranded DNA-binding (SSB proteins is essential for organisms and has various applications. However, no study has simultaneously and systematically compared the characteristics of SSB proteins. In addition, SSB proteins may bind RNA and play an unknown biological role in RNA metabolism. Here, we expressed a novel species of SSB protein derived from Thermococcus kodakarensis KOD1 (KOD, as well as SSB proteins from Thermus thermophilus (TTH, Escherichia coli, and Sulfolobus Solfataricus P2 (SSOB, abbreviated kod, tth, bl21, and ssob, respectively. These SSB proteins could bind ssDNA and viral RNA. bl21 resisted heat treatment for more than 9 h, Ssob and kod could withstand 95°C for 10 h and retain its ssDNA- and RNA-binding ability. Four SSB proteins promoted the specificity of the DNA polymerase in PCR-based 5- and 9-kb genome fragment amplification. kod also increased the amplification of a 13-kb PCR product, and SSB protein-bound RNA resisted Benzonase digestion. The SSB proteins could also enter the host cell bound to RNA, which resulted in modulation of viral RNA metabolism, particularly ssob and bl21.

  17. Associations of hormone-related factors with breast cancer risk according to hormone receptor status among white and African American women.

    Science.gov (United States)

    Cui, Yong; Deming-Halverson, Sandra L; Shrubsole, Martha J; Beeghly-Fadiel, Alicia; Fair, Alecia M; Sanderson, Maureen; Shu, Xiao-Ou; Kelley, Mark C; Zheng, Wei

    2014-12-01

    Causes of racial disparities in breast cancer incidence and mortality between white and African American women remain unclear. This study evaluated associations of menstrual and reproductive factors with breast cancer risk by race and cancer subtypes. Included in the study were 1866 breast cancer cases and 2306 controls recruited in the Nashville Breast Health Study, a population-based case-control study. Multivariable logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs). African American women were more likely to have estrogen receptor-negative (ER(-)), progesterone receptor-negative (PR(-)), and triple-negative (ER(-)PR(-)HER2(-)) breast cancer than white women. Age at menarche (≥ 14 years) and multiparity (≥ 3 live births) were inversely associated with ER(+) tumors only, whereas late age at first live birth (> 30 years) and nulliparity were associated with elevated risk; such associations were predominantly seen in white women (OR = 0.70, 95% CI = 0.55-0.88; OR = 0.72, 95% CI = 0.56-0.92; OR = 1.42, 95% CI = 1.13-1.79; OR = 1.32, 95% CI = 1.06-1.63, respectively). Age at menopause between 47 and 51 years was associated with elevated risk of ER(-) tumors in both white and African American women. Among women who had natural menopause, positive association between ever-use of hormone replacement therapy and breast cancer risk was seen in white women only (OR = 1.39, 95% CI = 1.03-1.87). This study suggests that certain hormone-related factors are differentially associated with risk of breast cancer subtypes, and these associations also differ by race. Copyright © 2014 Elsevier Inc. All rights reserved.

  18. Modulation of protein fermentation does not affect fecal water toxicity: a randomized cross-over study in healthy subjects.

    Directory of Open Access Journals (Sweden)

    Karen Windey

    Full Text Available Protein fermentation results in production of metabolites such as ammonia, amines and indolic, phenolic and sulfur-containing compounds. In vitro studies suggest that these metabolites might be toxic. However, human and animal studies do not consistently support these findings. We modified protein fermentation in healthy subjects to assess the effects on colonic metabolism and parameters of gut health, and to identify metabolites associated with toxicity.After a 2-week run-in period with normal protein intake (NP, 20 healthy subjects followed an isocaloric high protein (HP and low protein (LP diet for 2 weeks in a cross-over design. Protein fermentation was estimated from urinary p-cresol excretion. Fecal metabolite profiles were analyzed using GC-MS and compared using cluster analysis. DGGE was used to analyze microbiota composition. Fecal water genotoxicity and cytotoxicity were determined using the Comet assay and the WST-1-assay, respectively, and were related to the metabolite profiles.Dietary protein intake was significantly higher during the HP diet compared to the NP and LP diet. Urinary p-cresol excretion correlated positively with protein intake. Fecal water cytotoxicity correlated negatively with protein fermentation, while fecal water genotoxicity was not correlated with protein fermentation. Heptanal, 3-methyl-2-butanone, dimethyl disulfide and 2-propenyl ester of acetic acid are associated with genotoxicity and indole, 1-octanol, heptanal, 2,4-dithiapentane, allyl-isothiocyanate, 1-methyl-4-(1-methylethenyl-benzene, propionic acid, octanoic acid, nonanoic acid and decanoic acid with cytotoxicity.This study does not support a role of protein fermentation in gut toxicity. The identified metabolites can provide new insight into colonic health.ClinicalTrial.gov NCT01280513.

  19. Source of protein supplementation during in vitro culture does not affect the quality of resulting blastocysts in the domestic cat.

    Science.gov (United States)

    Nestle, E; Graves-Herring, J; Keefer, C; Comizzoli, P

    2012-12-01

    The objective of this study was to assess and compare the quality of cat blastocysts produced in vitro using commercial blastocyst growth media supplemented with different sources of proteins (serum protein substitute from in vitro maturation through embryo development vs 4 mg/ml of bovine serum albumin for maturation and 5% foetal calf serum for fertilization and embryo development). Impact was specifically examined on the proportion of blastocyst formation, total number of blastomeres, proportion of inner cell mass and expression of pluripotency marker proteins NANOG and OCT-4. Blastocyst formation per total cleaved embryos was similar (p > 0.05) regardless of the protein supplementation. There were no differences (p > 0.05) between culture conditions regarding average number of blastomeres and proportion of inner cell mass in each embryo. Presence of OCT-4 protein was detected in nuclei of both trophectoderm and inner cell mass region, with a stronger signal in the latter regardless of the culture medium. NANOG protein also was present in the inner cell mass regardless of the in vitro culture condition. We therefore demonstrated that serum protein substitute was as good as semi-defined protein sources for the production of good-quality blastocysts and embryonic stem cells. In addition, a single defined medium could be successfully used for cat oocyte maturation, in vitro fertilization and embryo development. © 2012 Blackwell Verlag GmbH.

  20. Protein source and quality in therapeutic foods affect the immune response and outcome in severe acute malnutrition

    Science.gov (United States)

    Protein is a vital component of therapeutic foods designed to treat severe acute malnutrition (SAM) in children; however there are still unknowns about the quality and quantity of the proteins to use in these foods. This review examines two recent studies investigating several different qualities an...

  1. Extraction of Pathogenesis-Related Proteins and Phenolics in Sauvignon Blanc as Affected by Grape Harvesting and Processing Conditions.

    Science.gov (United States)

    Tian, Bin; Harrison, Roland; Morton, James; Jaspers, Marlene; Hodge, Simon; Grose, Claire; Trought, Mike

    2017-07-12

    Thaumatin-like proteins (TLPs) and chitinases are the two main groups of pathogenesis-related (PR) proteins found in wine that cause protein haze formation. Previous studies have found that phenolics are also involved in protein haze formation. In this study, Sauvignon Blanc grapes were harvested and processed in two vintages (2011 and 2012) by three different treatments: (1) hand harvesting with whole bunch press (H-WB); (2) hand harvesting with destem/crush and 3 h skin contact (H-DC-3); and (3) machine harvesting with destem/crush and 3 h skin contact (M-DC-3). The juices were collected at three pressure levels (0.4 MPa, 0.8 MPa and 1.6 MPa), some juices were fermented in 750 mL of wine bottles to determine the bentonite requirement for the resulting wines. Results showed juices of M-DC-3 had significantly lower concentration of proteins, including PR proteins, compared to those of H-DC-3, likely due to the greater juice yield of M-DC-3 and interactions between proteins and phenolics. Juices from the 0.8-1.6 MPa pressure and resultant wines had the highest concentration of phenolics but the lowest concentration of TLPs. This supported the view that TLPs are released at low pressure as they are mainly present in grape pulp but additional extraction of phenolics largely present in skin occurs at higher pressing pressure. Wine protein stability tests showed a positive linear correlation between bentonite requirement and the concentration of chitinases, indicating the possibility of predicting bentonite requirement by quantification of chitinases. This study contributes to an improved understanding of extraction of haze-forming PR proteins and phenolics that can influence bentonite requirement for protein stabilization.

  2. Redox agents and N-ethylmaleimide affect protein polymerization during laboratory scale dry pasta production and cooking.

    Science.gov (United States)

    Bruneel, Charlotte; Buggenhout, Joke; Lagrain, Bert; Brijs, Kristof; Delcour, Jan A

    2016-04-01

    Durum wheat (Triticum durum Desf.) semolina gluten proteins consist of monomeric gliadin and polymeric glutenin and determine the quality of pasta products made therefrom. During pasta drying, glutenin starts polymerizing already below 60 °C (65% relative humidity (RH)), whereas gliadin only is incorporated in the protein network at temperatures exceeding 68 °C (68% RH) through thiol (SH)/disulfide (SS) exchange reactions. Removal of free SH groups in glutenin by adding 2.3 μmol KBrO3 or KIO3 per g dry matter semolina protein (g protein) or 13.8 μmol N-ethylmaleimide/g protein reduces gliadin-glutenin cross-linking during pasta drying and/or cooking and yields cooked pasta of high quality. Introducing free SH groups by adding 13.8 μmol glutathione/g protein increases gliadin-glutenin cross-linking during pasta processing, resulting in cooked pasta of lower quality. We hypothesize that too much gliadin incorporation in the glutenin network during pasta processing tightens the protein network and results in lower cooking quality. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Magnolol causes alterations in the cell cycle in androgen insensitive human prostate cancer cells in vitro by affecting expression of key cell cycle regulatory proteins.

    Science.gov (United States)

    McKeown, Brendan T; McDougall, Luke; Catalli, Adriana; Hurta, Robert A R

    2014-01-01

    Prostate cancer, one of the most common cancers in the Western world, affects many men worldwide. This study investigated the effects of magnolol, a compound found in the roots and bark of the magnolia tree Magnolia officinalis, on the behavior of 2 androgen insensitive human prostate cancer cell lines, DU145 and PC3, in vitro. Magnolol, in a 24-h exposure at 40 and 80 μM, was found to be cytotoxic to cells. Magnolol also affected cell cycle progression of DU145 and PC3 cells, resulting in alterations to the cell cycle and subsequently decreasing the proportion of cells entering the G2/M-phase of the cell cycle. Magnolol inhibited the expression of cell cycle regulatory proteins including cyclins A, B1, D1, and E, as well as CDK2 and CDK4. Protein expression levels of pRBp107 decreased and pRBp130 protein expression levels increased in response to magnolol exposure, whereas p16(INK4a), p21, and p27 protein expression levels were apparently unchanged post 24-h exposure. Magnolol exposure at 6 h did increase p27 protein expression levels. This study has demonstrated that magnolol can alter the behavior of androgen insensitive human prostate cancer cells in vitro and suggests that magnolol may have potential as a novel anti-prostate cancer agent.

  4. Energy metabolism in young mink kits (Neovison vison) affected by protein and carbohydrate level in the diet

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Hansen, NE; Tauson, A-H

    The mink is a strict carnivore and mink diets usually have a high content of protein. The energy metabolism in young minks in the transition period from milk to solid food is not investigated in detail, and the protein requirement is poorly defined. The substrate oxidation can give useful...... information about the relative contribution of different nutrients to the total heat production (HE; Tauson et al. 1997). The aim of the study was to examine the effect of different provision of protein and carbohydrate on the energy metabolism and substrate oxidation of mink kits between 6 and 12 weeks...

  5. Factors Affecting the Binding of a Recombinant Heavy Metal-Binding Domain (CXXC motif) Protein to Heavy Metals

    OpenAIRE

    Kamala Boonyodying; Thanakorn Watcharasupat; Waranan Yotpanya; Thawatchai Kitti; Wanna Kawang; Duangkamol Kunthalert; Sutthirat Sitthisak

    2012-01-01

    A number of heavy metal-binding proteins have been used to study bioremediation. CXXC motif, a metal binding domain containing Cys-X-X-Cys motif, has been identified in various organisms. These proteins are capable of binding various types of heavy metals. In this study, heavy metal binding domain (CXXC motif) recombinant protein encoded from mcsA gene of S. aureus were cloned and overexpressed in Escherichia coli. The factors involved in the metal-binding activity were determined in order to...

  6. Protein and carbohydrate composition of larval food affects tolerance tothermal stress and desiccation in adult Drosophila melanogaster

    DEFF Research Database (Denmark)

    Andersen, Laila H; Kristensen, Torsten N; Loeschcke, Volker

    2010-01-01

    as well as egg production and egg-to-adult viability. We raised larvae on a carbohydrate-enriched and a protein-enriched growth medium. We found that flies developed on the high protein medium had increased heat and desiccation tolerance compared to flies developed on the carbohydrate-enriched medium....... In contrast, flies developed on the carbohydrate-enriched growth medium recovered faster from chill coma stress compared to flies developed on a protein-enriched medium. We also found gender differences in stress tolerance, with female flies being more tolerant to chill coma, heat knockdown and desiccation...... when developing on the protein-enriched medium. Our study indicates that larval nutrition has a strong impact on the ability to cope with stress, and that the optimal nutrient composition varies with the type of stress...

  7. In Vivo pharmacokinetics of hesperidin are affected by treatment with glucosidase-like BglA protein isolated from yeasts

    OpenAIRE

    Li, Yong-mei; Li, Xiao-mian; Li, Guang-ming; Du, Wen-cai; Zhang, Jing; Li, Wei-xia; Xu, Jianshe; Hu, Ming; Zhu, Ze

    2008-01-01

    Hesperidin is an abundant flavanone glycoside in citrus fruits and has been reported to possess a wide range of biological activities. However, hesperidin has poor bioavailability. Here, we test the hypothesis that hesperetin found in Chenpi will have a better bioavailability than hesperidin and that treatment of hesperidin with glucosidase-like yeast Bg1A protein will increase its bioavailability. The results indicate that hesperidin in pure or extract form is hydrolyzed by BglA protein extr...

  8. Lipid-Based Nutrient Supplements During Pregnancy and Lactation Did Not Affect Human Milk Oligosaccharides and Bioactive Proteins in a Randomized Trial.

    Science.gov (United States)

    Jorgensen, Josh M; Arnold, Charles; Ashorn, Per; Ashorn, Ulla; Chaima, David; Cheung, Yin Bun; Davis, Jasmine Cc; Fan, Yue-Mei; Goonatilleke, Elisha; Kortekangas, Emma; Kumwenda, Chiza; Lebrilla, Carlito B; Maleta, Kenneth; Totten, Sarah M; Wu, Lauren D; Dewey, Kathryn G

    2017-08-09

    Background: Human milk oligosaccharides (HMOs) and bioactive proteins are beneficial to infant health. Recent evidence suggests that maternal nutrition may affect the amount of HMOs and proteins in breast milk; however, the effect of nutrient supplementation on HMOs and bioactive proteins has not yet been well studied.Objective: We aimed to determine whether lipid-based nutrient supplements (LNSs) affect milk bioactive protein and HMO concentrations at 6 mo postpartum in women in rural Malawi. These are secondary outcomes of a previously published randomized controlled trial.Methods: Women were randomly assigned to consume either an iron and folic acid capsule (IFA) daily from ≤20 wk gestation until delivery, followed by placebo daily from delivery to 6 mo postpartum, or a multiple micronutrient (MMN) capsule or LNS daily from ≤20 wk gestation to 6 mo postpartum. Breast milk concentrations of total HMOs, sialylated HMOs, fucosylated HMOs, lactoferrin, lactalbumin, lysozymes, antitrypsin, immunoglobulin A, and osteopontin were analyzed at 6 mo postpartum (n = 647). Between-group differences in concentrations and in proportions of women classified as having low concentrations were tested.Results: HMO and bioactive protein concentrations did not differ between groups (P > 0.10 for all comparisons). At 6 mo postpartum, the proportions of women with low HMOs or bioactive proteins were not different between groups except for osteopontin. A lower proportion of women in the IFA group had low osteopontin compared with the LNS group after adjusting for covariates (OR: 0.5; 95% CI: 0.3, 0.9; P = 0.016).Conclusion: The study findings do not support the hypothesis that supplementation with an LNS or MMN capsule during pregnancy and postpartum would increase HMO or bioactive milk proteins at 6 mo postpartum among Malawian women. This trial was registered at clinicaltrials.gov as NCT01239693. © 2017 American Society for Nutrition.

  9. Nuclear LSm8 affects number of cytoplasmic processing bodies via controlling cellular distribution of Like-Sm proteins.

    Science.gov (United States)

    Novotny, Ivan; Podolská, Katerina; Blazíková, Michaela; Valásek, Leos Shivaya; Svoboda, Petr; Stanek, David

    2012-10-01

    Processing bodies (P-bodies) are dynamic cytoplasmic structures involved in mRNA degradation, but the mechanism that governs their formation is poorly understood. In this paper, we address a role of Like-Sm (LSm) proteins in formation of P-bodies and provide evidence that depletion of nuclear LSm8 increases the number of P-bodies, while LSm8 overexpression leads to P-body loss. We show that LSm8 knockdown causes relocalization of LSm4 and LSm6 proteins to the cytoplasm and suggest that LSm8 controls nuclear accumulation of all LSm2-7 proteins. We propose a model in which redistribution of LSm2-7 to the cytoplasm creates new binding sites for other P-body components and nucleates new, microscopically visible structures. The model is supported by prolonged residence of two P-body proteins, DDX6 and Ago2, in P-bodies after LSm8 depletion, which indicates stronger interactions between these proteins and P-bodies. Finally, an increased number of P-bodies has negligible effects on microRNA-mediated translation repression and nonsense mediated decay, further supporting the view that the function of proteins localized in P-bodies is independent of visible P-bodies.

  10. SPA Proteins Affect the Subcellular Localization of COP1 in the COP1/SPA Ubiquitin Ligase Complex during Photomorphogenesis.

    Science.gov (United States)

    Balcerowicz, Martin; Kerner, Konstantin; Schenkel, Christian; Hoecker, Ute

    2017-07-01

    The Arabidopsis ( Arabidopsis thaliana ) COP1/SPA ubiquitin ligase is a central repressor that suppresses light signaling in darkness by targeting positive regulators of the light response, mainly transcription factors, for degradation. Light inactivates COP1/SPA, in part by excluding COP1 from the nucleus. SPA proteins are essential cofactors of COP1, but their exact role in the COP1/SPA complex is thus far unknown. To unravel a potential role of SPA proteins in COP1 nucleocytoplasmic partitioning, we monitored the subcellular localization of COP1 in a spa1234 quadruple mutant ( spaQn ). We analyzed a YFP-COP1-expressing transgenic line and endogenous COP1 after subcellular fractionation. In dark-grown seedlings, both YFP-COP1 and endogenous COP1 accumulated in the nucleus in the absence and presence of SPA proteins, indicating that SPA proteins are not required for nuclear localization of COP1 in darkness. In contrast, in white light-grown seedlings, spaQn mutants failed to relocalize COP1 from the nucleus to the cytoplasm. Hence, SPA proteins are necessary for the light-controlled change in COP1 subcellular localization. We conclude that SPA proteins have a dual role: (1) they are required for light-responsiveness of COP1 subcellular localization, and (2) they promote COP1 activity in darkness in a fashion that is independent of the nuclear import/nuclear retention of COP1. © 2017 American Society of Plant Biologists. All Rights Reserved.

  11. Proteins associated with heat-induced leaf senescence in creeping bentgrass as affected by foliar application of nitrogen, cytokinins, and an ethylene inhibitor.

    Science.gov (United States)

    Jespersen, David; Huang, Bingru

    2015-02-01

    Heat stress causes premature leaf senescence in cool-season grass species. The objective of this study was to identify proteins regulated by nitrogen, cytokinins, and ethylene inhibitor in relation to heat-induced leaf senescence in creeping bentgrass (Agrostis stolonifera). Plants (cv. Penncross) were foliar sprayed with 18 mM carbonyldiamide (N source), 25 μM aminoethoxyvinylglycine (AVG, ethylene inhibitor), 25 μM zeatin riboside (ZR, cytokinin), or a water control, and then exposed to 20/15°C (day/night) or 35/30°C (heat stress) in growth chambers. All treatments suppressed heat-induced leaf senescence, as shown by higher turf quality and chlorophyll content, and lower electrolyte leakage in treated plants compared to the untreated control. A total of 49 proteins were responsive to N, AVG, or ZR under heat stress. The abundance of proteins in photosynthesis increased, with ribulose-1,5-bisphosphate carboxylase/oxygenase affected by all three treatments, chlorophyll a/b-binding protein by AVG and N or Rubisco activase by AVG. Proteins for amino acid metabolism were upregulated, including alanine aminotransferase by three treatments and ferredoxin-dependent glutamate synthase by AVG and N. Upregulated proteins also included catalase by AVG and N and heat shock protein by ZR. Exogenous applications of AVG, ZR, or N downregulated proteins in respiration (enolase, glyceraldehyde 3-phosphate dehydrogenase, and succinate dehygrogenase) under heat stress. Alleviation of heat-induced senescence by N, AVG, or ZR was associated with enhanced protein abundance in photosynthesis and amino acid metabolism and stress defense systems (heat shock protection and antioxidants), as well as suppression of those imparting respiration metabolism. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Protein Turnover and Cellular Stress in Mildly and Severely Affected Muscles from Patients with Limb Girdle Muscular Dystrophy Type 2I

    DEFF Research Database (Denmark)

    Hauerslev, Simon; Sveen, Marie-Louise; Vissing, John

    2013-01-01

    Patients with Limb girdle muscular dystrophy type 2I (LGMD2I) are characterized by progressive muscle weakness and wasting primarily in the proximal muscles, while distal muscles often are spared. Our aim was to investigate if wasting could be caused by impaired regeneration in the proximal...... highly increased in severely affected muscles compared to mildly affected muscles. Our results indicate that alterations in the protein turnover and myostatin levels could progressively impair the muscle mass maintenance and/or regeneration resulting in gradual muscular atrophy....

  13. The Oncogenic Fusion Proteins SET-Nup214 and Sequestosome-1 (SQSTM1)-Nup214 Form Dynamic Nuclear Bodies and Differentially Affect Nuclear Protein and Poly(A)+ RNA Export.

    Science.gov (United States)

    Port, Sarah A; Mendes, Adélia; Valkova, Christina; Spillner, Christiane; Fahrenkrog, Birthe; Kaether, Christoph; Kehlenbach, Ralph H

    2016-10-28

    Genetic rearrangements are a hallmark of several forms of leukemia and can lead to oncogenic fusion proteins. One example of an affected chromosomal region is the gene coding for Nup214, a nucleoporin that localizes to the cytoplasmic side of the nuclear pore complex (NPC). We investigated two such fusion proteins, SET-Nup214 and SQSTM1 (sequestosome)-Nup214, both containing C-terminal portions of Nup214. SET-Nup214 nuclear bodies containing the nuclear export receptor CRM1 were observed in the leukemia cell lines LOUCY and MEGAL. Overexpression of SET-Nup214 in HeLa cells leads to the formation of similar nuclear bodies that recruit CRM1, export cargo proteins, and certain nucleoporins and concomitantly affect nuclear protein and poly(A) + RNA export. SQSTM1-Nup214, although mostly cytoplasmic, also forms nuclear bodies and inhibits nuclear protein but not poly(A) + RNA export. The interaction of the fusion proteins with CRM1 is RanGTP-dependent, as shown in co-immunoprecipitation experiments and binding assays. Further analysis revealed that the Nup214 parts mediate the inhibition of nuclear export, whereas the SET or SQSTM1 part determines the localization of the fusion protein and therefore the extent of the effect. SET-Nup214 nuclear bodies are highly mobile structures, which are in equilibrium with the nucleoplasm in interphase and disassemble during mitosis or upon treatment of cells with the CRM1-inhibitor leptomycin B. Strikingly, we found that nucleoporins can be released from nuclear bodies and reintegrated into existing NPC. Our results point to nuclear bodies as a means of preventing the formation of potentially insoluble and harmful protein aggregates that also may serve as storage compartments for nuclear transport factors. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. Degradation kinetics of fisetin and quercetin in solutions affected by medium pH, temperature and co-existed proteins

    Directory of Open Access Journals (Sweden)

    Wang Jing

    2016-01-01

    Full Text Available Impacts of medium pH, temperature and coexisted proteins on the degradation of two flavonoids fisetin and quercetin were assessed by spectroscopic method in the present study. Based on the measured degradation rate constants (k, fisetin was more stable than quercetin in all cases. Increasing medium pH from 6.0 to 7.5 at 37°C enhanced respective k values of fisetin and quercetin from 8.30x10−3 and 2.81x10−2 to 0.202 and 0.375 h-1 (P<0.05. In comparison with their degradation at 37°C, fisetin and quercetin showed larger k values at higher temperature (0.124 and 0.245 h−1 at 50°C, or 0.490 and 1.42 h−1 at 65°C. Four protein products in medium could stabilize the two flavonoids (P<0.05, as these proteins at 0.10 g L-1 decreased respective k values of fisetin and quercetin to 2.28x10−2-2.98x10−2 and 4.37´10−2-5.97x10−2 h−1. Hydrophobic interaction between the proteins and the two flavonoids was evidenced responsible for the stabilization, as sodium dodecyl sulfate could destroy the stabilization significantly (P<0.05. Casein and soybean protein provided greater stabilization than whey protein isolate. It is thus concluded that higher temperature and alkaline pH can enhance flavonoid loss, whereas coexisted proteins as flavonoid stabilizers can inhibit flavonoid degradation.

  15. Using Coexpression Protein Interaction Network Analysis to Identify Mechanisms of Danshensu Affecting Patients with Coronary Heart Disease

    Directory of Open Access Journals (Sweden)

    Mengqi Huo

    2017-06-01

    Full Text Available Salvia miltiorrhiza, known as Danshen, has attracted worldwide interest for its substantial effects on coronary heart disease (CHD. Danshensu (DSS is one of the main active ingredients of Danshen on CHD. Although it has been proven to have a good clinical effect on CHD, the action mechanisms remain elusive. In the current study, a coexpression network-based approach was used to illustrate the beneficial properties of DSS in the context of CHD. By integrating the gene expression profile data and protein-protein interactions (PPIs data, two coexpression protein interaction networks (CePIN in a CHD state (CHD CePIN and a non-CHD state (non-CHD CePIN were generated. Then, shared nodes and unique nodes in CHD CePIN were attained by conducting a comparison between CHD CePIN and non-CHD CePIN. By calculating the topological parameters of each shared node and unique node in the networks, and comparing the differentially expressed genes, target proteins involved in disease regulation were attained. Then, Gene Ontology (GO enrichment was utilized to identify biological processes associated to target proteins. Consequently, it turned out that the treatment of CHD with DSS may be partly attributed to the regulation of immunization and blood circulation. Also, it indicated that sodium/hydrogen exchanger 3 (SLC9A3, Prostaglandin G/H synthase 2 (PTGS2, Oxidized low-density lipoprotein receptor 1 (OLR1, and fibrinogen gamma chain (FGG may be potential therapeutic targets for CHD. In summary, this study provided a novel coexpression protein interaction network approach to provide an explanation of the mechanisms of DSS on CHD and identify key proteins which maybe the potential therapeutic targets for CHD.

  16. Reduced Abundance and Subverted Functions of Proteins in Prion-Like Diseases: Gained Functions Fascinate but Lost Functions Affect Aetiology

    Science.gov (United States)

    Nguyen-Phuoc, Kim; Leighton, Patricia L. A.

    2017-01-01

    Prions have served as pathfinders that reveal many aspects of proteostasis in neurons. The recent realization that several prominent neurodegenerative diseases spread via a prion-like mechanism illuminates new possibilities for diagnostics and therapeutics. Thus, key proteins in Alzheimer Disease and Amyotrophic lateral sclerosis (ALS), including amyloid-β precursor protein, Tau and superoxide dismutase 1 (SOD1), spread to adjacent cells in their misfolded aggregated forms and exhibit template-directed misfolding to induce further misfolding, disruptions to proteostasis and toxicity. Here we invert this comparison to ask what these prion-like diseases can teach us about the broad prion disease class, especially regarding the loss of these key proteins’ function(s) as they misfold and aggregate. We also consider whether functional amyloids might reveal a role for subverted protein function in neurodegenerative disease. Our synthesis identifies SOD1 as an exemplar of protein functions being lost during prion-like protein misfolding, because SOD1 is inherently unstable and loses function in its misfolded disease-associated form. This has under-appreciated parallels amongst the canonical prion diseases, wherein the normally folded prion protein, PrPC, is reduced in abundance in fatal familial insomnia patients and during the preclinical phase in animal models, apparently via proteostatic mechanisms. Thus while template-directed misfolding and infectious properties represent gain-of-function that fascinates proteostasis researchers and defines (is required for) the prion(-like) diseases, loss and subversion of the functions attributed to hallmark proteins in neurodegenerative disease needs to be integrated into design towards effective therapeutics. We propose experiments to uniquely test these ideas. PMID:29064456

  17. Dietary N-Carbamylglutamate Supplementation in a Reduced Protein Diet Affects Carcass Traits and the Profile of Muscle Amino Acids and Fatty Acids in Finishing Pigs.

    Science.gov (United States)

    Ye, Changchuan; Zeng, Xiangzhou; Zhu, Jinlong; Liu, Ying; Ye, Qianhong; Qiao, Shiyan; Zeng, Xiangfang

    2017-07-19

    The aim of this study was to investigate whether dietary N-carbamylglutamate (NCG) supplementation in a reduced protein diet affected carcass traits and meat quality in finishing pigs. A total of 120 gilts were randomly assigned to one of four treatments for 40 days, including a standard protein diet (SP), a reduced protein diet supplemented with 1.7% l-alanine (RP + Ala), a reduced protein diet supplemented with 1.0% l-arginine (RP + Arg), and a reduced protein diet supplemented with 0.1% NCG and 1.7% l-alanine (RP + NCG). NCG supplementation increased the endogenous synthesis of l-arginine. The RP + NCG diet significantly increased the loin eye area (p protein diet was effective to produce functional pork with a high content of leucine (p dietary NCG supplementation. In conclusion, the RP + NCG diet is effective to increase the longissimus dorsi muscle area, decrease back fat accretion, and produce functional pork with a high content of leucine but without a negative impact on the muscle fatty acid profile in finishing pigs.

  18. Xylo-Oligosaccharides and Inulin Affect Genotoxicity and Bacterial Populations Differently in a Human Colonic Simulator Challenged with Soy Protein

    Science.gov (United States)

    Christophersen, Claus T.; Petersen, Anne; Licht, Tine R.; Conlon, Michael A.

    2013-01-01

    High dietary intakes of some protein sources, including soy protein, can increase colonic DNA damage in animals, whereas some carbohydrates attenuate this. We investigated whether inulin and xylo-oligosaccharides (XOS) could be protective against DNA strand breaks by adding them to a human colonic simulator consisting of a proximal vessel (PV) (pH 5.5) and a distal vessel (DV) (pH 6.8) inoculated with human faeces and media containing soy protein. Genotoxicity of the liquid phase and microbial population changes in the vessels were measured. Soy protein (3%) was fermented with 1% low amylose cornstarch for 10 day followed by soy protein with 1% XOS or 1% inulin for 10 day. Inulin did not alter genotoxicity but XOS significantly reduced PV genotoxicity and increased DV genotoxicity. Inulin and XOS significantly increased butyrate concentration in the DV but not PV. Numbers of the key butyrate-producing bacterium Faecalibacterium prausnitzii were significantly increased in the PV and DV by inulin but significantly decreased by XOS in both vessels. Other bacteria examined were also significantly impacted by the carbohydrate treatments or by the vessel (i.e., pH). There was a significant overall inverse correlation between levels of damage induced by the ferments and levels of sulphate-reducing bacteria, Bacteroides fragilis, and acetate. In conclusion, dietary XOS can potentially modulate the genotoxicity of the colonic environment and specific bacterial groups and short chain fatty acids may mediate this. PMID:24064573

  19. Uncoupling protein 2 haplotype does not affect human brain structure and function in a sample of community-dwelling older adults.

    Directory of Open Access Journals (Sweden)

    Verena Heise

    Full Text Available Uncoupling protein 2 (UCP2 is a mitochondrial membrane protein that plays a role in uncoupling electron transport from adenosine triphosphate (ATP formation. Polymorphisms of the UCP2 gene in humans affect protein expression and function and have been linked to survival into old age. Since UCP2 is expressed in several brain regions, we investigated in this study whether UCP2 polymorphisms might 1 affect occurrence of neurodegenerative or mental health disorders and 2 affect measures of brain structure and function. We used structural magnetic resonance imaging (MRI, diffusion-weighted MRI and resting-state functional MRI in the neuroimaging sub-study of the Whitehall II cohort. Data from 536 individuals aged 60 to 83 years were analyzed. No association of UCP2 polymorphisms with the occurrence of neurodegenerative disorders or grey and white matter structure or resting-state functional connectivity was observed. However, there was a significant effect on occurrence of mood disorders in men with the minor alleles of -866G>A (rs659366 and Ala55Val (rs660339 being associated with increasing odds of lifetime occurrence of mood disorders in a dose dependent manner. This result was not accompanied by effects of UCP2 polymorphisms on brain structure and function, which might either indicate that the sample investigated here was too small and underpowered to find any significant effects, or that potential effects of UCP2 polymorphisms on the brain are too subtle to be picked up by any of the neuroimaging measures used.

  20. Xylo-oligosaccharides and inulin affect genotoxicity and bacterial populations differently in a human colonic simulator challenged with soy protein

    DEFF Research Database (Denmark)

    Christophersen, C. T.; Petersen, Anne; Licht, Tine Rask

    2013-01-01

    High dietary intakes of some protein sources, including soy protein, can increase colonic DNA damage in animals, whereas some carbohydrates attenuate this. We investigated whether inulin and xylo-oligosaccharides (XOS) could be protective against DNA strand breaks by adding them to a human colonic...... cornstarch for 10 day followed by soy protein with 1% XOS or 1% inulin for 10 day. Inulin did not alter genotoxicity but XOS significantly reduced PV genotoxicity and increased DV genotoxicity. Inulin and XOS significantly increased butyrate concentration in the DV but not PV. Numbers of the key butyrate......-producing bacterium Faecalibacterium prausnitzii were significantly increased in the PV and DV by inulin but significantly decreased by XOS in both vessels. Other bacteria examined were also significantly impacted by the carbohydrate treatments or by the vessel (i.e., pH). There was a significant overall inverse...

  1. Polymorphism in SFTPD gene affects assembly and constitutional serum levels of surfactant protein D in a Lebanese population

    DEFF Research Database (Denmark)

    Fakih, Dalia; Chamat, Soulaima; Medlej-Hashim, Myrna

    2014-01-01

    to a threonine or methionine at position 11 of the SP-D protein. SP-D was measured by ELISA in serum taken from 97 healthy subjects. The p.Met11Thr SNP was genotyped. Different forms of SP-D were separated by gel filtration chromatography. Basal serum SP-D levels varied extensively, over a range of 108.3 ng......Surfactant protein D (SP-D), an oligomeric lung-derived lectin, has essential roles in innate immunity. It can be measured in serum. Previous studies have shown that constitutional SP-D serum levels and the protein degree of multimerization are genetically influenced. We aimed to establish...

  2. Protein-enriched meal replacements do not adversely affect liver, kidney or bone density: an outpatient randomized controlled trial

    Directory of Open Access Journals (Sweden)

    Yan Eric

    2010-12-01

    Full Text Available Abstract Background There is concern that recommending protein-enriched meal replacements as part of a weight management program could lead to changes in biomarkers of liver or renal function and reductions in bone density. This study was designed as a placebo-controlled clinical trial utilizing two isocaloric meal plans utilizing either a high protein-enriched (HP or a standard protein (SP meal replacement in an outpatient weight loss program. Subjects/methods 100 obese men and women over 30 years of age with a body mass index (BMI between 27 to 40 kg/m2 were randomized to one of two isocaloric weight loss meal plans 1. HP group: providing 2.2 g protein/kg of lean body mass (LBM/day or 2. SP group: providing 1.1 g protein/kg LBM/day. Meal replacement (MR was used twice daily (one meal, one snack for 3 months and then once a day for 9 months. Body weight, lipid profiles, liver function, renal function and bone density were measured at baseline and 12 months. Results Seventy subjects completed the study. Both groups lost weight (HP -4.29 ± 5.90 kg vs. SP -4.66 ± 6.91 kg, p 2, p = 0.210; SP -0.03 ± 0.17 g/cm2, p = 0.320 in either group over one year. Conclusions These studies demonstrate that protein-enriched meals replacements as compared to standard meal replacements recommended for weight management do not have adverse effects on routine measures of liver function, renal function or bone density at one year. Clinicaltrial.gov: NCT01030354.

  3. Magnolol Affects Cellular Proliferation, Polyamine Biosynthesis and Catabolism-Linked Protein Expression and Associated Cellular Signaling Pathways in Human Prostate Cancer Cells in vitro

    Directory of Open Access Journals (Sweden)

    Brendan T. McKeown

    2015-01-01

    Full Text Available Background: Prostate cancer is the most commonly diagnosed form of cancer in men in Canada and the United States. Both genetic and environmental factors contribute to the development and progression of many cancers, including prostate cancer. Context and purpose of this study: This study investigated the effects of magnolol, a compound found in the roots and bark of the magnolia tree Magnolia officinalis, on cellular proliferation and proliferation-linked activities of PC3 human prostate cancer cells in vitro. Results: PC3 cells exposed to magnolol at a concentration of 80 μM for 6 hours exhibited decreased protein expression of ornithine decarboxylase, a key regulator in polyamine biosynthesis, as well as affecting the expression of other proteins involved in polyamine biosynthesis and catabolism. Furthermore, protein expression of the R2 subunit of ribonucleotide reductase, a key regulatory protein associated with DNA synthesis, was significantly decreased. Finally, the MAPK (mitogen-activated protein kinase, PI3K (phosphatidylinositol 3-kinase, NFκB (nuclear factor of kappa-light-chain-enhancer of activated B cells and AP-1 (activator protein 1 cellular signaling pathways were assayed to determine which, if any, of these pathways magnolol exposure would alter. Protein expressions of p-JNK-1 and c-jun were significantly increased while p-p38, JNK-1/2, PI3Kp85, p-PI3Kp85, p-Akt, NFκBp65, p-IκBα and IκBα protein expressions were significantly decreased. Conclusions: These alterations further support the anti-proliferative effects of magnolol on PC3 human prostate cancer cells in vitro and suggest that magnolol may have potential as a novel anti-prostate cancer agent.

  4. Nuclear LSm8 affects number of cytoplasmic processing bodies via controlling cellular distribution of Like-Sm proteins

    OpenAIRE

    Novotný, Ivan; Podolská, Kateřina; Blažíková, Michaela; Valášek, Leoš Shivaya; Svoboda, Petr; Staněk, David

    2012-01-01

    Processing bodies (P-bodies) are dynamic cytoplasmic structures involved in mRNA degradation, but the mechanism that governs their formation is poorly understood. In this paper, we address a role of Like-Sm (LSm) proteins in formation of P-bodies and provide evidence that depletion of nuclear LSm8 increases the number of P-bodies, while LSm8 overexpression leads to P-body loss. We show that LSm8 knockdown causes relocalization of LSm4 and LSm6 proteins to the cytoplasm and suggest that LSm8 c...

  5. Induced lung inflammation and dietary protein supply affect nitrogen retention and amino acid metabolism in growing pigs

    NARCIS (Netherlands)

    Kampman-van de Hoek, E.; Sakkas, P.; Gerrits, W.J.J.; Borne, van den J.J.G.C.; Peet-Schwering, van der C.M.C.; Jansman, A.J.M.

    2015-01-01

    It is hypothesised that during immune system activation, there is a competition for amino acids (AA) between body protein deposition and immune system functioning. The aim of the present study was to quantify the effect of immune system activation on N retention and AA metabolism in growing pigs,

  6. Skeletal muscle myofibrillar and sarcoplasmic protein synthesis rates are affected differently by altitude-induced hypoxia in native lowlanders

    DEFF Research Database (Denmark)

    Holm, Lars; Haslund, Mads Lyhne; Robach, Paul

    2010-01-01

    As a consequence to hypobaric hypoxic exposure skeletal muscle atrophy is often reported. The underlying mechanism has been suggested to involve a decrease in protein synthesis in order to conserve O(2). With the aim to challenge this hypothesis, we applied a primed, constant infusion of 1-(13)C-...

  7. How age and sex affect the erythrocyte sedimentation rate and C-reactive protein in early rheumatoid arthritis

    NARCIS (Netherlands)

    Siemons, L.; Klooster, P.M. ten; Vonkeman, H.E.; Riel, P.L.C.M. van; Glas, C.A.; Laar, M.A. van der

    2014-01-01

    BACKGROUND: The erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) are two commonly used measures of inflammation in rheumatoid arthritis (RA). As current RA treatment guidelines strongly emphasize early and aggressive treatment aiming at fast remission, optimal measurement of

  8. Hepatitis B Virus Core Protein Phosphorylation Sites Affect Capsid Stability and Transient Exposure of the C-terminal Domain.

    Science.gov (United States)

    Selzer, Lisa; Kant, Ravi; Wang, Joseph C-Y; Bothner, Brian; Zlotnick, Adam

    2015-11-20

    Hepatitis B virus core protein has 183 amino acids divided into an assembly domain and an arginine-rich C-terminal domain (CTD) that regulates essential functions including genome packaging, reverse transcription, and intracellular trafficking. Here, we investigated the CTD in empty hepatitis B virus (HBV) T=4 capsids. We examined wild-type core protein (Cp183-WT) and a mutant core protein (Cp183-EEE), in which three CTD serines are replaced with glutamate to mimic phosphorylated protein. We found that Cp183-WT capsids were less stable than Cp183-EEE capsids. When we tested CTD sensitivity to trypsin, we detected two different populations of CTDs differentiated by their rate of trypsin cleavage. Interestingly, CTDs from Cp183-EEE capsids exhibited a much slower rate of proteolytic cleavage when compared with CTDs of Cp183-WT capsids. Cryo-electron microscopy studies of trypsin-digested capsids show that CTDs at five-fold symmetry vertices are most protected. We hypothesize that electrostatic interactions between glutamates and arginines in Cp183-EEE, particularly at five-fold, increase capsid stability and reduce CTD exposure. Our studies show that quasi-equivalent CTDs exhibit different rates of exposure and thus might perform distinct functions during the hepatitis B virus lifecycle. Our results demonstrate a structural role for CTD phosphorylation and indicate crosstalk between CTDs within a capsid particle. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  9. Binding of hydrogen-citrate to photoactive yellow protein is affected by the structural changes related to signaling state formation

    NARCIS (Netherlands)

    Hospes, M.; Ippel, J.H.|info:eu-repo/dai/nl/115343415; Boelens, R.|info:eu-repo/dai/nl/070151407; Hellingwerf, K.J.; Hendriks, Johnny

    2012-01-01

    The tricarboxylic acid citric acid is a key intermediary metabolite in organisms from all domains of the tree of life. Surprisingly, this metabolite specifically interacts with the light-induced signaling state of the photoactive yellow protein (PYP), such that, at 30 mM, it retards recovery of this

  10. Binding of hydrogen-citrate to photoactive yellow protein is affected by the structural changes related to signaling state formation.

    NARCIS (Netherlands)

    Hospes, M.; Ippel, J.H.; Boelens, R.; Hellingwerf, K.J.; Hendriks, J.

    2012-01-01

    The tricarboxylic acid citric acid is a key intermediary metabolite in organisms from all domains of the tree of life. Surprisingly, this metabolite specifically interacts with the light-induced signaling state of the photoactive yellow protein (PYP), such that, at 30 mM, it retards recovery of this

  11. Peptide production and decay rates affect the quantitative accuracy of protein cleavage isotope dilution mass spectrometry (PC-IDMS).

    Science.gov (United States)

    Shuford, Christopher M; Sederoff, Ronald R; Chiang, Vincent L; Muddiman, David C

    2012-09-01

    No consensus has been reached on the proper time to add stable-isotope labeled (SIL) peptides in protein cleavage isotope dilution mass spectrometry workflows. While quantifying 24 monolignol pathway enzymes in the xylem tissue of Populus trichocarpa, we compared the protein concentrations obtained when adding the SIL standard peptides concurrently with the enzyme or after quenching of the digestion (i.e. postdigestion) and observed discrepancies for nearly all tryptic peptides investigated. In some cases, greater than 30-fold differences were observed. To explain these differences and potentially correct for them, we developed a mathematical model based on pseudo-first-order kinetics to account for the dynamic production and decay (e.g. degradation and precipitation) of the native peptide targets in conjunction with the decay of the SIL peptide standards. A time course study of the digests confirmed the results predicted by the proposed model and revealed that the discrepancy between concurrent and postdigestion introduction of the SIL standards was related to differential decay experienced by the SIL peptide and the native peptide in each method. Given these results, we propose concurrent introduction of the SIL peptide is most appropriate, though not free from bias. Mathematical modeling of this method reveals that overestimation of protein quantities would still result when rapid peptide decay occurs and that this bias would be further exaggerated by slow proteolysis. We derive a simple equation to estimate the bias for each peptide based on the relative rates of production and decay. According to this equation, nearly half of the peptides evaluated here were estimated to have quantitative errors greater than 10% and in a few cases over 100%. We conclude that the instability of peptides can often significantly bias the protein quantities measured in protein cleavage isotope dilution mass spectrometry-based assays and suggest peptide stability be made a priority

  12. Dietary protein during gestation affects maternal insulin-like growth factor, insulin-like growth factor binding protein, leptin concentrations, and fetal growth in heifers.

    Science.gov (United States)

    Sullivan, T M; Micke, G C; Perkins, N; Martin, G B; Wallace, C R; Gatford, K L; Owens, J A; Perry, V E A

    2009-10-01

    The influence of supplemental protein during gestation on maternal hormones and fetal growth was determined in composite beef heifers. At AI, 118 heifers were stratified by BW within each composite genotype (BeefX = 1/2 Senepol, 1/4 Brahman, 1/8 Charolais, 1/8 Red Angus and CBX = 1/2 Senepol, 1/4 Brahman, 1/4 Charolais) into 4 treatment groups: high high (HH = 1.4 kg CP/d for first and second trimesters of gestation), high low (HL = 1.4 kg of CP/d for first trimester and 0.4 kg of CP/d for second trimester), low high (lowH = 0.4 kg CP/d for first trimester and 1.4 kg of CP/d and for second trimester), or low low (LL = 0.4 kg CP/d for first and second trimesters). Maternal plasma IGF-I and -II, total IGFBP, and leptin concentrations were determined at 14 d before AI and at d 28, 82, 179, and 271 post-AI (mean gestation length 286 d), and leptin concentrations were also determined at calving. Increased dietary protein increased maternal plasma IGF-I (P < 0.001 on d 28, 82, and 179), IGF-II (P = 0.01 on d 82; P = 0.04 on d 271), and total IGFBP (P = 0.002 on d 82; P = 0.005 on d 179; P = 0.03 on d 271). Maternal plasma IGF-I at d 271 was negatively associated with calf crown-rump length at birth (P = 0.003). BeefX had greater birth weight calves (P = 0.01), greater IGF-II (P < 0.001), increased ratios of IGF-I:total IGFBP (P = 0.008) and IGF-II:total IGFBP (P < 0.001), and reduced total IGFBP compared with CBX (P = 0.02). Increased dietary protein during second trimester increased maternal plasma leptin at calving (P = 0.005). Maternal plasma leptin near term was positively associated with heifer BCS (P = 0.02) and with calf birth weight (P = 0.04), and at calving was positively associated with heifer age at AI (P = 0.02). These findings suggest that maternal dietary protein, age, and genotype influence plasma concentrations of metabolic hormones and fetal growth in Bos indicus-influenced heifers.

  13. TOX4 and NOVA1 proteins are partners of the LEDGF PWWP domain and affect HIV-1 replication.

    Directory of Open Access Journals (Sweden)

    Mehdi Morchikh

    Full Text Available PWWP domains are involved in the chromatin attachment of several proteins. They bind to both DNA and proteins and their interaction with specific histone methylation marks define them as a new class of histone code readers. The lens epithelium derived growth factor (LEDGF/p75 contains an N-terminal PWWP domain necessary for its interaction with chromatin but also a C-terminal domain which interacts with several proteins, such as lentiviral integrases. These two domains confer a chromatin-tethering function to LEDGF/p75 and in the case of lentiviral integrases, this tethering participates in the efficiency and site selectivity of integration. Although proteins interacting with LEDGF/p75 C-terminal domain have been extensively studied, no data exist about partners of its PWWP domain regulating its interaction with chromatin. In this study, we report the identification by yeast-two-hybrid of thirteen potential partners of the LEDGF PWWP domain. Five of these interactions were confirmed in mammalian cells, using both a protein complementation assay and co-immunoprecipitation approaches. Three of these partners interact with full length LEDGF/p75, they are specific for PWWP domains of the HDGF family and they require PWWP amino acids essential for the interaction with chromatin. Among them, the transcription activator TOX4 and the splicing cofactor NOVA1 were selected for a more extensive study. These two proteins or their PWWP interacting regions (PIR colocalize with LEDGF/p75 in Hela cells and interact in vitro in the presence of DNA. Finally, single round VSV-G pseudotyped HIV-1 but not MLV infection is inhibited in cells overexpressing these two PIRs. The observed inhibition of infection can be attributed to a defect in the integration step. Our data suggest that a regulation of LEDGF interaction with chromatin by cellular partners of its PWWP domain could be involved in several processes linked to LEDGF tethering properties, such as lentiviral

  14. TRH receptor mobility in the plasma membrane is strongly affected by agonist binding and by interaction with some cognate signaling proteins.

    Science.gov (United States)

    Moravcova, Radka; Melkes, Barbora; Novotny, Jiri

    2017-11-14

    Extensive research has been dedicated to elucidating the mechanisms of signal transduction through different G protein-coupled receptors (GPCRs). However, relatively little is known about the regulation of receptor movement within the cell membrane upon ligand binding. In this study we focused our attention on the thyrotropin-releasing hormone (TRH) receptor that typically couples to Gq/11 proteins. We monitored receptor diffusion in the plasma membrane of HEK293 cells stably expressing yellow fluorescent protein (YFP)-tagged TRH receptor (TRHR-YFP) by fluorescence recovery after photobleaching (FRAP). FRAP analysis indicated that the lateral movement of the TRH receptor was markedly reduced upon TRH binding as the value of its diffusion coefficient fell down by 55%. This effect was prevented by the addition of the TRH receptor antagonist midazolam. We also found that siRNA-mediated knockdown of Gq/11α, Gβ, β-arrestin2 and phospholipase Cβ1, but not of Giα1, β-arrestin1 or G protein-coupled receptor kinase 2, resulted in a significant decrease in the rate of TRHR-YFP diffusion, indicating the involvement of the former proteins in the regulation of TRH receptor behavior. The observed partial reduction of the TRHR-YFP mobile fraction caused by down-regulation of Giα1 and β-arrestin1 suggests that these proteins may also play distinct roles in THR receptor-mediated signaling. These results demonstrate for the first time that not only agonist binding but also abundance of some signaling proteins may strongly affect TRH receptor dynamics in the plasma membrane.

  15. Diets high in protein or saturated fat do not affect insulin sensitivity or plasma concentrations of lipids and lipoproteins in overweight and obese adults.

    Science.gov (United States)

    Chiu, Sally; Williams, Paul T; Dawson, Taylor; Bergman, Richard N; Stefanovski, Darko; Watkins, Steven M; Krauss, Ronald M

    2014-11-01

    Previous human studies reported inconsistent effects of dietary protein and branched-chain amino acids (BCAAs) on insulin action and glucose metabolism. Similarly, it is unclear whether saturated fat (SF) intake influences these metabolic variables. The objective of this study was to test the effects of high [30% of energy (%E)] vs. moderate (20%E) intakes of protein (primarily whey) on insulin action and lipid and lipoprotein concentrations in the context of both high (15%E) and low (7%E) SF diets. The study was conducted as a randomized controlled trial in 158 overweight and obese men and women. After a 4-wk baseline diet [55%E carbohydrate, 15%E protein, 30%E fat (7%E SF)], participants were randomly assigned to 4 wk of either the baseline diet or 1 of 4 test diets containing 35%E carbohydrate and either 20%E or 30%E protein and either 7%E or 15%E SF. Frequently sampled i.v. glucose tolerance tests were administered after each dietary period. Other than significantly higher fasting glucose concentrations for high vs. moderate protein intakes with a low-fat diet (difference ± SE: 0.47 ± 0.14 mmol/L; P = 0.001), there were no significant effects of dietary protein or SF on glucose metabolism, plasma insulin, or concentrations of lipids and lipoproteins. Changes in plasma BCAAs across all diets were negatively correlated with changes in the metabolic clearance rate of insulin (ρ = -0.18, P = 0.03) and positively correlated with changes in the acute insulin response to glucose (ρ = 0.15, P = 0.05). These findings suggest that short-term intake of BCAAs can influence insulin dynamics. However, in this group of overweight and obese individuals, neither high protein nor SF intake affected insulin sensitivity or plasma concentrations of lipids and lipoproteins. This trial was registered at clinicaltrials.gov as NCT00508937. © 2014 American Society for Nutrition.

  16. Down-Regulation of Small Rubber Particle Protein Expression Affects Integrity of Rubber Particles and Rubber Content in Taraxacum brevicorniculatum

    Czech Academy of Sciences Publication Activity Database

    Hillebrand, A.; Post, J. J.; Wurbs, D.; Wahler, D.; Lenders, D.; Krzyžánek, Vladislav; Pruefer, D.; Gronover, CH. S.

    2012-01-01

    Roč. 7, č. 7 (2012), e41874:1-9 E-ISSN 1932-6203 Institutional support: RVO:68081731 Keywords : Hevea-Brasiliensis * Parthenium-Argentatum * Elongation-Factor * Silencing Affects * Surface- Structure * Oil Bodies * Latex * Prenyltransferase * Biosynthesis * Stability Subject RIV: CE - Biochemistry Impact factor: 3.730, year: 2012

  17. Exposure to brominated flame retardant PBDE-99 affects cytoskeletal protein expression in the neonatal mouse cerebral cortex

    DEFF Research Database (Denmark)

    Alm, Henrik; Kultima, Kim; Scholz, Birger

    2008-01-01

    Polybrominated diphenyl ethers (PBDEs) are environmental contaminants found in human and animal tissues worldwide. Neonatal exposure to the flame retardant 2,2', 4,4',5-pentabromodiphenyl ether (PBDE-99) disrupts normal brain development in mice, and results in disturbed spontaneous behavior....... Protein resolution was enhanced by sample pre-fractionation. In the cell model, we determined cell viability using the trypan blue exclusion assay, and apoptosis using immunocytochemical detection of cleaved caspase-3. We determined the identity of 111 differentially expressed proteins, 32 (29%) of which...... activity. These results indicate that the permanent neurological damage induced by PBDE-99 during the brain growth spurt involve detrimental effects on cytoskeletal regulation and neuronal maturation in the developing cerebral cortex....

  18. Growth hormone stimulates the collagen synthesis in human tendon and skeletal muscle without affecting myofibrillar protein synthesis

    DEFF Research Database (Denmark)

    Doessing, Simon; Heinemeier, Katja; Holm, Lars

    2010-01-01

    matrix collagen synthesis in skeletal muscle and tendon, but without any effect upon myofibrillar protein synthesis. The results suggest that GH is more important in strengthening the matrix tissue than for muscle cell hypertrophy in adult human musculotendinous tissue.......In skeletal muscle and tendon the extracellular matrix confers important tensile properties and is crucially important for tissue regeneration after injury. Musculoskeletal tissue adaptation is influenced by mechanical loading, which modulates the availability of growth factors, including growth...... young individuals. rhGH administration caused an increase in serum GH, serum IGF-I, and IGF-I mRNA expression in tendon and muscle. Tendon collagen I mRNA expression and tendon collagen protein synthesis increased by 3.9-fold and 1.3-fold, respectively (P muscle collagen I m...

  19. SIRT1 affects DNA methylation of polycomb group protein target genes, a hotspot of the epigenetic shift observed in ageing

    OpenAIRE

    Wakeling, Luisa; Ions, Laura; Escolme, Suzanne; Cockell, Simon,; Su, Tianhong; Dey, Madhurima; Hampton, Emily; Jenkins, Gail; Wainwright, Linda; McKay, Jill; Ford, Dianne

    2015-01-01

    Background SIRT1 is likely to play a role in the extension in healthspan induced by dietary restriction. Actions of SIRT1 are pleiotropic, and effects on healthspan may include effects on DNA methylation. Polycomb group protein target genes (PCGTs) are suppressed by epigenetic mechanisms in stem cells, partly through the actions of the polycomb repressive complexes (PRCs), and have been shown previously to correspond with loci particularly susceptible to age-related changes in DNA methylation...

  20. The predictive accuracy of secondary chemical shifts is more affected by protein secondary structure than solvent environment

    Energy Technology Data Exchange (ETDEWEB)

    Tremblay, Marie-Laurence; Banks, Aaron W.; Rainey, Jan K., E-mail: jan.rainey@dal.c [Dalhousie University, Department of Biochemistry and Molecular Biology (Canada)

    2010-04-15

    Biomolecular NMR spectroscopy frequently employs estimates of protein secondary structure using secondary chemical shift ({Delta}{delta}) values, measured as the difference between experimental and random coil chemical shifts (RCCS). Most published random coil data have been determined in aqueous conditions, reasonable for non-membrane proteins, but potentially less relevant for membrane proteins. Two new RCCS sets are presented here, determined in dimethyl sulfoxide (DMSO) and chloroform:methanol:water (4:4:1 by volume) at 298 K. A web-based program, CS-CHEMeleon, has been implemented to determine the accuracy of secondary structure assessment by calculating and comparing {Delta}{delta} values for various RCCS datasets. Using CS-CHEMeleon, {Delta}{delta} predicted versus experimentally determined secondary structures were compared for large datasets of membrane and non-membrane proteins as a function of RCCS dataset, {Delta}{delta} threshold, nucleus, localized parameter averaging and secondary structure type. Optimized {Delta}{delta} thresholds are presented both for published and for the DMSO and chloroform:methanol:water derived RCCS tables. Despite obvious RCCS variations between datasets, prediction of secondary structure was consistently similar. Strikingly, predictive accuracy seems to be most dependent upon the type of secondary structure, with helices being the most accurately predicted by {Delta}{delta} values using five different RCCS tables. We suggest caution when using {Delta}{delta}-based restraints in structure calculations as the underlying dataset may be biased. Comparative assessment of multiple RCCS datasets should be performed, and resulting {Delta}{delta}-based restraints weighted appropriately relative to other experimental restraints.

  1. Curcumin causes DNA damage and affects associated protein expression in HeLa human cervical cancer cells.

    Science.gov (United States)

    Shang, Hung-Sheng; Chang, Chuan-Hsun; Chou, Yu-Ru; Yeh, Ming-Yang; Au, Man-Kuan; Lu, Hsu-Feng; Chu, Yung-Lin; Chou, Hsiao-Min; Chou, Hsiu-Chen; Shih, Yung-Luen; Chung, Jing-Gung

    2016-10-01

    Cervical cancer is one of the most common cancers in women worldwide and it is a prominent cause of cancer mortality. Curcumin is one of the major compounds from Turmeric and has been shown to induce cytotoxic cell death in human cervical cancer cells. However, there is no study to show curcumin induced DNA damage action via the effect on the DNA damage and repair protein in cervical cancer cells in detail. In this study, we investigated whether or not curcumin induced cell death via DNA damage, chromatin condensation in human cervical cancer HeLa cells by using comet assay and DAPI staining, respectively, we found that curcumin induced cell death through the induction of DNA damage, and chromatin condensation. Western blotting and confocal laser microscopy examination were used to examine the effects of curcumin on protein expression associated with DNA damage, repair and translocation of proteins. We found that curcumin at 13 µM increased the protein levels associated with DNA damage and repair, such as O6-methylguanine-DNA methyltransferase, early-onset breast cancer 1 (BRCA1), mediator of DNA damage checkpoint 1, p-p53 and p-H2A.XSer140 in HeLa cells. Results from confocal laser systems microscopy indicated that curcumin increased the translocation of p-p53 and p-H2A.XSer140 from cytosol to nuclei in HeLa cells. In conclusion, curcumin induced cell death in HeLa cells via induction of DNA damage, and chromatin condensation in vitro.

  2. Insulin-like growth factor binding protein-3 affects osteogenic efficacy on dental implants in rat mandible

    Energy Technology Data Exchange (ETDEWEB)

    Bhattarai, Govinda; Lee, Young-Hee [Department of Oral Biochemistry, Institute of Oral Bioscience, School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Lee, Min-Ho [Department of Dental Materials, Institute of Oral Bioscience, School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of); Park, Il-Song [Division of Advanced Materials Engineering, Research Center for Advanced Materials, Development and Institute of Biodegradable Materials, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Yi, Ho-Keun, E-mail: yihokn@chonbuk.ac.kr [Department of Oral Biochemistry, Institute of Oral Bioscience, School of Dentistry, Chonbuk National University, Jeonju (Korea, Republic of)

    2015-10-01

    Insulin like growth factor binding protein-3 (IGFBP-3) in bone cells and its utilization in dental implants have not been well studied. The aim of this study was to determine the osteogenic efficacy of chitosan gold nanoparticles (Ch-GNPs) conjugated with IGFBP-3 coated titanium (Ti) implants. Ch-GNPs were conjugated with IGFBP-3 plasmid DNA through a coacervation process. Conjugation was cast over Ti surfaces, and cells were seeded on coated surfaces. For in vitro analysis the expression of different proteins was analyzed by immunoblotting. For in vivo analysis, Ch-GNP/IGFBP-3 coated implants were installed in rat mandibles. Four weeks post-implantation, mandibles were examined by microcomputed tomography (μCT), immunohistochemistry, hematoxylin & eosin and tartrate resistance acid phosphatase staining. In vitro overexpressed Ch-GNP/IGFBP-3 coated Ti surfaces was associated with activation of extracellular signal related kinase (ERK), inhibition of the stress activated protein c-Jun N-terminal kinase (JNK) and enhanced bone morphogenetic protein (BMP)-2 and 7 compared to control. Further, in vivo, Ch-GNP/IGFBP-3 coated implants were associated with inhibition of implant induced osteoclastogenesis molecules, receptor activator of nuclear factor kappa-B ligand (RANKL) and enhanced expression of osteogenic molecules including BMP2/7 and osteopontin (OPN). The μCT analysis demonstrated that IGFBP-3 increased the volume of newly formed bone surrounding the implants compared to control (n = 5; p < 0.05). These results support the view that IGFBP-3 overexpression diminishes osteoclastogenesis and enhances osteogenesis of Ti implants, and can serve as a potent molecule for the development of good implantation. - Highlights: • Chitosan gold nanoparticles were conjugated with IGFBP-3 and coated onto surface of the titanium implants for gene delivery to bone. • Implants were inserted in rat mandible for 4 weeks. • Parameters studied: histopathology and radiology.

  3. Yersinia outer protein YopE affects the actin cytoskeleton in Dictyostelium discoideum through targeting of multiple Rho family GTPases

    LENUS (Irish Health Repository)

    Vlahou, Georgia

    2009-07-14

    Abstract Background All human pathogenic Yersinia species share a virulence-associated type III secretion system that translocates Yersinia effector proteins into host cells to counteract infection-induced signaling responses and prevent phagocytosis. Dictyostelium discoideum has been recently used to study the effects of bacterial virulence factors produced by internalized pathogens. In this study we explored the potential of Dictyostelium as model organism for analyzing the effects of ectopically expressed Yersinia outer proteins (Yops). Results The Yersinia pseudotuberculosis virulence factors YopE, YopH, YopM and YopJ were expressed de novo within Dictyostelium and their effects on growth in axenic medium and on bacterial lawns were analyzed. No severe effect was observed for YopH, YopJ and YopM, but expression of YopE, which is a GTPase activating protein for Rho GTPases, was found to be highly detrimental. GFP-tagged YopE expressing cells had less conspicuous cortical actin accumulation and decreased amounts of F-actin. The actin polymerization response upon cAMP stimulation was impaired, although chemotaxis was unaffected. YopE also caused reduced uptake of yeast particles. These alterations are probably due to impaired Rac1 activation. We also found that YopE predominantly associates with intracellular membranes including the Golgi apparatus and inhibits the function of moderately overexpressed RacH. Conclusion The phenotype elicited by YopE in Dictyostelium can be explained, at least in part, by inactivation of one or more Rho family GTPases. It further demonstrates that the social amoeba Dictyostelium discoideum can be used as an efficient and easy-to-handle model organism in order to analyze the function of a translocated GAP protein of a human pathogen.

  4. Ultra-sensitive detection of prion protein fibrils by flow cytometry in blood from cattle affected with bovine spongiform encephalopathy

    Directory of Open Access Journals (Sweden)

    Maas Elke

    2005-10-01

    Full Text Available Abstract Background The definite diagnosis of prion diseases such as Creutzfeldt-Jakob disease (CJD in humans or bovine spongiform encephalopathy (BSE in cattle currently relies on the post mortem detection of the pathological form of the prion protein (PrPSc in brain tissue. Infectivity studies indicate that PrPSc may also be present in body fluids, even at presymptomatic stages of the disease, albeit at concentrations well below the detection limits of currently available analytical methods. Results We developed a highly sensitive method for detecting prion protein aggregates that takes advantage of kinetic differences between seeded and unseeded polymerization of prion protein monomers. Detection of the aggregates was carried out by flow cytometry. In the presence of prion seeds, the association of labelled recombinant PrP monomers in plasma and serum proceeds much more efficiently than in the absence of seeds. In a diagnostic model system, synthetic PrP aggregates were detected down to a concentration of approximately 10-8 nM [0.24 fg/ml]. A specific signal was detected in six out of six available serum samples from BSE-positive cattle. Conclusion We have developed a method based on seed-dependent PrP fibril formation that shows promising results in differentiating a small number of BSE-positive serum samples from healthy controls. This method may provide the basis for an ante mortem diagnostic test for prion diseases.

  5. Phosphorproteome Changes of Myofibrillar Proteins at Early Post-mortem Time in Relation to Pork Quality As Affected by Season.

    Science.gov (United States)

    Li, Xiao; Fang, Tian; Zong, Menghuan; Shi, Xiaoqin; Xu, Xinglian; Dai, Chen; Li, Chunbao; Zhou, Guanghong

    2015-12-02

    The effect of season on phosphorylation of myofibrillar proteins and meat quality of pork longissimus muscles was investigated. Muscle samples were obtained from 40 pork carcasses (10 for each season) at 45 min and 3 and 9 h post-mortem. Myofibrillar proteins were extracted, separated by SDS-PAGE, quantified by phosphor-specific staining, and finally identified by LC-MS/MS. Muscle pH, glycogen, and ATP were measured, and pale, soft, and exudative (PSE) meat was identified by pH value at 45 min post-mortem. A total of 23 bands were detected on SDS-PAGE gels. The phosphorylation levels of bands did not differ between PSE and normal meat. However, the phosphorylation levels of 22 bands were significantly changed by season. Nine of them showed different changes from 45 min to 9 h post-mortem, which were identified to be involved in energy metabolism and sarcomere contraction. Correlation analysis indicated the regulatory progress of these proteins during rigor mortis. These observations contribute to a better understanding of the biochemical processes for the conversion of muscle to meat varying with season.

  6. Nuclear localization of CPI-17, a protein phosphatase-1 inhibitor protein, affects histone H3 phosphorylation and corresponds to proliferation of cancer and smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Eto, Masumi, E-mail: masumi.eto@jefferson.edu [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kirkbride, Jason A.; Chugh, Rishika; Karikari, Nana Kofi [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Kim, Jee In [Department of Molecular Physiology and Biophysics, and Kimmel Cancer Center, Thomas Jefferson University, 1020 Locust Street, PA 19107 (United States); Cardiovascular Research Institute, Kyungpook National University School of Medicine, Daegu 700-422 (Korea, Republic of)

    2013-04-26

    Highlights: •Non-canonical roles of the myosin phosphatase inhibitor (CPI-17) were studied. •CPI-17 is localized in the nucleus of hyperplastic cancer and smooth muscle cells. •CPI-17 Ser12 phosphorylation may regulate the nuclear import. •CPI-17 regulates histone H3 phosphorylation and cell proliferation. •The nuclear CPI-17-PP1 axis plays a proliferative role in cells. -- Abstract: CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17 kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear import. CPI-17 phosphorylated at Ser12 was not localized at nuclei, suggesting a suppressive role of Ser12 phosphorylation in the nuclear import. Activated CPI-17 bound to all three isoforms of PP1 catalytic subunit in Panc1 nuclear extracts. CPI-17 knockdown in Panc1 resulted in dephosphorylation of histone H3 at Thr3, Ser10 and Thr11, whereas it had no effects on the phosphorylation of myosin light chain and merlin, the known targets of MLCP. In parallel, CPI-17 knockdown suppressed Panc1 proliferation. We propose that CPI-17 accumulated in the nucleus through the N-terminal tail targets multiple PP1 signaling pathways regulating cell proliferation.

  7. Nitrite-cured color and phosphate-mediated water binding of pork muscle proteins as affected by calcium in the curing solution.

    Science.gov (United States)

    Zhao, Jing; Xiong, Youling L

    2012-07-01

    Calcium is a mineral naturally present in water and may be included into meat products during processing thereby influencing meat quality. Phosphates improve myofibril swelling and meat water-holding capacity (WHC) but can be sensitive to calcium precipitation. In this study, pork shoulder meat was used to investigate the impact of calcium at 0, 250, and 500 ppm and phosphate type [sodium pyrophosphate (PP), tripolyphosphate (TPP), and hexametaphopshate (HMP)] at 10 mM on nitrite-cured protein extract color at various pH levels (5.5, 6.0, and 6.5) and crude myofibril WHC at pH 6.0. Neither calcium nor phosphates present in the curing brines significantly affected the cured color. Increasing the pH tended to promote the formation of metmyoglobin instead of nitrosylmyoglobin. The ability of PP to enhance myofibril WHC was hampered (P water binding by myofibrils. The depressed muscle fiber swelling responding to added calcium as evidenced by phase contrast microscopy substantiated, to a certain extent, the deleterious effect of calcium, suggesting that hardness of curing water can significantly affect the quality of cured meat products. Although not affecting nitrite-cured color, calcium hampers the efficacy of phosphates to promote water binding by muscle proteins, underscoring the importance of water quality for brine-enhanced meat products. © 2012 Institute of Food Technologists®

  8. Maternal periconceptional and gestational low protein diet affects mouse offspring growth, cardiovascular and adipose phenotype at 1 year of age.

    Directory of Open Access Journals (Sweden)

    Adam J Watkins

    Full Text Available Human and animal studies have revealed a strong association between periconceptional environmental factors, such as poor maternal diet, and an increased propensity for cardiovascular and metabolic disease in adult offspring. Previously, we reported cardiovascular and physiological effects of maternal low protein diet (LPD fed during discrete periods of periconceptional development on 6-month-old mouse offspring. Here, we extend the analysis in 1 year aging offspring, evaluating mechanisms regulating growth and adiposity. Isocaloric LPD (9% casein or normal protein diet (18% casein; NPD was fed to female MF-1 mice either exclusively during oocyte maturation (for 3.5 days prior to mating; Egg-LPD, Egg-NPD, respectively, throughout gestation (LPD, NPD or exclusively during preimplantation development (for 3.5 days post mating; Emb-LPD. LPD and Emb-LPD female offspring were significantly lighter and heavier than NPD females respectively for up to 52 weeks. Egg-LPD, LPD and Emb-LPD offspring displayed significantly elevated systolic blood pressure at 52 weeks compared to respective controls (Egg-NPD, NPD. LPD females had significantly reduced inguinal and retroperitoneal fat pad: body weight ratios compared to NPD females. Expression of the insulin receptor (Insr and insulin-like growth factor I receptor (Igf1r in retroperitoneal fat was significantly elevated in Emb-LPD females (P<0.05, whilst Emb-LPD males displayed significantly decreased expression of the mitochondrial uncoupling protein 1 (Ucp1 gene compared to NPD offspring. LPD females displayed significantly increased expression of Ucp1 in interscapular brown adipose tissue when compared to NPD offspring. Our results demonstrate that aging offspring body weight, cardiovascular and adiposity homeostasis can be programmed by maternal periconceptional nutrition. These adverse outcomes further exemplify the criticality of dietary behaviour around the time of conception on long-term offspring

  9. Ischemia in tumors induces early and sustained phosphorylation changes in stress kinase pathways but does not affect global protein levels

    Energy Technology Data Exchange (ETDEWEB)

    Mertins, Philipp; Yang, Feng; Liu, Tao; Mani, DR; Petyuk, Vladislav A.; Gillette, Michael; Clauser, Karl; Qiao, Jana; Gritsenko, Marina A.; Moore, Ronald J.; Levine, Douglas; Townsend, Reid; Erdmann-Gilmore, Petra; Snider, Jacqueline E.; Davies, Sherri; Ruggles, Kelly; Fenyo, David; Kitchens, R. T.; Li, Shunqiang; Olvera, Narcisco; Dao, Fanny; Rodriguez, Henry; Chan, Daniel W.; Liebler, Daniel; White, Forest; Rodland, Karin D.; Mills, Gordon; Smith, Richard D.; Paulovich, Amanda G.; Ellis, Matthew; Carr, Steven A.

    2014-07-01

    Advances in quantitative mass spectrometry (MS)-based proteomics have sparked efforts to characterize the proteomes of tumor samples to provide complementary and unique information inaccessible by genomics. Tumor samples are usually not accrued with proteomic characterization in mind, raising concerns regarding effects of undocumented sample ischemia on protein abundance and phosphosite stoichiometry. Here we report the effects of cold ischemia time on clinical ovarian cancer samples and patient-derived basal and luminal breast cancer xenografts. Tumor tissues were excised and collected prior to vascular ligation, subjected to accurately defined ischemia times up to 60 min, and analyzed by quantitative proteomics and phosphoproteomics using isobaric tags and high-performance, multidimensional LC-MS/MS. No significant changes were detected at the protein level in each tumor type after 60 minutes of ischemia, and the majority of the >25,000 phosphosites detected were also stable. However, large, reproducible increases and decreases in protein phosphorylation at specific sites were observed in up to 24% of the phosphoproteome starting as early as 5 minutes post-excision. Early and sustained activation of stress response, transcriptional regulation and cell death pathways were observed in common across tumor types. Tissue-specific changes in phosphosite stability were also observed suggesting idiosyncratic effects of ischemia in particular lineages. Our study provides insights into the information that may be obtained by proteomic characterization of tumor samples after undocumented periods of ischemia, and suggests caution especially in interpreting activation of stress pathways in such samples as they may reflect sample handling rather than tumor physiology.

  10. Metabolic induction and early responses of mouse blastocyst developmental programming following maternal low protein diet affecting life-long health.

    Science.gov (United States)

    Eckert, Judith J; Porter, Richard; Watkins, Adam J; Burt, Elizabeth; Brooks, Suzanne; Leese, Henry J; Humpherson, Peter G; Cameron, Iain T; Fleming, Tom P

    2012-01-01

    Previously, we have shown that a maternal low protein diet, fed exclusively during the preimplantation period of mouse development (Emb-LPD), is sufficient to induce by the blastocyst stage a compensatory growth phenotype in late gestation and postnatally, correlating with increased risk of adult onset cardiovascular disease and behavioural dysfunction. Here, we examine mechanisms of induction of maternal Emb-LPD programming and early compensatory responses by the embryo. Emb-LPD induced changes in maternal serum metabolites at the time of blastocyst formation (E3.5), notably reduced insulin and increased glucose, together with reduced levels of free amino acids (AAs) including branched chain AAs leucine, isoleucine and valine. Emb-LPD also caused reduction in the branched chain AAs within uterine fluid at the blastocyst stage. These maternal changes coincided with an altered content of blastocyst AAs and reduced mTORC1 signalling within blastocysts evident in reduced phosphorylation of effector S6 ribosomal protein and its ratio to total S6 protein but no change in effector 4E-BP1 phosphorylated and total pools. These changes were accompanied by increased proliferation of blastocyst trophectoderm and total cells and subsequent increased spreading of trophoblast cells in blastocyst outgrowths. We propose that induction of metabolic programming following Emb-LPD is achieved through mTORC1signalling which acts as a sensor for preimplantation embryos to detect maternal nutrient levels via branched chain AAs and/or insulin availability. Moreover, this induction step associates with changes in extra-embryonic trophectoderm behaviour occurring as early compensatory responses leading to later nutrient recovery.

  11. Central action of xenin affects the expression of lipid metabolism-related genes and proteins in mouse white adipose tissue.

    Science.gov (United States)

    Bhavya, Sharma; Lew, Pei San; Mizuno, Tooru M

    2017-06-01

    Xenin is a gastrointestinal hormone that reduces food intake when administered centrally and it has been hypothesized that central action of xenin participates in the regulation of whole-body metabolism. The present study was performed to address this hypothesis by investigating the central effect of xenin on the expression of genes and proteins that are involved in the regulation of lipid metabolism in white adipose tissue (WAT). Male obese ob/ob mice received intracerebroventricular (i.c.v.) injections of xenin (5μg) twice 12h apart. Food intake and body weight change during a 24-h period after the first injection were measured. Epididymal WAT was collected at the end of the 24-h treatment period and levels of lipid metabolism-related genes and proteins were measured. Xenin treatment caused significant reductions in food intake and body weight compared to control vehicle treatment. Levels of fatty acid synthase (FASN) protein were significantly reduced by xenin treatment, while levels of adipose triglyceride lipase (Atgl) and beta-3 adrenergic receptor (Adrb3) mRNA and phosphorylated hormone sensitive lipase (Ser(660)-pHSL and Ser(563)-pHSL) were significantly increased by xenin treatment. These findings suggest that central action of xenin causes alterations in lipid metabolism in adipose tissue toward reduced lipogenesis and increased lipolysis, possibly contributing to xenin-induced body weight reduction. Thus, enhancing central action of xenin and its downstream targets may be possible targets for the treatment of obesity by reducing the amount of stored fat in adipose tissue. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Identification of residues in ABCG2 affecting protein trafficking and drug transport, using co-evolutionary analysis of ABCG sequences.

    Science.gov (United States)

    Haider, Ameena J; Cox, Megan H; Jones, Natalie; Goode, Alice J; Bridge, Katherine S; Wong, Kelvin; Briggs, Deborah; Kerr, Ian D

    2015-07-17

    ABCG2 is an ABC (ATP-binding cassette) transporter with a physiological role in urate transport in the kidney and is also implicated in multi-drug efflux from a number of organs in the body. The trafficking of the protein and the mechanism by which it recognizes and transports diverse drugs are important areas of research. In the current study, we have made a series of single amino acid mutations in ABCG2 on the basis of sequence analysis. Mutant isoforms were characterized for cell surface expression and function. One mutant (I573A) showed disrupted glycosylation and reduced trafficking kinetics. In contrast with many ABC transporter folding mutations which appear to be 'rescued' by chemical chaperones or low temperature incubation, the I573A mutation was not enriched at the cell surface by either treatment, with the majority of the protein being retained in the endoplasmic reticulum (ER). Two other mutations (P485A and M549A) showed distinct effects on transport of ABCG2 substrates reinforcing the role of TM helix 3 in drug recognition and transport and indicating the presence of intracellular coupling regions in ABCG2. © 2015 Authors.

  13. Okara as a protein supplement affects feed intake and milk composition of ewes and growth performance of lambs

    Directory of Open Access Journals (Sweden)

    Laura B. Harthan

    2017-06-01

    Full Text Available Evaluating the feeding value of wet okara as a protein supplement for lactating ewes with twin lambs was the objective. A 4 × 4 Latin square replicated 2× (4 sheep, 4 treatments, 4 periods per square; 2 squares was conducted to examine the influence of concentrate mix (okara or not and type of forage (silage or hay on ewe milk composition and growth of their lactating lambs. Treatment periods were 14 days (7 days adaptation and 7 days collection. Ewes (55 to 74.8 kg BW were fed 1 of 4 diets: wheat middling and corn concentrate with mixed grass hay (TSH, okara and corn with mixed grass hay (OSH, soybean and wheat middlings with hay crop silage (TSS, and okara and corn with hay crop silage (OSS. Ewes fed hay diets had lower forage dry matter intakes than ewes fed silage. Intake of okara supplement was higher (P < 0.05 with OSH (3.64 kg/d than with OSS (1.70 kg/d. There was no difference in supplement intake between TSH and TSS. There were no differences among diets for lamb daily gains or in ewe milk compositions among the diets. Okara is an effective source of protein for lactating ewes and their twin lambs.

  14. Nuclear magnetic resonance-based metabolomics reveals that dairy protein fractions affect urinary urea excretion differently in overweight adolescents

    DEFF Research Database (Denmark)

    Zheng, Hong; Yde, Christian C.; Dalsgaard, Trine K.

    2015-01-01

    Dairy proteins are an important part of our diet, and recently, there is considerable focus on understanding the effects of the two major dairy proteins fractions constituted by casein and whey. In the present study, the impact of a dietary intervention with casein, whey, and skim milk...... was investigated by using NMR-based urine metabolomics. Overweight adolescents (n = 192; age = 12–15 years; BMI = 25.4 ± 2.3 kg/m2) were randomly assigned to 1 L/day of casein (citrate content: 3.27 mol/L), whey (citrate content: 0.04 mol/L), skim milk, or water for 12 weeks. A significant increase in the urinary...... excretion of urea was found after the 12-week casein and skim milk interventions, while the 12-week whey intervention had no significant effect on the urea excretion. In addition, NMR-based metabolomics revealed a decreased urinary citrate excretion in the whey group and thereby demonstrated its potential...

  15. Comparison of gel properties and biochemical characteristics of myofibrillar protein from bighead carp (Aristichthys nobilis) affected by frozen storage and a hydroxyl radical-generation oxidizing system.

    Science.gov (United States)

    Lu, Han; Zhang, Longteng; Li, Qingzheng; Luo, Yongkang

    2017-05-15

    We wanted to clarify whether gel properties can be affected by in vivo or in vitro myofibrillar protein oxidation and, thus, to provide relevant information and a scientific foundation for the processing of gel products. To accomplish this, we measured the changes in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), total disulfide (SS) content, surface hydrophobicity (So-ANS), carbonyl content, and gel texture and water-holding capacity (WHC) of isolated myofibrillar protein from bighead carp fillets during frozen storage and under different H2O2 concentrations, which were used to represent in vivo and in vitro conditions, respectively. The results indicated that a certain range in content of disulfide crosslinks (0.91mol/105g protein) would promote gel hardness. Mild protein oxidation caused by a certain degree of frozen storage and hydroxyl radicals can promote gel texture and WHC. Based on those results, freezing bighead carp for a certain period can be used to produce gel products. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Human I-mfa domain proteins specifically interact with KSHV LANA and affect its regulation of Wnt signaling-dependent transcription

    Energy Technology Data Exchange (ETDEWEB)

    Kusano, Shuichi, E-mail: skusano@m2.kufm.kagoshima-u.ac.jp [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan); Eizuru, Yoshito [Division of Persistent and Oncogenic Viruses, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima 890-8544 (Japan)

    2010-06-04

    Kaposi's sarcoma-associated herpes virus (KSHV)-encoded latency-associated nuclear antigen (LANA) protein has been reported to interact with glycogen synthase kinase 3{beta} (GSK-3{beta}) and to negatively regulate its activity, leading to stimulation of GSK-3{beta}-dependent {beta}-catenin degradation. We show here that the I-mfa domain proteins, HIC (human I-mfa domain-containing protein) and I-mfa (inhibitor of MyoD family a), interacted in vivo with LANA through their C-terminal I-mfa domains. This interaction affected the intracellular localization of HIC, inhibited the LANA-dependent transactivation of a {beta}-catenin-regulated reporter construct, and decreased the level of the LANA.GSK-3{beta} complex. These data reveal for the first time that I-mfa domain proteins interact with LANA and negatively regulate LANA-mediated activation of Wnt signaling-dependent transcription by inhibiting the formation of the LANA.GSK-3{beta} complex.

  17. Arabidopsis BPM proteins function as substrate adaptors to a cullin3-based E3 ligase to affect fatty acid metabolism in plants.

    Science.gov (United States)

    Chen, Liyuan; Lee, Joo Hyun; Weber, Henriette; Tohge, Takayuki; Witt, Sandra; Roje, Sanja; Fernie, Alisdair R; Hellmann, Hanjo

    2013-06-01

    Regulation of transcriptional processes is a critical mechanism that enables efficient coordination of the synthesis of required proteins in response to environmental and cellular changes. Transcription factors require accurate activity regulation because they play a critical role as key mediators assuring specific expression of target genes. In this work, we show that cullin3-based E3 ligases have the potential to interact with a broad range of ethylene response factor (ERF)/APETALA2 (AP2) transcription factors, mediated by Math-BTB/POZ (for Meprin and TRAF [tumor necrosis factor receptor associated factor] homolog)-Broad complex, Tramtrack, Bric-a-brac/Pox virus and Zinc finger) proteins. The assembly with an E3 ligase causes degradation of their substrates via the 26S proteasome, as demonstrated for the wrinkled1 ERF/AP2 protein. Furthermore, loss of Math-BTB/POZ proteins widely affects plant development and causes altered fatty acid contents in mutant seeds. Overall, this work demonstrates a link between fatty acid metabolism and E3 ligase activities in plants and establishes CUL3-based E3 ligases as key regulators in transcriptional processes that involve ERF/AP2 family members.

  18. Arabidopsis BPM Proteins Function as Substrate Adaptors to a CULLIN3-Based E3 Ligase to Affect Fatty Acid Metabolism in Plants[W

    Science.gov (United States)

    Chen, Liyuan; Lee, Joo Hyun; Weber, Henriette; Tohge, Takayuki; Witt, Sandra; Roje, Sanja; Fernie, Alisdair R.; Hellmann, Hanjo

    2013-01-01

    Regulation of transcriptional processes is a critical mechanism that enables efficient coordination of the synthesis of required proteins in response to environmental and cellular changes. Transcription factors require accurate activity regulation because they play a critical role as key mediators assuring specific expression of target genes. In this work, we show that CULLIN3-based E3 ligases have the potential to interact with a broad range of ETHYLENE RESPONSE FACTOR (ERF)/APETALA2 (AP2) transcription factors, mediated by MATH-BTB/POZ (for Meprin and TRAF [tumor necrosis factor receptor associated factor] homolog)-Broad complex, Tramtrack, Bric-a-brac/Pox virus and Zinc finger) proteins. The assembly with an E3 ligase causes degradation of their substrates via the 26S proteasome, as demonstrated for the WRINKLED1 ERF/AP2 protein. Furthermore, loss of MATH-BTB/POZ proteins widely affects plant development and causes altered fatty acid contents in mutant seeds. Overall, this work demonstrates a link between fatty acid metabolism and E3 ligase activities in plants and establishes CUL3-based E3 ligases as key regulators in transcriptional processes that involve ERF/AP2 family members. PMID:23792371

  19. Treatment with cefotaxime affects expression of conjugation associated proteins and conjugation transfer frequency of an IncI1 plasmid in Escherichia coli

    DEFF Research Database (Denmark)

    Møller, Thea S.B.; Liu, Gang; Boysen, Anders

    2017-01-01

    Horizontal gene transfer (HGT) is the major mechanism responsible for spread of antibiotic resistance. Antibiotic treatment has been suggested to promote HGT, either by directly affecting the conjugation process itself or by selecting for conjugations subsequent to DNA transfer. However, recent......-regulation of the transfer machinery was confirmed at the transcriptional level for five selected genes. The CTX treatment did not cause induction of the SOS-response as revealed by absence of significantly regulated SOS associated proteins in the proteome and no significant up-regulation of recA and sfiA genes...... research suggests that the effect of antibiotic treatment on plasmid conjugation frequencies, and hence the spread of resistance plasmids, may have been overestimated. We addressed the question by quantifying transfer proteins and conjugation frequencies of a blaCTX-M-1 encoding IncI1 resistance plasmid...

  20. Amino acid profile of metabolisable protein in lactating dairy cows is affected by dry matter concentration in grass-clover silage

    DEFF Research Database (Denmark)

    Johansen, Marianne; Lund, Peter; Weisbjerg, Martin Riis

    2018-01-01

    Our previous study showed that supply of metabolisable protein (MP) to lactating dairy cows increased with increasing dry matter (DM) concentration in grass-clover silage. The aim of this study was to examine how amino acid (AA) profile of MP was affected by silage DM concentration. Eight grass......-clover silages with DM concentrations ranging from 283 to 725 g/kg were fed ad libitum to four multi-fistulated dairy cows in an incomplete balanced scheme over five periods. Individual AA were analysed in silages, in microbes isolated from the rumen, and in duodenal and ileal chyme, respectively. Proportion...... of arginine in silage AA increased, proportions of cysteine and proline in silage AA tended to increase, whereas proportions of alanine, histidine, isoleucine, lysine, methionine, ornithine, serine, threonine and valine in silage AA decreased with increased silage DM concentration. Crude protein (CP) and AA...

  1. Signal peptide-CUB-EGF domain-containing protein 1 (SCUBE1) level in hemodialysis patients and parameters affecting that level.

    Science.gov (United States)

    Ulusoy, Sukru; Ozkan, Gulsum; Menteşe, Ahmet; Yavuz, Adnan; Karahan, S Caner; Sümer, Ayşegül Uzun

    2012-11-01

    Signal peptide-CUB (complement C1r/C1s, Uegf, and Bmp1)-EGF (epidermal growth factor)-domain-containing protein 1 (SCUBE1) is a cell surface protein belonging to the SCUBE gene family. SCUBE1 has been shown to rise in parallel with platelet activation in acute ischemic events. However, there are no studies showing levels in the hemodialysis patient group, in which there is known to be an increase in platelet function impairment and activation. The purpose of this study was to investigate SCUBE1 levels in a hemodialysis patient group and the factors affecting those levels. One hundred three hemodialysis patients and 21 age-matched healthy controls were included. SCUBE1 and sCD40L levels were investigated from blood specimens collected on pre- and post-hemodialysis sessions. We investigated the correlation between SCUBE1 levels and sCD40L, patients' demographic data, parameters with hemodialysis treatment and routine biochemical tests. SCUBE1 levels were significantly higher in the hemodialysis patient group compared with the controls (p=0.000). There was a significant rise in SCUBE1 levels in the post-hemodialysis session (p=0.000). We determined a positive correlation between SCUBE1 and sCD40L (p=0.016, r=0.215). Gender, blood pressure, BUN, creatinine, hematocrit and high-sensitivity C-reactive protein (hsCRP) levels, hemodialysis membrane surface area, amount of ultrafiltration, blood flow rate, dialysis flow rate and carnitine use significantly affected SCUBE1 levels. We have shown, for the first time in the literature, that SCUBE1 level, a potential acute ischemia marker, is elevated in hemodialysis patients with no clinical ischemic event, and that various factors affect this elevation. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  2. Long-Term Intake of a High-Protein Diet Affects Body Phenotype, Metabolism, and Plasma Hormones in Mice.

    Science.gov (United States)

    Vu, John P; Luong, Leon; Parsons, William F; Oh, Suwan; Sanford, Daniel; Gabalski, Arielle; Lighton, John Rb; Pisegna, Joseph R; Germano, Patrizia M

    2017-12-01

    Background: High-protein diets (HPDs) recently have been used to obtain body weight and fat mass loss and expand muscle mass. Several studies have documented that HPDs reduce appetite and food intake.Objective: Our goal was to determine the long-term effects of an HPD on body weight, energy intake and expenditure, and metabolic hormones.Methods: Male C57BL/6 mice (8 wk old) were fed either an HPD (60% of energy as protein) or a control diet (CD; 20% of energy as protein) for 12 wk. Body composition and food intakes were determined, and plasma hormone concentrations were measured in mice after being fed and after overnight feed deprivation at several time points.Results: HPD mice had significantly lower body weight (in means ± SEMs; 25.73 ± 1.49 compared with 32.5 ± 1.31 g; P = 0.003) and fat mass (9.55% ± 1.24% compared with 15.78% ± 2.07%; P = 0.05) during the first 6 wk compared with CD mice, and higher lean mass throughout the study starting at week 2 (85.45% ± 2.25% compared with 75.29% ± 1.90%; P = 0.0001). Energy intake, total energy expenditure, and respiratory quotient were significantly lower in HPD compared with CD mice as shown by cumulative energy intake and eating rate. Water vapor was significantly higher in HPD mice during both dark and light phases. In HPD mice, concentrations of leptin [feed-deprived: 41.31 ± 11.60 compared with 3041 ± 683 pg/mL (P = 0.0004); postprandial: 112.5 ± 102.0 compared with 8273 ± 1415 pg/mL (P weight and fat mass loss, but throughout the study preserved body lean mass and significantly reduced energy intake and expenditure as well as leptin and GLP-1 concentrations while elevating postprandial glucagon concentrations. This study suggests that long-term use of HPDs may be an effective strategy to decrease energy intake and expenditure and to maintain body lean mass. © 2017 American Society for Nutrition.

  3. Codon 141 polymorphisms of the ovine prion protein gene affect the phenotype of classical scrapie transmitted from goats to sheep.

    Science.gov (United States)

    Konold, Timm; Phelan, Laura J; Donnachie, Ben R; Chaplin, Melanie J; Cawthraw, Saira; González, Lorenzo

    2017-05-04

    A study to investigate transmission of classical scrapie via goat milk was carried out in sheep: firstly, lambs were challenged orally with goat scrapie brain homogenate to confirm transmission of scrapie from goats to sheep. In the second study phase, milk from scrapie-infected goats was fed to lambs. Lambs were selected according to their prion protein gene (PRNP) genotype, which was either VRQ/VRQ or ARQ/ARQ, with or without additional polymorphisms at codon 141 (FF141, LF141 or LL141) of the ovine PRNP. This report describes the clinical, pathological and molecular phenotype of goat scrapie in those sheep that progressed to clinical end-stage. Ten sheep (six VRQ/VRQ and four ARQ/ARQ, of which three FF141 and one LL141) challenged with one of two scrapie brain homogenates, and six pairs of sheep (ARQ, of which five LL141 and seven LF141) fed milk from six different goats, developed clinical disease, which was characterised by a pruritic (all VRQ/VRQ and LL141 sheep) or a non-pruritic form (all LF141 and FF141 sheep). Immunohistochemical (IHC) examination revealed that the pattern of intra- and extracellular accumulation of disease-associated prion protein in the brain was also dependent on PRNP polymorphisms at codon 141, which was similar in VRQ and LL141 sheep but different from LF141 and FF141 sheep. The influence of codon 141 was also seen in discriminatory Western blot (WB), with LF141 and FF141 sheep showing a bovine spongiform encephalopathy-like profile (diminished reactivity with P4 antibody) on brain tissue. However, discriminatory WB in lymphoid tissues, and IHC pattern and profile both in lymphoid and brain tissue was consistent with classical scrapie in all sheep. This study provided further evidence that the clinical presentation and the pathological and molecular phenotypes of scrapie in sheep are influenced by PRNP polymorphisms, particularly at codon 141. Differences in the truncation of disease-associated prion protein between LL141 sheep and

  4. Nuclear location of tumor suppressor protein maspin inhibits proliferation of breast cancer cells without affecting proliferation of normal epithelial cells

    Science.gov (United States)

    2014-01-01

    Background Maspin, which is classified as a tumor suppressor protein, is downregulated in many types of cancer. Several studies have suggested potential anti-proliferative activity of maspin as well as sensitizing activity of maspin for therapeutic cytotoxic agents in breast cancer tissue culture and animal models. All of the experimental data gathered so far have been based on studies with maspin localized cytoplasmically, while maspin in breast cancer tumor cells may be located in the cytoplasm, nucleus or both. In this study, the effect of maspin cytoplasmic and nuclear location and expression level on breast cancer proliferation and patient survival was studied. Methods Tissue sections from 166 patients with invasive ductal breast cancer were stained by immunohistochemistry for maspin and Ki-67 protein. The localization and expression level of maspin were correlated with estimated patient overall survival and percent of Ki-67-positive cells. In further studies, we created constructs for transient transfection of maspin into breast cancer cells with targeted cytoplasmic and nuclear location. We analyzed the effect of maspin location in normal epithelial cell line MCF10A and three breast cancer cell lines - MCF-7, MDA-MB-231 and SKBR-3 - by immunofluorescence and proliferation assay. Results We observed a strong positive correlation between moderate and high nuclear maspin level and survival of patients. Moreover, a statistically significant negative relationship was observed between nuclear maspin and Ki-67 expression in patients with invasive ductal breast cancer. Spearman’s correlation analysis showed a negative correlation between level of maspin localized in nucleus and percentage of Ki-67 positive cells. No such differences were observed in cells with cytoplasmic maspin. We found a strong correlation between nuclear maspin and loss of Ki-67 protein in breast cancer cell lines, while there was no effect in normal epithelial cells from breast. The anti

  5. The DCR protein TTC3 affects differentiation and Golgi compactness in neurons through specific actin-regulating pathways.

    Directory of Open Access Journals (Sweden)

    Gaia Elena Berto

    Full Text Available In neuronal cells, actin remodeling plays a well known role in neurite extension but is also deeply involved in the organization of intracellular structures, such as the Golgi apparatus. However, it is still not very clear which mechanisms may regulate actin dynamics at the different sites. In this report we show that high levels of the TTC3 protein, encoded by one of the genes of the Down Syndrome Critical Region (DCR, prevent neurite extension and disrupt Golgi compactness in differentiating primary neurons. These effects largely depend on the capability of TTC3 to promote actin polymerization through signaling pathways involving RhoA, ROCK, CIT-N and PIIa. However, the functional relationships between these molecules differ significantly if considering the TTC3 activity on neurite extension or on Golgi organization. Finally, our results reveal an unexpected stage-dependent requirement for F-actin in Golgi organization at different stages of neuronal differentiation.

  6. StarD7 Protein Deficiency Adversely Affects the Phosphatidylcholine Composition, Respiratory Activity, and Cristae Structure of Mitochondria.

    Science.gov (United States)

    Horibata, Yasuhiro; Ando, Hiromi; Zhang, Peixiang; Vergnes, Laurent; Aoyama, Chieko; Itoh, Masahiko; Reue, Karen; Sugimoto, Hiroyuki

    2016-11-25

    Phosphatidylcholine (PC) is a major phospholipid of mitochondria, comprising 40-50% of both the outer and the inner membranes. However, PC must be imported from its production organelles because mitochondria lack the enzymes essential for PC biosynthesis. In a previous study, we found that StarD7 mediates the intracellular transfer of PC to mitochondria. Therefore, in this study, we analyzed the contribution of StarD7 to the maintenance of mitochondrial phospholipid content and function using siRNA-mediated knockdown and knock-out (KO) of the StarD7 gene in HEPA-1 cells. Real time analysis of respiratory activity demonstrated that the oxygen consumption rate and activity of mitochondrial complexes were impaired in StarD7-KD cells. To confirm these results, we established StarD7-KO HEPA-1 cells by double nicking using CRISPR/Cas9n. As expected, StarD7-KD and -KO cells showed a significant reduction in mitochondrial PC content. The ATP level and growth rate of KO cells were notably lower compared with wild-type cells when cultured in glucose-free galactose-containing medium to force cells to rely on mitochondrial ATP production. In KO cells, the level of the MTCO1 protein, a primary subunit of complex IV, was reduced without a concomitant decrease in its mRNA, but the level was restored when StarD7-I was overexpressed. StarD7-KO cells showed impaired formation of the mitochondrial supercomplexes and exhibited a disorganized cristae structure, with no changes in optic atrophy 1 protein. These findings indicate that StarD7 plays important roles in maintaining the proper composition of mitochondrial phospholipids as well as mitochondrial function and morphogenesis. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Overexpression or silencing of FOXO3a affects proliferation of endothelial progenitor cells and expression of cell cycle regulatory proteins.

    Directory of Open Access Journals (Sweden)

    Tiantian Sang

    Full Text Available Endothelial dysfunction is involved in the pathogenesis of many cardiovascular diseases such as atherosclerosis. Endothelial progenitor cells (EPCs have been considered to be of great significance in therapeutic angiogenesis. Furthermore, the Forkhead box O (FOXO transcription factors are known to be important regulators of cell cycle. Therefore, we investigated the effects of changes in FOXO3a activity on cell proliferation and cell cycle regulatory proteins in EPCs. The constructed recombinant adenovirus vectors Ad-TM (triple mutant-FOXO3a, Ad-shRNA-FOXO3a and the control Ad-GFP were transfected into EPCs derived from human umbilical cord blood. Assessment of transfection efficiency using an inverted fluorescence microscope and flow cytometry indicated a successful transfection. Additionally, the expression of FOXO3a was markedly increased in the Ad-TM-FOXO3a group but was inhibited in the Ad-shRNA-FOXO3a group as seen by western blotting. Overexpression of FOXO3a suppressed EPC proliferation and modulated expression of the cell cycle regulatory proteins including upregulation of the cell cycle inhibitor p27(kip1 and downregulation of cyclin-dependent kinase 2 (CDK2, cyclin D1 and proliferating cell nuclear antigen (PCNA. In the Ad-shRNA-FOXO3a group, the results were counter-productive. Furthermore, flow cytometry for cell cycle analysis suggested that the active mutant of FOXO3a caused a noticeable increase in G1- and S-phase frequencies, while a decrease was observed after FOXO3a silencing. In conclusion, these data demonstrated that FOXO3a could possibly inhibit EPC proliferation via cell cycle arrest involving upregulation of p27(kip1 and downregulation of CDK2, cyclin D1 and PCNA.

  8. Polymorphism rs11085226 in the gene encoding polypyrimidine tract-binding protein 1 negatively affects glucose-stimulated insulin secretion.

    Directory of Open Access Journals (Sweden)

    Martin Heni

    Full Text Available OBJECTIVE: Polypyrimidine tract-binding protein 1 (PTBP1 promotes stability and translation of mRNAs coding for insulin secretion granule proteins and thereby plays a role in β-cells function. We studied whether common genetic variations within the PTBP1 locus influence insulin secretion, and/or proinsulin conversion. METHODS: We genotyped 1,502 healthy German subjects for four tagging single nucleotide polymorphisms (SNPs within the PTBP1 locus (rs351974, rs11085226, rs736926, and rs123698 covering 100% of genetic variation with an r(2≥0.8. The subjects were metabolically characterized by an oral glucose tolerance test with insulin, proinsulin, and C-peptide measurements. A subgroup of 320 subjects also underwent an IVGTT. RESULTS: PTBP1 SNP rs11085226 was nominally associated with lower insulinogenic index and lower cleared insulin response in the OGTT (p≤0.04. The other tested SNPs did not show any association with the analyzed OGTT-derived secretion parameters. In the IVGTT subgroup, SNP rs11085226 was accordingly associated with lower insulin levels within the first ten minutes following glucose injection (p = 0.0103. Furthermore, SNP rs351974 was associated with insulin levels in the IVGTT (p = 0.0108. Upon interrogation of MAGIC HOMA-B data, our rs11085226 result was replicated (MAGIC p = 0.018, but the rs351974 was not. CONCLUSIONS: We conclude that common genetic variation in PTBP1 influences glucose-stimulated insulin secretion. This underlines the importance of PTBP1 for beta cell function in vivo.

  9. Aire knockdown in medullary thymic epithelial cells affects Aire protein, deregulates cell adhesion genes and decreases thymocyte interaction.

    Science.gov (United States)

    Pezzi, Nicole; Assis, Amanda Freire; Cotrim-Sousa, Larissa Cotrim; Lopes, Gabriel Sarti; Mosella, Maritza Salas; Lima, Djalma Sousa; Bombonato-Prado, Karina F; Passos, Geraldo Aleixo

    2016-09-01

    We demonstrate that even a partial reduction of Aire mRNA levels by siRNA-induced Aire knockdown (Aire KD) has important consequences to medullary thymic epithelial cells (mTECs). Aire knockdown is sufficient to reduce Aire protein levels, impair its nuclear location, and cause an imbalance in large-scale gene expression, including genes that encode cell adhesion molecules. These genes drew our attention because adhesion molecules are implicated in the process of mTEC-thymocyte adhesion, which is critical for T cell development and the establishment of central self-tolerance. Accordingly, we consider the following: 1) mTECs contribute to the elimination of self-reactive thymocytes through adhesion; 2) Adhesion molecules play a crucial role during physical contact between these cells; and 3) Aire is an important transcriptional regulator in mTECs. However, its role in controlling mTEC-thymocyte adhesion remains unclear. Because Aire controls adhesion molecule genes, we hypothesized that the disruption of its expression could influence mTEC-thymocyte interaction. To test this hypothesis, we used a murine Aire(+) mTEC cell line as a model system to reproduce mTEC-thymocyte adhesion in vitro. Transcriptome analysis of the mTEC cell line revealed that Aire KD led to the down-modulation of more than 800 genes, including those encoding for proteins involved in cell adhesion, i.e., the extracellular matrix constituent Lama1, the CAM family adhesion molecules Vcam1 and Icam4, and those that encode peripheral tissue antigens. Thymocytes co-cultured with Aire KD mTECs had a significantly reduced capacity to adhere to these cells. This finding is the first direct evidence that Aire also plays a role in controlling mTEC-thymocyte adhesion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Adaptor Protein-1 Complex Affects the Endocytic Trafficking and Function of Peptidylglycine α-Amidating Monooxygenase, a Luminal Cuproenzyme*

    Science.gov (United States)

    Bonnemaison, Mathilde L.; Bäck, Nils; Duffy, Megan E.; Ralle, Martina; Mains, Richard E.; Eipper, Betty A.

    2015-01-01

    The adaptor protein-1 complex (AP-1), which transports cargo between the trans-Golgi network and endosomes, plays a role in the trafficking of Atp7a, a copper-transporting P-type ATPase, and peptidylglycine α-amidating monooxygenase (PAM), a copper-dependent membrane enzyme. Lack of any of the four AP-1 subunits impairs function, and patients with MEDNIK syndrome, a rare genetic disorder caused by lack of expression of the σ1A subunit, exhibit clinical and biochemical signs of impaired copper homeostasis. To explore the role of AP-1 in copper homeostasis in neuroendocrine cells, we used corticotrope tumor cells in which AP-1 function was diminished by reducing expression of its μ1A subunit. Copper levels were unchanged when AP-1 function was impaired, but cellular levels of Atp7a declined slightly. The ability of PAM to function was assessed by monitoring 18-kDa fragment-NH2 production from proopiomelanocortin. Reduced AP-1 function made 18-kDa fragment amidation more sensitive to inhibition by bathocuproine disulfonate, a cell-impermeant Cu(I) chelator. The endocytic trafficking of PAM was altered, and PAM-1 accumulated on the cell surface when AP-1 levels were reduced. Reduced AP-1 function increased the Atp7a presence in early/recycling endosomes but did not alter the ability of copper to stimulate its appearance on the plasma membrane. Co-immunoprecipitation of a small fraction of PAM and Atp7a supports the suggestion that copper can be transferred directly from Atp7a to PAM, a process that can occur only when both proteins are present in the same subcellular compartment. Altered luminal cuproenzyme function may contribute to deficits observed when the AP-1 function is compromised. PMID:26170456

  11. Alzheimer-specific variants in the 3'UTR of Amyloid precursor protein affect microRNA function.

    Science.gov (United States)

    Delay, Charlotte; Calon, Frédéric; Mathews, Paul; Hébert, Sébastien S

    2011-10-07

    APP expression misregulation can cause genetic Alzheimer's disease (AD). Recent evidences support the hypothesis that polymorphisms located in microRNA (miRNA) target sites could influence the risk of developing neurodegenerative disorders such as Parkinson's disease (PD) and frontotemporal dementia. Recently, a number of single nucleotide polymorphisms (SNPs) located in the 3'UTR of APP have been found in AD patients with family history of dementia. Because miRNAs have previously been implicated in APP expression regulation, we set out to determine whether these polymorphisms could affect miRNA function and therefore APP levels. Bioinformatics analysis identified twelve putative miRNA bindings sites located in or near the APP 3'UTR variants T117C, A454G and A833C. Among those candidates, seven miRNAs, including miR-20a, miR-17, miR-147, miR-655, miR-323-3p, miR-644, and miR-153 could regulate APP expression in vitro and under physiological conditions in cells. Using luciferase-based assays, we could show that the T117C variant inhibited miR-147 binding, whereas the A454G variant increased miR-20a binding, consequently having opposite effects on APP expression. Taken together, our results provide proof-of-principle that APP 3'UTR polymorphisms could affect AD risk through modulation of APP expression regulation, and set the stage for further association studies in genetic and sporadic AD.

  12. Palmitoylcarnitine affects localization of growth associated protein GAP-43 in plasma membrane subdomains and its interaction with Gα(o) in neuroblastoma NB-2a cells.

    Science.gov (United States)

    Tułodziecka, Karolina; Czeredys, Magdalena; Nałęcz, Katarzyna A

    2013-03-01

    Palmitoylcarnitine was observed previously to promote differentiation of neuroblastoma NB-2a cells, and to affect protein kinase C (PKC). Palmitoylcarnitine was also observed to increase palmitoylation of several proteins, including a PKC substrate, whose expression augments during differentiation of neural cells-a growth associated protein GAP-43, known to bind phosphatidylinositol 4,5-bisphosphate [PI(4,5)P(2)]. Since palmitoylated proteins are preferentially localized in sphingolipid- and cholesterol-rich microdomains of plasma membrane, the present study has been focused on a possible effect of palmitoylcarnitine on GAP-43 localization in these microdomains. Palmitoylcarnitine treatment resulted in GAP-43 appearance in floating fractions (rafts) in sucrose gradient and increased co-localization with cholesterol and with PI(4,5)P(2), although co-localization of both lipids decreased. GAP-43 disappeared from raft fraction upon treatment with 2-bromopalmitate (an inhibitor of palmitoylating enzymes) and after treatment with etomoxir (carnitine palmitoyltransferase I inhibitor). Raft localization of GAP-43 was completely abolished by treatment with methyl-β-cyclodextrin, a cholesterol binding agent, while there was no change upon sequestration of PI(4,5)P(2) with neomycin. GAP-43 co-precipitated with a monomeric form of Gα(o), a phenomenon diminished after palmitoylcarnitine treatment and paralleled by a decrease of Gα(o) in the raft fraction. These observations point to palmitoylation of GAP-43 as a mechanism leading to an increased localization of this protein in microdomains of plasma membrane rich in cholesterol, in majority different, however, from microdomains in which PI(4,5)P(2) is present. This localization correlates with decreased interaction with Gα(o) and suppression of its activity-an important step regulating neural cell differentiation.

  13. A gestational high protein diet affects the abundance of muscle transcripts related to cell cycle regulation throughout development in porcine progeny.

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    Michael Oster

    Full Text Available BACKGROUND: In various animal models pregnancy diets have been shown to affect offspring phenotype. Indeed, the underlying programming of development is associated with modulations in birth weight, body composition, and continual diet-dependent modifications of offspring metabolism until adulthood, producing the hypothesis that the offspring's transcriptome is permanently altered depending on maternal diet. METHODOLOGY/PRINCIPAL FINDINGS: To assess alterations of the offspring's transcriptome due to gestational protein supply, German Landrace sows were fed isoenergetic diets containing protein levels of either 30% (high protein--HP or 12% (adequate protein--AP throughout their pregnancy. Offspring muscle tissue (M. longissimus dorsi was collected at 94 days post conception (dpc, and 1, 28, and 188 days post natum (dpn for use with Affymetrix GeneChip Porcine Genome Arrays and subsequent statistical and Ingenuity pathway analyses. Numerous transcripts were found to have altered abundance at 94 dpc and 1 dpn; at 28 dpn no transcripts were altered, and at 188 dpn only a few transcripts showed a different abundance between diet groups. However, when assessing transcriptional changes across developmental time points, marked differences were obvious among the dietary groups. Depending on the gestational dietary exposure, short- and long-term effects were observed for mRNA expression of genes related to cell cycle regulation, energy metabolism, growth factor signaling pathways, and nucleic acid metabolism. In particular, the abundance of transcripts related to cell cycle remained divergent among the groups during development. CONCLUSION: Expression analysis indicates that maternal protein supply induced programming of the offspring's genome; early postnatal compensation of the slight growth retardation obvious at birth in HP piglets resulted, as did a permanently different developmental alteration and responsiveness to the common environment of the

  14. Dietary protein sources differentially affect microbiota, mTOR activity and transcription of mTOR signaling pathways in the small intestine.

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    Soumya K Kar

    Full Text Available Dietary protein sources can have profound effects on host-microbe interactions in the gut that are critically important for immune resilience. However more knowledge is needed to assess the impact of different protein sources on gut and animal health. Thirty-six wildtype male C57BL/6J mice of 35 d age (n = 6/group; mean ± SEM body weight 21.9 ± 0.25 g were randomly assigned to groups fed for four weeks with semi synthetic diets prepared with one of the following protein sources containing (300 g/kg as fed basis: soybean meal (SBM, casein, partially delactosed whey powder, spray dried plasma protein, wheat gluten meal and yellow meal worm. At the end of the experiment, mice were sacrificed to collect ileal tissue to acquire gene expression data, and mammalian (mechanistic target of rapamycin (mTOR activity, ileal digesta to study changes in microbiota and serum to measure cytokines and chemokines. By genome-wide transcriptome analysis, we identified fourteen high level regulatory genes that are strongly affected in SBM-fed mice compared to the other experimental groups. They mostly related to the mTOR pathway. In addition, an increased (P < 0.05 concentration of granulocyte colony-stimulating factor was observed in serum of SBM-fed mice compared to other dietary groups. Moreover, by 16S rRNA sequencing, we observed that SBM-fed mice had higher (P < 0.05 abundances of Bacteroidales family S24-7, compared to the other dietary groups. We showed that measurements of genome-wide expression and microbiota composition in the mouse ileum reveal divergent responses to diets containing different protein sources, in particular for a diet based on SBM.

  15. Alpha7-nicotinic acetylcholine receptors affect growth regulation of human mesothelioma cells: role of mitogen-activated protein kinase pathway.

    Science.gov (United States)

    Trombino, Sonya; Cesario, Alfredo; Margaritora, Stefano; Granone, PierLuigi; Motta, Giovanni; Falugi, Carla; Russo, Patrizia

    2004-01-01

    This study presents data suggesting that both human mesothelioma (cell lines and human mesothelioma biopsies) and human normal mesothelial cells express receptors for acetylcholine and that stimulation of these receptors by nicotine prompted cell growth via activation of nicotinic cholinergic receptors. Thus, these data demonstrate that: (a) human mesothelioma cells and human biopsies of mesothelioma as well as of normal pleural mesothelial cells express functionally alpha-7 nicotinic acethlycholine receptors, evaluated by alpha-bungarotoxin-FITC binding, receptor binding assay, Western blot, and reverse transcription-PCR; (b) choline acetyltransferase immunostaining is present in mesothelioma cells; (c) mesothelioma cell growth is modulated by the cholinergic system in which agonists (i.e., nicotine) has a proliferative effect, and antagonists (i.e., curare) has an inhibitory effect, evaluated by cell cloning, DNA synthesis and cell cycle; (d) nicotine induces Ca(+2) influx, evaluated by [(45)Ca(2+)] uptake, and consequently activation of mitogen-activated protein kinase pathway (extracellular signal-regulated kinase and p90(RSK) phosphorylation), evaluated by Western blot; and (e) apoptosis mechanisms in mesothelioma cells are under the control of the cholinergic system (nicotine antiapoptotic via induction of nuclear factor-kappaB complexes and phosphorylation of Bad at Ser(112); curare proapoptotic via G(0)-G(1) arrest p21(waf-1) dependent but p53 independent). The involvement of the nonneuronal cholinergic system in mesothelioma appears reasonable and open up new therapeutic strategies.

  16. Cellular Retinoic Acid-Binding Protein 1 Modulates Stem Cell Proliferation to Affect Learning and Memory in Male Mice.

    Science.gov (United States)

    Lin, Yu-Lung; Persaud, Shawna D; Nhieu, Jennifer; Wei, Li-Na

    2017-09-01

    Retinoic acid (RA) is the active ingredient of vitamin A. It exerts its canonical activity by binding to nuclear RA receptors (RARs) to regulate gene expression. Increasingly, RA is also known to elicit nongenomic RAR-independent activities, most widely detected in activating extracellular regulated kinase (ERK)1/2. This study validated the functional role of cellular retinoic acid-binding protein 1 (Crabp1) in mediating nongenomic activity in RA, specifically activating ERK1/2 to rapidly augment the cell cycle by expanding the growth 1 phase and slowing down embryonic stem cell and neural stem cell (NSC) proliferation. The study further uncovered the physiological activity of Crabp1 in modulating NSC proliferation and animal behavior. In the Crabp1 knockout mouse hippocampus, where Crabp1 is otherwise detected in the subgranular zone, neurogenesis and NSC proliferation increased and hippocampus-dependent brain functions such as learning and memory correspondingly improved. This study established the physiological role of Crabp1 in modulating stem cell proliferation and hippocampus-dependent brain activities such as learning and memory. Copyright © 2017 Endocrine Society.

  17. Mouse Prion Protein Polymorphism Phe-108/Val-189 Affects the Kinetics of Fibril Formation and the Response to Seeding

    Science.gov (United States)

    Cortez, Leonardo M.; Kumar, Jitendra; Renault, Ludovic; Young, Howard S.; Sim, Valerie L.

    2013-01-01

    Prion diseases are fatal neurodegenerative disorders associated with the polymerization of the cellular form of prion protein (PrPC) into an amyloidogenic β-sheet infectious form (PrPSc). The sequence of host PrP is the major determinant of host prion disease susceptibility. In mice, the presence of allele a (Prnpa, encoding the polymorphism Leu-108/Thr-189) or b (Prnpb, Phe-108/Val-189) is associated with short or long incubation times, respectively, following infection with PrPSc. The molecular bases linking PrP sequence, infection susceptibility, and convertibility of PrPC into PrPSc remain unclear. Here we show that recombinant PrPa and PrPb aggregate and respond to seeding differently in vitro. Our kinetic studies reveal differences during the nucleation phase of the aggregation process, where PrPb exhibits a longer lag phase that cannot be completely eliminated by seeding the reaction with preformed fibrils. Additionally, PrPb is more prone to propagate features of the seeds, as demonstrated by conformational stability and electron microscopy studies of the formed fibrils. We propose a model of polymerization to explain how the polymorphisms at positions 108 and 189 produce the phenotypes seen in vivo. This model also provides insight into phenomena such as species barrier and prion strain generation, two phenomena also influenced by the primary structure of PrP. PMID:23283973

  18. transparent, a gene affecting stripe formation in Zebrafish, encodes the mitochondrial protein Mpv17 that is required for iridophore survival

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    Jana Krauss

    2013-06-01

    In the skin of adult zebrafish, three pigment cell types arrange into alternating horizontal stripes, melanophores in dark stripes, xanthophores in light interstripes and iridophores in both stripes and interstripes. The analysis of mutants and regeneration studies revealed that this pattern depends on interactions between melanophores and xanthophores; however, the role of iridophores in this process is less understood. We describe the adult viable and fertile mutant transparent (tra, which shows a loss or strong reduction of iridophores throughout larval and adult stages. In addition, in adults only the number of melanophores is strongly reduced, and stripes break up into spots. Stripes in the fins are normal. By cell transplantations we show that tra acts cell-autonomously in iridophores, whereas the reduction in melanophores in the body occurs secondarily as a consequence of iridophore loss. We conclude that differentiated iridophores are required for the accumulation and maintenance of melanophores during pigment pattern formation. The tra mutant phenotype is caused by a small deletion in mpv17, an ubiquituously expressed gene whose protein product, like its mammalian and yeast homologs, localizes to mitochondria. Iridophore death might be the result of mitochondrial dysfunction, consistent with the mitochondrial DNA depletion syndrome observed in mammalian mpv17 mutants. The specificity of the tra phenotype is most likely due to redundancy after gene multiplication, making this mutant a valuable model to understand the molecular function of Mpv17 in mitochondria.

  19. SpyB, a Small Heme-Binding Protein, Affects the Composition of the Cell Wall in Streptococcus pyogenes.

    Science.gov (United States)

    Edgar, Rebecca J; Chen, Jing; Kant, Sashi; Rechkina, Elena; Rush, Jeffrey S; Forsberg, Lennart S; Jaehrig, Bernhard; Azadi, Parastoo; Tchesnokova, Veronika; Sokurenko, Evgeni V; Zhu, Haining; Korotkov, Konstantin V; Pancholi, Vijay; Korotkova, Natalia

    2016-01-01

    Streptococcus pyogenes (Group A Streptococcus or GAS) is a hemolytic human pathogen associated with a wide variety of infections ranging from minor skin and throat infections to life-threatening invasive diseases. The cell wall of GAS consists of peptidoglycan sacculus decorated with a carbohydrate comprising a polyrhamnose backbone with immunodominant N-acetylglucosamine side-chains. All GAS genomes contain the spyBA operon, which encodes a 35-amino-acid membrane protein SpyB, and a membrane-bound C3-like ADP-ribosyltransferase SpyA. In this study, we addressed the function of SpyB in GAS. Phenotypic analysis of a spyB deletion mutant revealed increased bacterial aggregation, and reduced sensitivity to β-lactams of the cephalosporin class and peptidoglycan hydrolase PlyC. Glycosyl composition analysis of cell wall isolated from the spyB mutant suggested an altered carbohydrate structure compared with the wild-type strain. Furthermore, we found that SpyB associates with heme and protoporphyrin IX. Heme binding induces SpyB dimerization, which involves disulfide bond formation between the subunits. Thus, our data suggest the possibility that SpyB activity is regulated by heme.

  20. Protein turnover and cellular stress in mildly and severely affected muscles from patients with limb girdle muscular dystrophy type 2I.

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    Hauerslev, Simon; Sveen, Marie L; Vissing, John; Krag, Thomas O

    2013-01-01

    Patients with Limb girdle muscular dystrophy type 2I (LGMD2I) are characterized by progressive muscle weakness and wasting primarily in the proximal muscles, while distal muscles often are spared. Our aim was to investigate if wasting could be caused by impaired regeneration in the proximal compared to distal muscles. Biopsies were simultaneously obtained from proximal and distal muscles of the same patients with LGMD2I (n = 4) and healthy subjects (n = 4). The level of past muscle regeneration was evaluated by counting internally nucleated fibers and determining actively regenerating fibers by using the developmental markers embryonic myosin heavy chain (eMHC) and neural cell adhesion molecule (NCAM) and also assessing satellite cell activation status by myogenin positivity. Severe muscle histopathology was occasionally observed in the proximal muscles of patients with LGMD2I whereas distal muscles were always relatively spared. No difference was found in the regeneration markers internally nucleated fibers, actively regenerating fibers or activation status of satellite cells between proximal and distal muscles. Protein turnover, both synthesis and breakdown, as well as cellular stress were highly increased in severely affected muscles compared to mildly affected muscles. Our results indicate that alterations in the protein turnover and myostatin levels could progressively impair the muscle mass maintenance and/or regeneration resulting in gradual muscular atrophy.

  1. The challenging environment on board the International Space Station affects endothelial cell function by triggering oxidative stress through thioredoxin interacting protein overexpression: the ESA-SPHINX experiment.

    Science.gov (United States)

    Versari, Silvia; Longinotti, Giulia; Barenghi, Livia; Maier, Jeanette Anne Marie; Bradamante, Silvia

    2013-11-01

    Exposure to microgravity generates alterations that are similar to those involved in age-related diseases, such as cardiovascular deconditioning, bone loss, muscle atrophy, and immune response impairment. Endothelial dysfunction is the common denominator. To shed light on the underlying mechanism, we participated in the Progress 40P mission with Spaceflight of Human Umbilical Vein Endothelial Cells (HUVECs): an Integrated Experiment (SPHINX), which consisted of 12 in-flight and 12 ground-based control modules and lasted 10 d. Postflight microarray analysis revealed 1023 significantly modulated genes, the majority of which are involved in cell adhesion, oxidative phosphorylation, stress responses, cell cycle, and apoptosis. Thioredoxin-interacting protein was the most up-regulated (33-fold), heat-shock proteins 70 and 90 the most down-regulated (5.6-fold). Ion channels (TPCN1, KCNG2, KCNJ14, KCNG1, KCNT1, TRPM1, CLCN4, CLCA2), mitochondrial oxidative phosphorylation, and focal adhesion were widely affected. Cytokine detection in the culture media indicated significant increased secretion of interleukin-1α and interleukin-1β. Nitric oxide was found not modulated. Our data suggest that in cultured HUVECs, microgravity affects the same molecular machinery responsible for sensing alterations of flow and generates a prooxidative environment that activates inflammatory responses, alters endothelial behavior, and promotes senescence.

  2. Lactation Affects Isolated Mitochondria and Its Fatty Acid Composition but Has No Effect on Tissue Protein Oxidation, Lipid Peroxidation or DNA-Damage in Laboratory Mice

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    Teresa G. Valencak

    2016-01-01

    Full Text Available Linking peak energy metabolism to lifespan and aging remains a major question especially when focusing on lactation in females. We studied, if and how lactation affects in vitro mitochondrial oxygen consumption and mitochondrial fatty acid composition. In addition, we assessed DNA damage, lipid peroxidation and protein carbonyls to extrapolate on oxidative stress in mothers. As model system we used C57BL/6NCrl mice and exposed lactating females to two ambient temperatures (15 °C and 22 °C while they nursed their offspring until weaning. We found that state II and state IV respiration rates of liver mitochondria were significantly higher in the lactating animals than in non-lactating mice. Fatty acid composition of isolated liver and heart mitochondria differed between lactating and non-lactating mice with higher n-6, and lower n-3 polyunsaturated fatty acids in the lactating females. Surprisingly, lactation did not affect protein carbonyls, lipid peroxidation and DNA damage, nor did moderate cold exposure of 15 °C. We conclude that lactation increases rates of mitochondrial uncoupling and alters mitochondrial fatty acid composition thus supporting the “uncoupling to survive” hypothesis. Regarding oxidative stress, we found no impact of lactation and lower ambient temperature and contribute to growing evidence that there is no linear relationship between oxidative damage and lactation.

  3. Whole-body electromyostimulation and protein supplementation favorably affect sarcopenic obesity in community-dwelling older men at risk: the randomized controlled FranSO study.

    Science.gov (United States)

    Kemmler, Wolfgang; Weissenfels, Anja; Teschler, Marc; Willert, Sebastian; Bebenek, Michael; Shojaa, Mahdieh; Kohl, Matthias; Freiberger, Ellen; Sieber, Cornel; von Stengel, Simon

    2017-01-01

    Sarcopenic obesity (SO) is a geriatric syndrome characterized by the disproportion between the amount of lean mass and fat mass. Exercise decreases fat and maintains muscle mass; however, older people fail to exercise at doses sufficient to affect musculoskeletal and cardiometabolic risk factors. The aim of this study was to evaluate the effect of whole-body electromyostimulation (WB-EMS), a time-efficient, joint-friendly and highly individualized exercise technology, on sarcopenia and SO in older men. A total of 100 community-dwelling northern Bavarian men aged ≥70 years with sarcopenia and obesity were randomly (1-1-1) assigned to either 16 weeks of 1) WB-EMS and protein supplementation (WB-EMS&P), 2) isolated protein supplementation or 3) nonintervention control. WB-EMS consisted of 1.5×20 min (85 Hz, 350 µs, 4 s of strain to 4 s of rest) applied with moderate-to-high intensity while moving. We further generated a daily protein intake of 1.7-1.8 g/kg/body mass per day. The primary study end point was Sarcopenia Z-Score, and the secondary study end points were body fat rate (%), skeletal muscle mass index (SMI) and handgrip strength. Intention-to-treat analysis determined a significantly favorable effect of WB-EMS&P (P<0.001) and protein (P=0.007) vs control. Both groups significantly (P<0.001) lost body fat (WB-EMS&P: 2.1%; protein: 1.1%) and differed significantly (P≤0.004) from control (0.3%). Differences between WB-EMS&P and protein were significant for the Sarcopenia Z-Score (P=0.39) and borderline nonsignificant (P=0.051) for body fat. SMI increased significantly in both groups (P<0.001 and P=0.043) and decreased significantly in the control group (CG; P=0.033); differences between the verum groups and control were significant (P≤0.009). Handgrip strength increased in the WB-EMS group (1.90 kg; P<0.001; P=0.050 vs control) only. No adverse effects of WB-EMS or protein supplementation were recorded. WB-EMS&P is a safe and efficient method for

  4. Whole-body electromyostimulation and protein supplementation favorably affect sarcopenic obesity in community-dwelling older men at risk: the randomized controlled FranSO study

    Science.gov (United States)

    Kemmler, Wolfgang; Weissenfels, Anja; Teschler, Marc; Willert, Sebastian; Bebenek, Michael; Shojaa, Mahdieh; Kohl, Matthias; Freiberger, Ellen; Sieber, Cornel; von Stengel, Simon

    2017-01-01

    Background Sarcopenic obesity (SO) is a geriatric syndrome characterized by the disproportion between the amount of lean mass and fat mass. Exercise decreases fat and maintains muscle mass; however, older people fail to exercise at doses sufficient to affect musculoskeletal and cardiometabolic risk factors. The aim of this study was to evaluate the effect of whole-body electromyostimulation (WB-EMS), a time-efficient, joint-friendly and highly individualized exercise technology, on sarcopenia and SO in older men. Materials and methods A total of 100 community-dwelling northern Bavarian men aged ≥70 years with sarcopenia and obesity were randomly (1–1–1) assigned to either 16 weeks of 1) WB-EMS and protein supplementation (WB-EMS&P), 2) isolated protein supplementation or 3) nonintervention control. WB-EMS consisted of 1.5×20 min (85 Hz, 350 µs, 4 s of strain to 4 s of rest) applied with moderate-to-high intensity while moving. We further generated a daily protein intake of 1.7–1.8 g/kg/body mass per day. The primary study end point was Sarcopenia Z-Score, and the secondary study end points were body fat rate (%), skeletal muscle mass index (SMI) and handgrip strength. Results Intention-to-treat analysis determined a significantly favorable effect of WB-EMS&P (P<0.001) and protein (P=0.007) vs control. Both groups significantly (P<0.001) lost body fat (WB-EMS&P: 2.1%; protein: 1.1%) and differed significantly (P≤0.004) from control (0.3%). Differences between WB-EMS&P and protein were significant for the Sarcopenia Z-Score (P=0.39) and borderline nonsignificant (P=0.051) for body fat. SMI increased significantly in both groups (P<0.001 and P=0.043) and decreased significantly in the control group (CG; P=0.033); differences between the verum groups and control were significant (P≤0.009). Handgrip strength increased in the WB-EMS group (1.90 kg; P<0.001; P=0.050 vs control) only. No adverse effects of WB-EMS or protein supplementation were recorded

  5. Tick Thioester-Containing Proteins and Phagocytosis Do Not Affect Transmission of Borrelia afzelii from the Competent Vector Ixodes ricinus.

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    Urbanová, Veronika; Hajdušek, Ondřej; Hönig Mondeková, Helena; Šíma, Radek; Kopáček, Petr

    2017-01-01

    The present concept of the transmission of Lyme disease from Borrelia-infected Ixodes sp. ticks to the naïve host assumes that a low number of spirochetes that manage to penetrate the midgut epithelium migrate through the hemocoel to the salivary glands and subsequently infect the host with the aid of immunomodulatory compounds present in tick saliva. Therefore, humoral and/or cellular immune reactions within the tick hemocoel may play an important role in tick competence to act as a vector for borreliosis. To test this hypothesis we have examined complement-like reactions in the hemolymph of the hard tick Ixodes ricinus against Borrelia afzelii (the most common vector and causative agent of Lyme disease in Europe). We demonstrate that I. ricinus hemolymph does not exhibit borreliacidal effects comparable to complement-mediated lysis of bovine sera. However, after injection of B. afzelii into the tick hemocoel, the spirochetes were efficiently phagocytosed by tick hemocytes and this cellular defense was completely eliminated by pre-injection of latex beads. As tick thioester-containing proteins (T-TEPs) are components of the tick complement system, we performed RNAi-mediated silencing of all nine genes encoding individual T-TEPs followed by in vitro phagocytosis assays. Silencing of two molecules related to the C3 complement component (IrC3-2 and IrC3-3) significantly suppressed phagocytosis of B. afzelii, while knockdown of IrTep (insect type TEP) led to its stimulation. However, RNAi-mediated silencing of T-TEPs or elimination of phagocytosis by injection of latex beads in B. afzelii-infected I. ricinus nymphs had no obvious impact on the transmission of spirochetes to naïve mice, as determined by B. afzelii infection of murine tissues following tick infestation. This result supports the concept that Borrelia spirochetes are capable of avoiding complement-related reactions within the hemocoel of ticks competent to transmit Lyme disease.

  6. Diabetes mellitus affects activity of calcium/calmodulin-dependent protein kinase II alpha in rat trigeminal ganglia.

    Science.gov (United States)

    Jerić, Milka; Vuica, Ana; Borić, Matija; Puljak, Livia; Jeličić Kadić, Antonia; Grković, Ivica; Filipović, Natalija

    2015-01-01

    The activity of calcium/calmodulin-dependent protein kinase II alpha (CaMKIIα) may play a critical role in the modulation of nociceptor activity and plasticity of primary sensory trigeminal neurons. The aim of this study was to investigate the immunoreactivity of phosphorylated CaMKIIα (pCaMKIIα) in subpopulations of trigeminal ganglion (TG) neurons in rat models of early diabetes type 1 (dm1) and 2 (dm2). DM1 model was induced with intraperitoneally (i.p.) injected streptozotocin (STZ) (55mg/kg). DM2 rats were fed with the high fat diet (HFD) for 2 weeks and then received 35mg/kg of STZ i.p. Two weeks and 2 months after the STZ-diabetes induction, rats were sacrificed and immunohistochemical analysis for detection of pCaMKIIα immunoreactivity and double immunofluorescence labelling with isolectin (IB4) was performed. Increased intensity of pCaMKIIα immunofluorescence, restricted to IB4-negative small-diameter neurons, was seen in TG neurons two months after STZ-DM1 induction. DM1 model, as well as the obesity (control dm2 groups) resulted in neuronal impaired growth while dm2 model led to neuron hypertrophy in TG. Observed changes may play a critical role in the modulation of nociceptor activity and plasticity of primary sensory trigeminal neurons. In future, innovative strategies for modulation of CaMKIIα activity in specific subpopulations of neurons could be a novel approach in therapy of diabetic trigeminal neuropathy. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Maternal high-protein or high-prebiotic-fiber diets affect maternal milk composition and gut microbiota in rat dams and their offspring.

    Science.gov (United States)

    Hallam, Megan C; Barile, Daniela; Meyrand, Mickael; German, J Bruce; Reimer, Raylene A

    2014-11-01

    Maternal gut microbiota and milk composition could modify offspring microbiota and therefore disease susceptibility. The effect of maternal high-protein (HP) or prebiotic diets on maternal milk composition and gut microbiota in rat dams and offspring was examined. Wistar rat dams were fed a control, HP (40% wt/wt), or high-prebiotic-fiber (21.6% wt/wt) (HF) diet throughout pregnancy and lactation. Pups were challenged with a high-fat/sucrose diet from 14.5 to 22.5 weeks of age. Dam milk was analyzed for fat, protein, and oligosaccharides (OS). Fecal microbiota was analyzed in dams at parturition and 2 weeks post-partum and in offspring at 5 and 22 weeks along with cecal digesta at termination. Maternal milk differed only in OS content, each diet group being distinguishable. HF1 and HP1 offspring had decreased plasma lipopolysaccharide compared with C1. Offspring sex, maternal diet, and time (5 weeks vs. 22 weeks of age) affected the microbial groups examined. Bifidobacteria was higher in HF dams and offspring. Increasing protein or fiber content in maternal diet during pregnancy and lactation modifies milk OS content and gut microbiota of dams which may influence establishment of gut microbiota in offspring. © 2014 The Obesity Society.

  8. Mps1 (Monopolar Spindle 1 Protein Inhibition Affects Cellular Growth and Pro-Embryogenic Masses Morphology in Embryogenic Cultures of Araucaria angustifolia (Araucariaceae.

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    Jackellinne C Douétts-Peres

    Full Text Available Somatic embryogenesis has been shown to be an efficient tool for studying processes based on cell growth and development. The fine regulation of the cell cycle is essential for proper embryo formation during the process of somatic embryogenesis. The aims of the present work were to identify and perform a structural and functional characterization of Mps1 and to analyze the effects of the inhibition of this protein on cellular growth and pro-embryogenic mass (PEM morphology in embryogenic cultures of A. angustifolia. A single-copy Mps1 gene named AaMps1 was retrieved from the A. angustifolia transcriptome database, and through a mass spectrometry approach, AaMps1 was identified and quantified in embryogenic cultures. The Mps1 inhibitor SP600125 (10 μM inhibited cellular growth and changed PEMs, and these effects were accompanied by a reduction in AaMps1 protein levels in embryogenic cultures. Our work has identified the Mps1 protein in a gymnosperm species for the first time, and we have shown that inhibiting Mps1 affects cellular growth and PEM differentiation during A. angustifolia somatic embryogenesis. These data will be useful for better understanding cell cycle control during somatic embryogenesis in plants.

  9. Analysis of polyglutamine-coding repeats in the TATA-binding protein in different human populations and in patients with schizophrenia an bipolar affective disorder

    Energy Technology Data Exchange (ETDEWEB)

    Rubinsztein, D.C.; Leggo, J. [Addenbrooke`s National Health Service Trust, Cambridge (United Kingdom); Crow, T.J. [Cambridge Univ. (United Kingdom)] [and others

    1996-09-20

    A new class of disease (including Huntington disease, Kennedy disease, and spinocerebellar ataxias types 1 and 3) results from abnormal expansions of CAG trinucleotides in the coding regions of genes. In all of these diseases the CAG repeats are thought to be translated into polyglutamine tracts. There is accumulating evidence arguing for CAG trinucleotide expansions as one of the causative disease mutations in schizophrenia and bipolar affective disorder. We and others believe that the TATA-binding protein (TBP) is an important candidate to investigate in these diseases as it contains a highly polymorphic stretch of glutamine codons, which are close to the threshold length where the polyglutamine tracts start to be associated with disease. Thus, we examined the lengths of this polyglutamine repeat in normal unrelated East Anglians, South African Blacks, sub-Saharan Africans mainly from Nigeria, and Asian Indians. We also examined 43 bipolar affective disorder patients and 65 schizophrenic patients. The range of polyglutamine tract-lengths that we found in humans was from 26-42 codons. No patients with bipolar affective disorder and schizophrenia had abnormal expansions at this locus. 22 refs., 1 tab.

  10. C4 Protein of Sweet Potato Leaf Curl Virus Regulates Brassinosteroid Signaling Pathway through Interaction with AtBIN2 and Affects Male Fertility in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Huiping Bi

    2017-09-01

    Full Text Available Sweepoviruses have been identified globally and cause substantial yield losses and cultivar decline in sweet potato. This study aimed to investigate the interaction between sweepovirus and plant host by analyzing the function of the viral protein C4 of Sweet potato leaf curl virus-Jiangsu (SPLCV-JS, a sweepovirus cloned from diseased sweet potato plants in East China. Ectopic expression of the C4 in Arabidopsis altered plant development drastically with phenotypic changes including leaf curling, seedling twisting, deformation of floral tissues and reduction of pollen fertility, and seed number. Using bimolecular fluorescence complementation analysis, this study demonstrated that the SPLCV-JS C4 protein interacted with brassinosteroid-insensitive 2 (AtBIN2 in the plasma membrane of Nicotiana benthamiana cells. The C4 AtBIN2 interaction was further confirmed by yeast two-hybrid assays. This interaction led to the re-localization of AtBIN2-interacting proteins AtBES1/AtBZR1 into the nucleus which altered the expression of brassinosteroid (BR-response genes, resulting in the activation of BR-signaling pathway. The interaction of SPLCV-JS C4 and AtBIN2 also led to the down-regulated expression of key genes involved in anther and pollen development, including SPROROCYTELESS/NOZZLE, DEFECTIVE IN TAPEL DEVELOPMENT AND FUNCTION 1, and ABORTED MICROSPORES, which caused abnormal tapetal development, followed by defective exine pattern formation of microspores and pollen release. Consequently, male fertility in the C4 transgenic Arabidopsis was reduced. The present study illustrated how the sweepovirus C4 protein functioned in host cells and affected male fertility by interacting with the key components of BR-signaling pathway.

  11. The protein precursors of peptides that affect the mechanics of connective tissue and/or muscle in the echinoderm Apostichopus japonicus.

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    Maurice R Elphick

    Full Text Available Peptides that cause muscle relaxation or contraction or that modulate electrically-induced muscle contraction have been discovered in the sea cucumber Apostichopus japonicus (Phylum Echinodermata; Class Holothuroidea. By analysing transcriptome sequence data, here the protein precursors of six of these myoactive peptides (the SALMFamides Sticho-MFamide-1 and -2, NGIWYamide, stichopin, GN-19 and GLRFA have been identified, providing novel insights on neuropeptide and endocrine-type signalling systems in echinoderms. The A. japonicus SALMFamide precursor comprises eight putative neuropeptides including both L-type and F-type SALMFamides, which contrasts with previous findings from the sea urchin Strongylocentrotus purpuratus where L-type and F-type SALMFamides are encoded by different genes. The NGIWYamide precursor contains five copies of NGIWYamide but, unlike other NG peptide-type neuropeptide precursors in deuterostomian invertebrates, the NGIWYamide precursor does not have a C-terminal neurophysin domain, indicating loss of this character in holothurians. NGIWYamide was originally discovered as a muscle contractant, but it also causes stiffening of mutable connective tissue in the body wall of A. japonicus, whilst holokinins (PLGYMFR and derivative peptides cause softening of the body wall. However, the mechanisms by which these peptides affect the stiffness of body wall connective tissue are unknown. Interestingly, analysis of the A. japonicus transcriptome reveals that the only protein containing the holokinin sequence PLGYMFR is an alpha-5 type collagen. This suggests that proteolysis of collagen may generate peptides (holokinins that affect body wall stiffness in sea cucumbers, providing a novel perspective on mechanisms of mutable connective tissue in echinoderms.

  12. BolA affects cell growth, and binds to the promoters of penicillin-binding proteins 5 and 6 and regulates their expression.

    Science.gov (United States)

    Guinote, Inês Batista; Matos, Rute Gonçalves; Freire, Patrick; Arraiano, Cecília Maria

    2011-03-01

    The gene bolA was discovered in the 80's, but unraveling its function in the cell has proven to be a complex task. The BolA protein has pleiotropic effects over cell physiology, altering growth and morphology, inducing biofilm formation, and regulating the balance of several membrane proteins. Recently, BolA was shown to be a transcription factor by repressing the expression of the mreB gene. The present report shows that BolA is a transcriptional regulator of the dacA and dacC genes, thus regulating both DD-carboxypeptidases PBP5 and PBP6 and thereby demonstrating the versatility of BolA as a cellular regulator. In this work, we also demonstrate that reduction of cell growth and survival can be connected to the overexpression of the bolA gene in different E. coli backgrounds, particularly in the exponential growth phase. The most interesting finding is that overproduction of BolA affects bacterial growth differently depending on whether the cells were inoculated directly from a plate culture or from an overnight batch culture. This strengthens the idea that BolA can be engaged in the coordination of genes that adapt the cell physiology in order to enhance cell adaptation and survival under stress conditions.

  13. Physico-chemical, functional and rheological properties of milk protein concentrate 60 as affected by disodium phosphate addition, diafiltration and homogenization.

    Science.gov (United States)

    Meena, Ganga Sahay; Singh, Ashish Kumar; Arora, Sumit; Borad, Sanket; Sharma, Rajan; Gupta, Vijay Kumar

    2017-05-01

    Ultrafiltration and diafiltration of skim milk altered delicate salt equilibrium and composition of 5× UF retentate (5× UFR), and thus adversely affected the reconstitutional and functional properties of milk protein concentrate (MPC) powders. It might be due to interaction and aggregation of proteins during spray drying. Therefore, this study was envisaged to investigate the effect of disodium phosphate (DSP) addition, diafiltration and homogenization of retentates on physico-chemical, functional and rheological properties of MPC60 powders. Solubility of fresh control powder was significantly lower than MPC60-H powder; at par with that of MPC60-DSP and MPC60-Na-K, but remained minimum after 60 days of storage at 25 ± 1 °C. The pH (6.6) adjustment of 5× UFR with DSP, significantly enhanced the dispersability, wettability, specific surface area (SSA), heat coagulation time (HCT), emulsification capacity and stability; buffer index of MPC60-DSP powder over control. Diafiltration of 5× UFR with NaCl and KCl, significantly ( P  pseudoplastic behavior, best explained by Hershel Bulkley model. These MPC60 powders with improved functional properties can be used for the improvement of quality attributes of various food formulations.

  14. The C. elegans maternal-effect gene clk-2 is essential for embryonic development, encodes a protein homologous to yeast Tel2p and affects telomere length.

    Science.gov (United States)

    Bénard, C; McCright, B; Zhang, Y; Felkai, S; Lakowski, B; Hekimi, S

    2001-10-01

    The Caenorhabditis elegans maternal-effect clk genes are involved in the temporal control of development and behavior. We report the genetic and molecular characterization of clk-2. A temperature-sensitive mutation in the gene clk-2 affects embryonic and post-embryonic development, reproduction, and rhythmic behaviors. Yet, virtually all phenotypes are fully maternally rescued. Embryonic development strictly requires the activity of maternal clk-2 during a narrow time window between oocyte maturation and the two- to four-cell embryonic stage. Positional cloning of clk-2 reveals that it encodes a protein homologous to S. cerevisiae Tel2p. In yeast, the gene TEL2 regulates telomere length and participates in gene silencing at subtelomeric regions. In C. elegans, clk-2 mutants have elongated telomeres, and clk-2 overexpression can lead to telomere shortening. Tel2p has been reported to bind to telomeric DNA repeats in vitro. However, we find that a functional CLK-2::GFP fusion protein is cytoplasmic in worms. We discuss how the phenotype of clk-2 mutants could be the result of altered patterns of gene expression.

  15. A protein with an unusually short PPR domain, MEF8, affects editing at over 60 Arabidopsis mitochondrial C targets of RNA editing.

    Science.gov (United States)

    Diaz, Michael F; Bentolila, Stephane; Hayes, Michael L; Hanson, Maureen R; Mulligan, R Michael

    2017-11-01

    An RNA-seq approach was used to investigate the role of a PLS-subfamily pentatricopeptide repeat protein, Mitochondrial Editing Factor 8 (MEF8), on editing in Arabidopsis mitochondria and plastids. MEF8 has an intact DYW domain, but possesses an unusually short PLS repeat region of only five repeats. The MEF8 T-DNA insertion (mef8) line exhibited reduced editing at 38 mitochondrial editing sites and increased editing at 24 sites; therefore the absence of MEF8 affects 11% of the mitochondrial editome. Notably, 60% of the matR transcripts' sites showed a decrease of editing extent in the mef8 mutant. An E549A substitution in the MEF8 protein replaced the putatively catalytic glutamate of the HXE motif in the DYW domain. Complementation with MEF8-E549A failed to restore editing at the main target sites but was able to restore editing at the matR transcript; it also decreased the editing extent of most of the C targets exhibiting an increase of editing extent in the mef8 mutant plant. Thus, MEF8 has two antagonistic effects on mitochondrial editing: stimulatory, which requires a catalytic glutamate for most of the targets except for the matR transcript, and inhibitory, for which glutamate is dispensable. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  16. Single nucleotide polymorphisms at 15 codons of the prion protein gene from a scrapie-affected herd of Suffolk sheep in Brazil

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    Caroline Pinto de Andrade

    2011-10-01

    Full Text Available Scrapie is a transmissible spongiform encephalopathy of sheeps and goats, associated with the deposition of a isoform of the prion protein (PrPsc. This isoform presents an altered conformation that leads to aggregation in the host's central nervous and lymphoreticular systems. Predisposition to the prion agent infection can be influenced by specific genotypes related to mutations in amino acids of the PrPsc gene. The most characterized mutations occur at codons 136, 154 and 171, with genotypes VRQ being the most susceptible and ARR the most resistant. In this study we have analyzed polymorphisms in 15 different codons of the PrPsc gene in sheeps from a Suffolk herd from Brazil affected by an outbreak of classical scrapie. Amplicons from the PrPsc gene, encompassing the most relevant altered codons in the protein, were sequenced in order to determine each animal's genotype. We have found polymorphisms at 3 of the 15 analyzed codons (136, 143 and 171. The most variable codon was 171, where all described alleles were identified. A rare polymorphism was found at the 143 codon in 4% of the samples analyzed, which has been described as increasing scrapie resistance in otherwise susceptible animals. No other polymorphisms were detected in the remaining 12 analyzed codons, all of them corresponding to the wild-type prion protein. Regarding the risk degree of developing scrapie, most of the animals (96% had genotypes corresponding to risk groups 1 to 3 (very low to moderate, with only 4% in the higher risks group. Our data is discussed in relation to preventive measures involving genotyping and positive selection to control the disease.

  17. Zebrafish anti-apoptotic gene Bcl-xL can prevent aquatic birnavirus-induced cell death in fish cells without affecting expression of viral proteins.

    Science.gov (United States)

    Huang, Hui-Ling; Liu, Yen-Ting; Chen, Ming-Chyuan; Wu, Jen-Leih; Hong, Jiann-Ruey

    2011-12-01

    The aquatic birnavirus induces mitochondria-mediated cell death in fish; however, the molecular mechanism remains unknown. In the present study, we demonstrated that aquatic birnavirus-induced mitochondria-mediated cell death is regulated by the anti-apoptotic Bcl-2 family member, zfBcl-xL, which is anti-apoptotic and enhances host cell viability. First, CHSE-214 cells carrying EGFP-zfBcl-xL fused genes were selected, established in culture, and used to examine the involvement of zfBcl-xL in host cell protection from the effects of viral infection. EGFP-zfBcl-xL was found to prevent infectious pancreatic necrosis virus (IPNV)-induced phosphatidylserine exposure up to 40% at 12 h and 24 h post-infection (p.i.), block IPNV-induced loss of mitochondrial membrane potential (ΔΨm), and enhance host viability at the middle and late replication stages. In addition, zfBcl-xL overexpression prevented IPNV-induced caspase-9 activation up to 25% and 85% at the middle (12 h p.i.) and late (24 h p.i.) replication stages without affecting expression of viral proteins such as VP3 (as a viral death protein) protein. In the present study, we demonstrated that aquatic birnavirus-induced cell death is prevented by the anti-apoptotic Bcl-2 family member, zfBcl-xL, which enhances host cell viability through blockage of mitochondrial disruption and caspase-9 activation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. Post-weaning selenium and folate supplementation affects gene and protein expression and global DNA methylation in mice fed high-fat diets

    Science.gov (United States)

    2013-01-01

    Background Consumption of high-fat diets has negative impacts on health and well-being, some of which may be epigenetically regulated. Selenium and folate are two compounds which influence epigenetic mechanisms. We investigated the hypothesis that post-weaning supplementation with adequate levels of selenium and folate in offspring of female mice fed a high-fat, low selenium and folate diet during gestation and lactation will lead to epigenetic changes of potential importance for long-term health. Methods Female offspring of mothers fed the experimental diet were either maintained on this diet (HF-low-low), or weaned onto a high-fat diet with sufficient levels of selenium and folate (HF-low-suf), for 8 weeks. Gene and protein expression, DNA methylation, and histone modifications were measured in colon and liver of female offspring. Results Adequate levels of selenium and folate post-weaning affected gene expression in colon and liver of offspring, including decreasing Slc2a4 gene expression. Protein expression was only altered in the liver. There was no effect of adequate levels of selenium and folate on global histone modifications in the liver. Global liver DNA methylation was decreased in mice switched to adequate levels of selenium and folate, but there was no effect on methylation of specific CpG sites within the Slc2a4 gene in liver. Conclusions Post-weaning supplementation with adequate levels of selenium and folate in female offspring of mice fed high-fat diets inadequate in selenium and folate during gestation and lactation can alter global DNA methylation in liver. This may be one factor through which the negative effects of a poor diet during early life can be ameliorated. Further research is required to establish what role epigenetic changes play in mediating observed changes in gene and protein expression, and the relevance of these changes to health. PMID:23497688

  19. DMSA-Coated Iron Oxide Nanoparticles Greatly Affect the Expression of Genes Coding Cysteine-Rich Proteins by Their DMSA Coating.

    Science.gov (United States)

    Zhang, Ling; Wang, Xin; Zou, Jinglu; Liu, Yingxun; Wang, Jinke

    2015-10-19

    The dimercaptosuccinic acid (DMSA) was widely used to coat iron oxide nanoparticles (FeNPs); however, its intracellular cytotoxicity remains to be adequately elucidated. This study analyzed the differentially expressed genes (DEGs) in four mammalian cells treated by a DMSA-coated magnetite FeNP at various doses at different times. The results revealed that about one-fourth of DEGs coded cysteine-rich proteins (CRPs) in all cells under each treatment, indicating that the nanoparticles greatly affected the expressions of CRP-coding genes. Additionally, about 26% of CRP-coding DEGs were enzyme genes in all cells, indicating that the nanoparticles greatly affected the expression of enzyme genes. Further experiments with the nanoparticles and a polyethylenimine (PEI)-coated magnetite FeNP revealed that the effect mainly resulted from DMSA carried into cells by the nanoparticles. This study thus first reported the cytotoxicity of DMSA at the gene transcription level as coating molecules of FeNPs. This study provides new insight into the molecular mechanism by which the DMSA-coated nanoparticles resulted in the transcriptional changes of many CRP-coding genes in cells. This study draws attention toward the intracellular cytotoxicity of DMSA as a coating molecule of nanoparticles, which has very low toxicity as an orally administered antidote due to its extracellular distribution.

  20. Processing of soybean meal and 00-rapeseed meal reduces protein digestibility and pig growth performance but does not affect nitrogen solubilization along the small intestine.

    Science.gov (United States)

    Hulshof, T G; van der Poel, A F B; Hendriks, W H; Bikker, P

    2016-06-01

    An experiment was conducted to determine the effects of processing of soybean meal (SBM) and 00-rapeseed meal (RSM) on N solubilization in chyme, CP digestibility along the small intestine, metabolic load as determined by organ weight, body composition, and growth performance in growing pigs. The SBM and RSM were processed by secondary toasting (at 95°C for 30 min) in the presence of lignosulfonate, resulting in processed SBM (pSBM) and processed RSM (pRSM) as a model for overprocessed protein sources. Fifty-four growing pigs were each fed 1 of the 6 experimental diets. Four of the diets contained SBM, pSBM, RSM, or pRSM as the sole protein source. The remaining 2 experimental diets contained pSBM or pRSM and were supplemented with crystalline AA to the same standardized ileal digestible AA levels as the SBM or RSM diet. Pigs were slaughtered at 40 kg, and organ weights were recorded. The organs plus blood and empty carcass were analyzed for CP content. The small intestine was divided into 3 segments, and chyme samples were taken from the last meter of each segment. Chyme of the SBM, pSBM, RSM, and pRSM diets was centrifuged to separate the soluble and insoluble fractions, and N content was determined in the latter. The amount of insoluble N as a fraction of N in chyme at each small intestinal segment was not affected by processing. Diet type, comprising effects of processing and supplementing crystalline AA, affected ( < 0.05) the G:F and standardized ileal digestibility (SID) of CP. Processing reduced G:F from 0.56 to 0.38 for SBM and 0.49 to 0.40 for RSM, whereas supplementing crystalline AA increased G:F to the level of the SBM and RSM diets. Processing reduced the SID of CP from 87.2% to 69.2% for SBM and 71.0% to 52.2% for RSM. Diet type affected ( < 0.05) the CP content in the empty body, with processing reducing this content from 170 to 144 g/kg empty BW for SBM and 157 to 149 g/kg empty BW for RSM and supplementing crystalline AA restoring this content

  1. Whole-body electromyostimulation and protein supplementation favorably affect sarcopenic obesity in community-dwelling older men at risk: the randomized controlled FranSO study

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    Kemmler W

    2017-09-01

    Full Text Available Wolfgang Kemmler,1 Anja Weissenfels,1 Marc Teschler,1 Sebastian Willert,1 Michael Bebenek,1 Mahdieh Shojaa,1 Matthias Kohl,2 Ellen Freiberger,3 Cornel Sieber,3 Simon von Stengel1 1Institute of Medical Physics, Friedrich-Alexander University of Erlangen-Nürnberg, Erlangen, Germany; 2Faculty of Medical and Life Science, University of Furtwangen, Schwenningen, Germany; 3Institute of Biomedicine of Aging, Friedrich-Alexander University of Erlangen-Nürnberg, Nürnberg, Germany Background: Sarcopenic obesity (SO is a geriatric syndrome characterized by the disproportion between the amount of lean mass and fat mass. Exercise decreases fat and maintains muscle mass; however, older people fail to exercise at doses sufficient to affect musculoskeletal and cardiometabolic risk factors. The aim of this study was to evaluate the effect of whole-body electromyostimulation (WB-EMS, a time-efficient, joint-friendly and highly individualized exercise technology, on sarcopenia and SO in older men. Materials and methods: A total of 100 community-dwelling northern Bavarian men aged ≥70 years with sarcopenia and obesity were randomly (1–1–1 assigned to either 16 weeks of 1 WB-EMS and protein supplementation (WB-EMS&P, 2 isolated protein supplementation or 3 nonintervention control. WB-EMS consisted of 1.5×20 min (85 Hz, 350 µs, 4 s of strain to 4 s of rest applied with moderate-to-high intensity while moving. We further generated a daily protein intake of 1.7–1.8 g/kg/body mass per day. The primary study end point was Sarcopenia Z-Score, and the secondary study end points were body fat rate (%, skeletal muscle mass index (SMI and handgrip strength. Results: Intention-to-treat analysis determined a significantly favorable effect of WB-EMS&P (P<0.001 and protein (P=0.007 vs control. Both groups significantly (P<0.001 lost body fat (WB-EMS&P: 2.1%; protein: 1.1% and differed significantly (P≤0.004 from control (0.3%. Differences between WB

  2. Progressive resistance-loaded voluntary wheel running increases hypertrophy and differentially affects muscle protein synthesis, ribosome biogenesis, and proteolytic markers in rat muscle.

    Science.gov (United States)

    Mobley, C B; Holland, A M; Kephart, W C; Mumford, P W; Lowery, R P; Kavazis, A N; Wilson, J M; Roberts, M D

    2018-02-01

    We examined if 6 weeks of progressive resistance-loaded voluntary wheel running in rats induced plantaris, soleus, and/or gastrocnemius hypertrophy and/or affected markers of translational efficiency, ribosome biogenesis, and markers of proteolysis. For 6 weeks, 8 male Sprague-Dawley rats (~9-10 weeks of age, ~300-325 g) rats were assigned to the progressive resistance-loaded voluntary wheel running model (EX), and ten rats were not trained (SED). For EX rats, the wheel-loading paradigm was as follows - days 1-7: free-wheel resistance, days 8-15: wheel resistance set to 20%-25% body mass, days 16-24: 40% body mass, days 25-32: 60% body mass, days 33-42: 40% body mass. Following the intervention, muscles were analysed for markers of translational efficiency, ribosome biogenesis, and muscle proteolysis. Raw gastrocnemius mass (+13%, p < .01), relative (body mass-corrected) gastrocnemius mass (+16%, p < .001), raw plantaris mass (+13%, p < .05), and relative plantaris mass (+15%, p < .01) were greater in EX vs. SED rats. In spite of gastrocnemius hypertrophy, EX animals presented a 54% decrease in basal muscle protein synthesis levels (p < .01), a 125% increase in pan 4EBP1 levels (p < .001) and a 31% decrease in pan eIF4E levels (p < .05). However, in relation to SED animals, EX animals presented a 70% increase in gastrocnemius c-Myc protein levels (p < .05). Most markers of translational efficiency and ribosome biogenesis were not altered in the plantaris or soleus muscles of EX vs. SED animals. Gastrocnemius F-box protein 32 and poly-ubiquinated protein levels were approximately 150% and 200% greater in SED vs. EX rats (p < .001). These data suggest that the employed resistance training model increases hind limb muscle hypertrophy, and this may be mainly facilitated through reductions in skeletal muscle proteolysis, rather than alterations in ribosome biogenesis or translational efficiency. © 2017 Blackwell Verlag GmbH.

  3. The Varicella-Zoster Virus Immediate-Early 63 protein affects chromatin controlled gene transcription in a cell-type dependent manner

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    Bontems Sébastien

    2007-10-01

    Full Text Available Abstract Background Varicella Zoster Virus Immediate Early 63 protein (IE63 has been shown to be essential for VZV replication, and critical for latency establishment. The activity of the protein as a transcriptional regulator is not fully clear yet. Using transient transfection assays, IE63 has been shown to repress viral and cellular promoters containing typical TATA boxes by interacting with general transcription factors. Results In this paper, IE63 regulation properties on endogenous gene expression were evaluated using an oligonucleotide-based micro-array approach. We found that IE63 modulates the transcription of only a few genes in HeLa cells including genes implicated in transcription or immunity. Furthermore, we showed that this effect is mediated by a modification of RNA POL II binding on the promoters tested and that IE63 phosphorylation was essential for these effects. In MeWo cells, the number of genes whose transcription was modified by IE63 was somewhat higher, including genes implicated in signal transduction, transcription, immunity, and heat-shock signalling. While IE63 did not modify the basal expression of several NF-κB dependent genes such as IL-8, ICAM-1, and IκBα, it modulates transcription of these genes upon TNFα induction. This effect was obviously correlated with the amount of p65 binding to the promoter of these genes and with histone H3 acetylation and HDAC-3 removal. Conclusion While IE63 only affected transcription of a small number of cellular genes, it interfered with the TNF-inducibility of several NF-κB dependent genes by the accelerated resynthesis of the inhibitor IκBα.

  4. Deciphering the Dynamics of Non-Covalent Interactions Affecting Thermal Stability of a Protein: Molecular Dynamics Study on Point Mutant of Thermus thermophilus Isopropylmalate Dehydrogenase.

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    Reetu Sharma

    Full Text Available Thermus thermophilius isopropylmalate dehydrogenase catalyzes oxidative decarboxylation and dehydrogenation of isopropylmalate. Substitution of leucine to alanine at position 172 enhances the thermal stability among the known point mutants. Exploring the dynamic properties of non-covalent interactions such as saltbridges, hydrogen bonds and hydrophobic interactions to explain thermal stability of a protein is interesting in its own right. In this study dynamic changes in the non-covalent interactions are studied to decipher the deterministic features of thermal stability of a protein considering a case study of a point mutant in Thermus thermophilus isopropylmalate dehydrogenase. A total of four molecular dynamic simulations of 0.2 μs were carried out on wild type and mutant's functional dimers at 300 K and 337 K. Higher thermal stability of the mutant as compared to wild type is revealed by root mean square deviation, root mean square fluctuations and Cα-Cα distance with an increase in temperature from 300 K to 337 K. Most of the regions of wild type fluctuate higher than the corresponding regions of mutant with an increase in temperature. Cα-Cα distance analysis suggests that long distance networks are significantly affected in wild type as compared to the mutant. Short lived contacts are higher in wild type, while long lived contacts are lost at 337 K. The mutant forms less hydrogen bonds with water as compared to wild type at 337 K. In contrast to wild type, the mutant shows significant increase in unique saltbridges, hydrogen bonds and hydrophobic contacts at 337 K. The current study indicates that there is a strong inter-dependence of thermal stability on the way in which non-covalent interactions reorganize, and it is rewarding to explore this connection in single mutant studies.

  5. Flowering-Related RING Protein 1 (FRRP1) Regulates Flowering Time and Yield Potential by Affecting Histone H2B Monoubiquitination in Rice (Oryza Sativa).

    Science.gov (United States)

    Du, Yiwei; He, Wei; Deng, Changwang; Chen, Xi; Gou, Lanming; Zhu, Fugui; Guo, Wei; Zhang, Jianfu; Wang, Tao

    2016-01-01

    Flowering time is a critical trait for crops cultivated under various temperature/photoperiod conditions around the world. To understand better the flowering time of rice, we used the vector pTCK303 to produce several lines of RNAi knockdown transgenic rice and investigated their flowering times and other agronomic traits. Among them, the heading date of FRRP1-RNAi knockdown transgenic rice was 23-26 days earlier than that of wild-type plants. FRRP1 is a novel rice gene that encodes a C3HC4-type Really Interesting Novel Gene (RING) finger domain protein. In addition to the early flowering time, FRRP1-RNAi knockdown transgenic rice caused changes on an array of agronomic traits, including plant height, panicle length and grain length. We analyzed the expression of some key genes associated with the flowering time and other agronomic traits in the FRRP1-RNAi knockdown lines and compared with that in wild-type lines. The expression of Hd3a increased significantly, which was the key factor in the early flowering time. Further experiments showed that the level of histone H2B monoubiquitination (H2Bub1) was noticeably reduced in the FRRP1-RNAi knockdown transgenic rice lines compared with wild-type plants and MBP-FRRP1-F1 was capable of self-ubiquitination. The results indicate that Flowering Related RING Protein 1 (FRRP1) is involved in histone H2B monoubiquitination and suggest that FRRP1 functions as an E3 ligase in vivo and in vitro. In conclusion, FRRP1 probably regulates flowering time and yield potential in rice by affecting histone H2B monoubiquitination, which leads to changes in gene expression in multiple processes.

  6. MicroRNA-186 affects the proliferation of tumor cells via yes-associated protein 1 in the occurrence and development of pancreatic cancer.

    Science.gov (United States)

    Niu, Qinghui; Li, Xiaoyu; Xia, Di; Jiang, Yueping; Tian, Zibin; Bian, Cheng; Zhang, Cuiping; Liu, Pei; Zhang, Fengjuan; Yang, Yuling; Wang, Guanglan

    2017-09-01

    The present study aimed to determine the expression of microRNA (miRNA or miR)-186 in tumor tissues and peripheral blood of patients with pancreatic cancer (PC), as well as its mechanism of regulation. A total of 65 patients with PC who underwent surgery between June 2013 and October 2015 were included. In addition, 59 healthy subjects were recruited as controls. Reverse transcription-quantitative polymerase chain reaction was used to measure the expression of mRNA and miRNA. Western blotting and enzyme-linked immunosorbent assay were used to determine protein expression. Bioinformatics was employed for the prediction of the target gene of miR-186, whereas dual luciferase reporter assay was performed to identify whether miR-186 directly bound to YAP1 mRNA. Human pulmonary aortic endothelial cells (HPACs) were transfected with ago-miR-186. YAP1 expression in HPACs was silenced by siRNA. MTT assay was used to evaluate the viability of HPACs. YAP1 mRNA and protein expression levels were elevated in PC. In addition, expression levels of miR-186 in PC were downregulated. miR-186 regulated the expression of YAP1 by binding with the 3'-untranslated region of YAP1. Elevated expression of miR-186 inhibited the proliferation of HPACs by downregulating the expression of YAP1. Decreased expression of YAP1 by siRNA reduced the viability of HPACs. The present study demonstrates that YAP1 is upregulated in the tumor tissues and blood of PC patients, and this may be associated with the downregulation of miR-186. In addition, miR-186 may affect the occurrence and development of PC by controlling the proliferation of PC cells via YAP1.

  7. Can the amount of digestible undegraded protein offered to ewes in late pregnancy affect the periparturient change in resistance to gastrointestinal nematodes?

    Science.gov (United States)

    Sebastiano, Rocco S; Sweeney, Torres; Keady, Timothy W J; Hanrahan, James P; Good, Barbara

    2017-02-15

    Ewes experience a temporary decline in resistance to gastrointestinal nematodes (GIN) during the periparturient period, characterised by a rise in faecal egg count (FEC) that represents a major source of pasture contamination for naïve progeny. The aim of this study was to assess the effect of level of supplementation with digestible undegraded protein (DUP) during the last 6 weeks of pregnancy on periparturient FEC and the performance of ewes with a naturally acquired parasite infection. Eighty-five Belclare and Belclare x Scottish Blackface twin/triplet-bearing ewes were allocated to 1 of 4 dietary groups representing the combination of 2 concentrates (DUP concentration 29 and 94g/kg dry matter) with 2 levels of concentrate during the final 6 weeks of gestation (18 and 30kg in total for ewes with twins; 24 and 35kg for ewes with triplets). All ewes were housed during the pre-partum feeding period and offered grass silage ad libitum; food intake was recorded daily. The intake of DUP varied from 26 to 72g/d among treatments and was reflected in variation of 0.76 to 1.20 in metabolizable protein supply as a proportion of requirements. After lambing, ewes and lambs grazed on permanent sheep pasture, without concentrate supplementation, until weaning (14 weeks post lambing). The variables studied, from week 6 pre-lambing up to week 10 post-lambing, included: FEC, serum pepsinogen concentration, body weight (BW) and body condition score (BCS). The effect of week (relative to lambing date) on FEC was highly significant (P0.05) at any stage either pre- or post-partum. Pepsinogen concentration also varied with time but was not influenced by dietary treatment (P>0.05). The changes in BW and BCS from 6 weeks before lambing to weaning were not affected by the concentration of DUP in the supplement but ewes on treatments involving the higher level of supplementation lost less BW and BCS (Ppregnancy does not affect FEC, BW or BCS of housed ewes with a naturally acquired GIN

  8. Human papillomavirus 16 E2 interacts with neuregulin receptor degradation protein 1 affecting ErbB-3 expression in vitro and in clinical samples of cervical lesions.

    Science.gov (United States)

    Paolini, Francesca; Curzio, Gianfranca; Melucci, Elisa; Terrenato, Irene; Antoniani, Barbara; Carosi, Mariantonia; Mottolese, Marcella; Vici, Patrizia; Mariani, Luciano; Venuti, Aldo

    2016-05-01

    The ErbB tyrosine kinase receptors play a key role in regulating many cellular functions and human papillomaviruses (HPVs) may interact with transductional pathway of different growth factor receptors. Here, these interactions were analysed in W12 cell line carrying HPV 16 genome and in clinical samples. W12 cells, in which HPV16 becomes integrated during passages, were utilised to detect viral and ErbB family expression at early (W12E) and late passages (W12G). Interestingly, a strong reduction of ErbB-3 expression was observed in W12G. Loss of the E2 and E5 viral genes occurs in W12G and this may affect ErbB-3 receptor expression. E2 and E5 rescue experiments demonstrated that only E2 gene was able to restore ErbB-3 expression. E2 is a transcriptional factor but the expression levels of ErbB3 were unaffected and ErbB-3 promoter did not show any consensus sequence for E2, thus E2 may interact in another way with ErbB3. Indeed, HPV 16 E2 can modulate ErbB-3 by interacting with the ubiquitin ligase neuregulin receptor degradation protein 1 (Nrdp-1) that is involved in the regulation of this receptor, via ubiquitination and degradation. E2 co-immunoprecipitated in a complex with Nrdp-1 leading to hypothesise an involvement of this interaction in ErbB-3 regulation. In addition, 90% of the clinical samples with integrated virus and E2 loss showed no or low ErbB-3 positivity, confirming in vitro results. In conclusion, the new discovered interaction of HPV-16 E2 with Nrdp-1 may affect ErbB-3 expression. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. A novel RNA binding protein affects rbcL gene expression and is specific to bundle sheath chloroplasts in C4 plants.

    Science.gov (United States)

    Bowman, Shaun M; Patel, Minesh; Yerramsetty, Pradeep; Mure, Christopher M; Zielinski, Amy M; Bruenn, Jeremy A; Berry, James O

    2013-09-22

    Plants that utilize the highly efficient C4 pathway of photosynthesis typically possess kranz-type leaf anatomy that consists of two morphologically and functionally distinct photosynthetic cell types, the bundle sheath (BS) and mesophyll (M) cells. These two cell types differentially express many genes that are required for C4 capability and function. In mature C4 leaves, the plastidic rbcL gene, encoding the large subunit of the primary CO2 fixation enzyme Rubisco, is expressed specifically within BS cells. Numerous studies have demonstrated that BS-specific rbcL gene expression is regulated predominantly at post-transcriptional levels, through the control of translation and mRNA stability. The identification of regulatory factors associated with C4 patterns of rbcL gene expression has been an elusive goal for many years. RLSB, encoded by the nuclear RLSB gene, is an S1-domain RNA binding protein purified from C4 chloroplasts based on its specific binding to plastid-encoded rbcL mRNA in vitro. Co-localized with LSU to chloroplasts, RLSB is highly conserved across many plant species. Most significantly, RLSB localizes specifically to leaf bundle sheath (BS) cells in C4 plants. Comparative analysis using maize (C4) and Arabidopsis (C3) reveals its tight association with rbcL gene expression in both plants. Reduced RLSB expression (through insertion mutation or RNA silencing, respectively) led to reductions in rbcL mRNA accumulation and LSU production. Additional developmental effects, such as virescent/yellow leaves, were likely associated with decreased photosynthetic function and disruption of associated signaling networks. Reductions in RLSB expression, due to insertion mutation or gene silencing, are strictly correlated with reductions in rbcL gene expression in both maize and Arabidopsis. In both plants, accumulation of rbcL mRNA as well as synthesis of LSU protein were affected. These findings suggest that specific accumulation and binding of the RLSB binding

  10. Meloidogyne incognita Fatty Acid- and Retinol- Binding Protein (Mi-FAR-1 Affects Nematode Infection of Plant Roots and the Attachment of Pasteuria penetrans Endospores

    Directory of Open Access Journals (Sweden)

    Victor Phani

    2017-11-01

    Full Text Available Root-knot nematode (RKN Meloidogyne incognita is an economically important pest of crops. Pasteuria penetrans, is a nematode hyperparasitic bacterium capable of suppressing the reproduction of RKN and thereby useful for its management. Secreted fatty acid and retinol-binding proteins are unique in nematodes and are engaged in nutrient acquisition, development and reproduction; they are also a component of the nematode cuticle and thought to be involved in the interface between hosts and parasites. Attachment of endospores to the cuticle of second stage juveniles of RKN is the primary step of infection and several factors have been identified to facilitate attachment. In this study, the full length of Mi-far-1 (573 bp was cloned from M. incognita and characterized. Analysis revealed that the Mi-far-1 was rich in α-helix structure, contained a predicted consensus casein kinase II phosphorylation site and a glycosylation site. Quantitative PCR showed the highest expression in the fourth stage juveniles and in situ hybridization revealed the presence of Mi-far-1 mRNA in the hypodermis below the cuticle. Single copy insertion pattern of Mi-far-1 in M. incognita genome was detected by Southern blotting. Knockdown of Mi-far-1 showed significantly increased attachment of P. penetrans’ endospores on juvenile cuticle surface and also affected host finding, root infection and nematode fecundity.

  11. Ketogenic diet delays the phase of circadian rhythms and does not affect AMP-activated protein kinase (AMPK) in mouse liver.

    Science.gov (United States)

    Genzer, Yoni; Dadon, Maayan; Burg, Chen; Chapnik, Nava; Froy, Oren

    2015-12-05

    Ketogenic diet (KD) is used for weight loss or to treat epilepsy. KD leads to liver AMP-activated protein kinase (AMPK) activation, which would be expected to inhibit gluconeogenesis. However, KD leads to increased hepatic glucose output. As AMPK and its active phosphorylated form (pAMPK) show circadian oscillation, this discrepancy could stem from wrong-time-of-day sampling. The effect of KD was tested on mouse clock gene expression, AMPK, mTOR, SIRT1 and locomotor activity for 2 months and compared to low-fat diet (LFD). KD led to 1.5-fold increased levels of blood glucose and insulin. Brain pAMPK/AMPK ratio was 40% higher under KD, whereas that in liver was not affected. KD led to 40% and 20% down-regulation of the ratio of pP70S6K/P70S6K, the downstream target of mTOR, in the brain and liver, respectively. SIRT1 levels were 40% higher in the brain, but 40% lower in the liver of KD-fed mice. Clock genes showed delayed rhythms under KD. In the brain of KD-fed mice, amplitudes of clock genes were down-regulated, whereas 6-fold up-regulation was found in the liver. The metabolic state under KD indicates reduced satiety in the brain and reduced anabolism alongside increased gluconeogenesis in the liver. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  12. Tamm-Horsfall Protein Regulates Circulating and Renal Cytokines by Affecting Glomerular Filtration Rate and Acting as a Urinary Cytokine Trap*

    Science.gov (United States)

    Liu, Yan; El-Achkar, Tarek M.; Wu, Xue-Ru

    2012-01-01

    Although few organ systems play a more important role than the kidneys in cytokine catabolism, the mechanism(s) regulating this pivotal physiological function and how its deficiency affects systemic cytokine homeostasis remain unclear. Here we show that elimination of Tamm-Horsfall protein (THP) expression from mouse kidneys caused a marked elevation of circulating IFN-γ, IL1α, TNF-α, IL6, CXCL1, and IL13. Accompanying this were enlarged spleens with prominent white-pulp macrophage infiltration. Lipopolysaccharide (LPS) exacerbated the increase of serum cytokines without a corresponding increase in their urinary excretion in THP knock-out (KO) mice. This, along with the rise of serum cystatin C and the reduced inulin and creatinine clearance from the circulation, suggested that diminished glomerular filtration may contribute to reduced cytokine clearance in THP KO mice both at the baseline and under stress. Unlike wild-type mice where renal and urinary cytokines formed specific in vivo complexes with THP, this “trapping” effect was absent in THP KO mice, thus explaining why cytokine signaling pathways were activated in renal epithelial cells in such mice. Our study provides new evidence implicating an important role of THP in influencing cytokine clearance and acting as a decoy receptor for urinary cytokines. Based on these and other data, we present a unifying model that underscores the role of THP as a major regulator of renal and systemic immunity. PMID:22451664

  13. Octamer-binding protein 4 affects the cell biology and phenotypic transition of lung cancer cells involving β-catenin/E-cadherin complex degradation.

    Science.gov (United States)

    Chen, Zhong-Shu; Ling, Dong-Jin; Zhang, Yang-De; Feng, Jian-Xiong; Zhang, Xue-Yu; Shi, Tian-Sheng

    2015-03-01

    Clinical studies have reported evidence for the involvement of octamer‑binding protein 4 (Oct4) in the tumorigenicity and progression of lung cancer; however, the role of Oct4 in lung cancer cell biology in vitro and its mechanism of action remain to be elucidated. Mortality among lung cancer patients is more frequently due to metastasis rather than their primary tumors. Epithelial‑mesenchymal transition (EMT) is a prominent biological event for the induction of epithelial cancer metastasis. The aim of the present study was to investigate whether Oct4 had the capacity to induce lung cancer cell metastasis via the promoting the EMT in vitro. Moreover, the effect of Oct4 on the β‑catenin/E‑cadherin complex, associated with EMT, was examined using immunofluorescence and immunoprecipitation assays as well as western blot analysis. The results demonstrated that Oct4 enhanced cell invasion and adhesion accompanied by the downregulation of epithelial marker cytokeratin, and upregulation of the mesenchymal markers vimentin and N‑cadherin. Furthermore, Oct4 induced EMT of lung cancer cells by promoting β‑catenin/E‑cadherin complex degradation and regulating nuclear localization of β‑catenin. In conclusion, the present study indicated that Oct4 affected the cell biology of lung cancer cells in vitro through promoting lung cancer cell metastasis via EMT; in addition, the results suggested that the association and degradation of the β‑catenin/E‑cadherin complex was regulated by Oct4 during the process of EMT.

  14. Chronic fatigue syndrome: acute infection and history of physical activity affect resting levels and response to exercise of plasma oxidant/antioxidant status and heat shock proteins.

    Science.gov (United States)

    Jammes, Y; Steinberg, J G; Delliaux, S

    2012-07-01

    A history of high-level physical activity and/or acute infection might constitute stress factors affecting the plasma oxidant-antioxidant status and levels of heat shock proteins (HSPs) in patients with chronic fatigue syndrome (CFS). This case-control study compared data from 43 CFS patients to results from a matched control group of 23 healthy sedentary subjects. Five patients had no relevant previous history (group I). Eighteen had practised high-level sport (group II), and severe acute infection had been diagnosed in nine patients (group III). A combination of sport practice and infection was noted in 11 patients (group IV). After examination at rest, all subjects performed a maximal cycling exercise test. Plasma levels of two markers of oxidative stress [thiobarbituric acid reactive substances (TBARS) and reduced ascorbic acid (RAA)] and both HSP27 and HSP70 were measured. At rest, compared with the control group, the TBARS level was higher in groups II, III and IV patients, and the RAA level was lower in groups III and IV. In addition, HSP70 levels were significantly lower in all CFS groups, compared with controls, but negative correlations were found between resting HSP27 and HSP70 levels and the history of physical activity. After exercise, the peak level of TBARS significantly increased in groups II, III and IV, and the variations in HSP27 and HSP70 were attenuated or suppressed, with the greatest effects in groups III and IV. The presence of stress factors in the history of CFS patients is associated with severe oxidative stress and the suppression of protective HSP27 and HSP70 responses to exercise. © 2011 The Association for the Publication of the Journal of Internal Medicine.

  15. Xenopus LAP2β protein knockdown affects location of lamin B and nucleoporins and has effect on assembly of cell nucleus and cell viability.

    Science.gov (United States)

    Dubińska-Magiera, Magda; Chmielewska, Magdalena; Kozioł, Katarzyna; Machowska, Magdalena; Hutchison, Christopher J; Goldberg, Martin W; Rzepecki, Ryszard

    2016-05-01

    Xenopus LAP2β protein is the single isoform expressed in XTC cells. The protein localizes on heterochromatin clusters both at the nuclear envelope and inside a cell nucleus. The majority of XLAP2β fraction neither colocalizes with TPX2 protein during interphase nor can be immunoprecipitated with XLAP2β antibody. Knockdown of the XLAP2β protein expression in XTC cells by synthetic siRNA and plasmid encoded siRNA resulted in nuclear abnormalities including changes in shape of nuclei, abnormal chromatin structure, loss of nuclear envelope, mislocalization of integral membrane proteins of INM such as lamin B2, mislocalization of nucleoporins, and cell death. Based on timing of cell death, we suggest mechanism associated with nucleus reassembly or with entry into mitosis. This confirms that Xenopus LAP2 protein is essential for the maintenance of cell nucleus integrity and the process of its reassembly after mitosis.

  16. Hippocampal protein expression is differentially affected by chronic paroxetine treatment in adolescent and adult rats: A possible mechanism of paradoxical antidepressant responses in young persons

    Directory of Open Access Journals (Sweden)

    Emily Aspasia Karanges

    2013-07-01

    Full Text Available Selective serotonin reuptake inhibitors (SSRIs are commonly recognised as the pharmacological treatment of choice for patients with depressive disorders, yet their use in adolescent populations has come under scrutiny following reports of minimal efficacy and an increased risk of suicidal ideation and behavior in this age group. The biological mechanisms underlying these effects are largely unknown. Accordingly, the current study examined changes in hippocampal protein expression following chronic administration of paroxetine in drinking water (target dose = 10 mg/kg for 22 days to adult and adolescent rats. Results indicated age-specific changes in protein expression, with paroxetine significantly altering expression of 8 proteins in adolescents only and 10 proteins solely in adults. A further 12 proteins were significantly altered in both adolescents and adults. In adults, protein changes were generally suggestive of a neurotrophic and neuroprotective effect of paroxetine, with significant downregulation of apoptotic proteins Galectin 7 and Cathepsin B, and upregulation of the neurotrophic factor Neurogenin 1 and the antioxidant proteins Aldose reductase and Carbonyl reductase 3. Phosphodiesterase 10A, a signalling protein associated with major depressive disorder, was also downregulated (−6.5 fold in adult rats. Adolescent rats failed to show the neurotrophic and neuroprotective effects observed in adults, instead displaying upregulation of the proapoptotic protein BH3-interacting domain death agonist (4.3 fold. Adolescent protein expression profiles also suggested impaired phosphoinositide signalling (Protein kinase C: −3.1 fold and altered neurotransmitter transport and release (Syntaxin 7: 5.7 fold; Dynamin 1: −6.9 fold. The results of the present study provide clues as to possible mechanisms underlying the atypical response of human adolescents to paroxetine treatment.

  17. Problems in early diagnosis of bladder cancer in a spinal cord injury patient: Report of a case of simultaneous production of granulocyte colony stimulating factor and parathyroid hormone-related protein by squamous cell carcinoma of urinary bladder

    Directory of Open Access Journals (Sweden)

    Singh Gurpreet

    2002-08-01

    Full Text Available Abstract Background Typical symptoms and signs of a clinical condition may be absent in spinal cord injury (SCI patients. Case presentation A male with paraplegia was passing urine through penile sheath for 35 years, when he developed urinary infections. There was no history of haematuria. Intravenous urography showed bilateral hydronephrosis. The significance of abnormal outline of bladder was not appreciated. As there was large residual urine, he was advised intermittent catheterisation. Serum urea: 3.5 mmol/L; creatinine: 77 umol/L. A year later, serum urea: 36.8 mmol/l; creatinine: 632 umol/l; white cell count: 22.2; neutrophils: 18.88. Ultrasound: bilateral hydronephrosis. Bilateral nephrostomy was performed. Subsequently, blood tests showed: Urea: 14.2 mmol/l; Creatinine: 251 umol/l; Adjusted Calcium: 3.28 mmol/l; Parathyroid hormone: A repeat ultrasound scan demonstrated a tumour arising from right lateral wall; biopsy revealed squamous cell carcinoma. In view of persistently high white cell count and high calcium level, immunohistochemistry for G-CSF and PTHrP was performed. Dense staining of tumour cells for G-CSF and faintly positive staining for C-terminal PTHrP were observed. This patient expired about five months later. Conclusion This case demonstrates how delay in diagnosis of bladder cancer could occur in a SCI patient due to absence of characteristic symptoms and signs.

  18. Changes in the effective gravitational field strength affect the state of phosphorylation of stress-related proteins in callus cultures of Arabidopsis thaliana

    Science.gov (United States)

    Barjaktarović, Žarko; Schütz, Wolfgang; Madlung, Johannes; Fladerer, Claudia; Nordheim, Alfred; Hampp, Rüdiger

    2009-01-01

    In a recent study it was shown that callus cell cultures of Arabidopsis thaliana respond to changes in gravitational field strengths by changes in protein expression. Using ESI-MS/MS for proteins with differential abundance after separation by 2D-PAGE, 28 spots which changed reproducibly and significantly in amount (P gravitational fields induce the production of ROS. Our data further indicate that responses toward RP are more by post-translational protein modulation (most changes in the degree of phosphorylation occur under RP-treatment) than by protein expression (hypergravity). PMID:19129159

  19. Dietary protein sources differentially affect microbiota, mTOR activity and transcription of mTOR signaling pathways in the small intestine

    NARCIS (Netherlands)

    Kar, Soumya K.; Jansman, Alfons J.M.; Benis, Nirupama; Ramiro-Garcia, Javier; Schokker, Dirkjan; Kruijt, Leo; Stolte, Ellen H.; Taverne-Thiele, Johanna J.; Smits, Mari A.; Wells, Jerry M.

    2017-01-01

    Dietary protein sources can have profound effects on host-microbe interactions in the gut that are critically important for immune resilience. However more knowledge is needed to assess the impact of different protein sources on gut and animal health. Thirty-six wildtype male C57BL/6J mice of 35 d

  20. Dietary fish protein hydrolysates containing bioactive motifs affect serum and adipose tissue fatty acid compositions, serum lipids, postprandial glucose regulation and growth in obese Zucker fa/fa rats.

    Science.gov (United States)

    Drotningsvik, Aslaug; Mjøs, Svein A; Pampanin, Daniela M; Slizyte, Rasa; Carvajal, Ana; Remman, Tore; Høgøy, Ingmar; Gudbrandsen, Oddrun A

    2016-10-01

    The world's fisheries and aquaculture industries produce vast amounts of protein-containing by-products that can be enzymatically hydrolysed to smaller peptides and possibly be used as additives to functional foods and nutraceuticals targeted for patients with obesity-related metabolic disorders. To investigate the effects of fish protein hydrolysates on markers of metabolic disorders, obese Zucker fa/fa rats consumed diets with 75 % of protein from casein/whey (CAS) and 25 % from herring (HER) or salmon (SAL) protein hydrolysate from rest raw material, or 100 % protein from CAS for 4 weeks. The fatty acid compositions were similar in the experimental diets, and none of them contained any long-chain n-3 PUFA. Ratios of lysine:arginine and methionine:glycine were lower in HER and SAL diets when compared with CAS, and taurine was detected only in fish protein hydrolysate diets. Motifs with reported hypocholesterolemic or antidiabetic activities were identified in both fish protein hydrolysates. Rats fed HER diet had lower serum HDL-cholesterol and LDL-cholesterol, and higher serum TAG, MUFA and n-3:n-6 PUFA ratio compared with CAS-fed rats. SAL rats gained more weight and had better postprandial glucose regulation compared with CAS rats. Serum lipids and fatty acids were only marginally affected by SAL, but adipose tissue contained less total SFA and more total n-3 PUFA when compared with CAS. To conclude, diets containing hydrolysed rest raw material from herring or salmon proteins may affect growth, lipid metabolism, postprandial glucose regulation and fatty acid composition in serum and adipose tissue in obese Zucker rats.

  1. Presence or absence of carbohydrates and the proportion of fat in a high-protein diet affect appetite suppression but not energy expenditure in normal-weight human subjects fed in energy balance.

    Science.gov (United States)

    Veldhorst, Margriet A B; Westerterp, Klaas R; van Vught, Anneke J A H; Westerterp-Plantenga, Margriet S

    2010-11-01

    Two types of relatively high-protein diets, with a normal or low proportion of carbohydrates, have been shown effective for weight loss. The objective was to assess the significance of the presence or absence of carbohydrates and the proportion of fat in high-protein diets for affecting appetite suppression, energy expenditure, and fat oxidation in normal-weight subjects in energy balance. Subjects (aged 23 (sd 3) years and BMI 22·0 (sd 1·9) kg/m2) were stratified in two groups. Each was offered two diets in a randomised cross-over design: group 1 (n 22) - normal protein (NP; 10, 60 and 30 % energy (En%) from protein, carbohydrate and fat), high protein (HP; 30, 40 and 30 En%); group 2 (n 23) - normal protein (NP-g; 10, 60 and 30 En%), high protein, carbohydrate-free (HP-0C; 30, 0 and 70 En%) for 2 d; NP-g and HP-0C were preceded by glycogen-lowering exercise (day 1). Appetite was measured throughout day 2 using visual analogue scales (VAS). Energy expenditure (EE) and substrate oxidation (respiratory quotient; RQ) were measured in a respiration chamber (08.00 hours on day 2 until 07.30 hours on day 3). Fasting plasma β-hydroxybutyrate (BHB) concentration was measured (day 3). NP-g and NP did not differ in hunger, EE, RQ and BHB. HP-0C and HP v. NP-g and NP, respectively, were lower in hunger (P carbohydrates exchanged for fat. Energy expenditure was not affected by the carbohydrate content of a high-protein diet.

  2. Application potential of ATR-FT/IR molecular spectroscopy in animal nutrition: revelation of protein molecular structures of canola meal and presscake, as affected by heat-processing methods, in relationship with their protein digestive behavior and utilization for dairy cattle.

    Science.gov (United States)

    Theodoridou, Katerina; Yu, Peiqiang

    2013-06-12

    Protein quality relies not only on total protein but also on protein inherent structures. The most commonly occurring protein secondary structures (α-helix and β-sheet) may influence protein quality, nutrient utilization, and digestive behavior. The objectives of this study were to reveal the protein molecular structures of canola meal (yellow and brown) and presscake as affected by the heat-processing methods and to investigate the relationship between structure changes and protein rumen degradations kinetics, estimated protein intestinal digestibility, degraded protein balance, and metabolizable protein. Heat-processing conditions resulted in a higher value for α-helix and β-sheet for brown canola presscake compared to brown canola meal. The multivariate molecular spectral analyses (PCA, CLA) showed that there were significant molecular structural differences in the protein amide I and II fingerprint region (ca. 1700-1480 cm(-1)) between the brown canola meal and presscake. The in situ degradation parameters, amide I and II, and α-helix to β-sheet ratio (R_a_β) were positively correlated with the degradable fraction and the degradation rate. Modeling results showed that α-helix was positively correlated with the truly absorbed rumen synthesized microbial protein in the small intestine when using both the Dutch DVE/OEB system and the NRC-2001 model. Concerning the protein profiles, R_a_β was a better predictor for crude protein (79%) and for neutral detergent insoluble crude protein (68%). In conclusion, ATR-FT/IR molecular spectroscopy may be used to rapidly characterize feed structures at the molecular level and also as a potential predictor of feed functionality, digestive behavior, and nutrient utilization of canola feed.

  3. Estimation of genetic parameters and detection of chromosomal regions affecting the major milk proteins and their post translational modifications in Danish Holstein and Danish Jersey cattle.

    Science.gov (United States)

    Buitenhuis, Bart; Poulsen, Nina A; Gebreyesus, Grum; Larsen, Lotte B

    2016-08-02

    In the Western world bovine milk products are an important protein source in human diet. The major proteins in bovine milk are the four caseins (CN), αS1-, αS2-, β-, and k-CN and the two whey proteins, β-LG and α-LA. It has been shown that both the amount of specific CN and their isoforms including post-translational modifications (PTM) influence technological properties of milk. Therefore, the aim of this study was to 1) estimate genetic parameters for individual proteins in Danish Holstein (DH) (n = 371) and Danish Jersey (DJ) (n = 321) milk, and 2) detect genomic regions associated with specific milk protein and their different PTM forms using a genome-wide association study (GWAS) approach. For DH, high heritability estimates were found for protein percentage (0.47), casein percentage (0.43), k-CN (0.77), β-LG (0.58), and α-LA (0.40). For DJ, high heritability estimates were found for protein percentage (0.70), casein percentage (0.52), and α-LA (0.44). The heritability for G-k-CN, U-k-CN and GD was higher in the DH compared to the DJ, whereas the heritability for the PD of αS1-CN was lower in DH compared to DJ, whereas the PD for αS2-CN was higher in DH compared to DJ. The GWAS results for the main milk proteins were in line what has been earlier published. However, we showed that there were SNPs specifically regulating G-k-CN in DH. Some of these SNPs were assigned to casein protein kinase genes (CSNK1G3, PRKCQ). The genetic analysis of the major milk proteins and their PTM forms revealed that these were heritable in both DH and DJ. In DH, genomic regions specific for glycosylation of k-CN were detected. Furthermore, genomic regions for the major milk proteins confirmed the regions on BTA6 (casein cluster), BTA11 (PEAP), and BTA14 (DGAT1) as important regions influencing protein composition in milk. The results from this study provide confidence that it is possible to breed for specific milk protein including the different PTM forms.

  4. Deciphering the Dynamics of Non-Covalent Interactions Affecting Thermal Stability of a Protein: Molecular Dynamics Study on Point Mutant of Thermus thermophilus Isopropylmalate Dehydrogenase

    National Research Council Canada - National Science Library

    Sharma, Reetu; Sastry, G Narahari

    2015-01-01

    .... Exploring the dynamic properties of non-covalent interactions such as saltbridges, hydrogen bonds and hydrophobic interactions to explain thermal stability of a protein is interesting in its own right...

  5. A mutation in the Arabidopsis HYL1 gene encoding a dsRNA binding protein affects responses to abscisic acid, auxin, and cytokinin

    Science.gov (United States)

    Lu, C.; Fedoroff, N.

    2000-01-01

    Both physiological and genetic evidence indicate interconnections among plant responses to different hormones. We describe a pleiotropic recessive Arabidopsis transposon insertion mutation, designated hyponastic leaves (hyl1), that alters the plant's responses to several hormones. The mutant is characterized by shorter stature, delayed flowering, leaf hyponasty, reduced fertility, decreased rate of root growth, and an altered root gravitropic response. It also exhibits less sensitivity to auxin and cytokinin and hypersensitivity to abscisic acid (ABA). The auxin transport inhibitor 2,3,5-triiodobenzoic acid normalizes the mutant phenotype somewhat, whereas another auxin transport inhibitor, N-(1-naph-thyl)phthalamic acid, exacerbates the phenotype. The gene, designated HYL1, encodes a 419-amino acid protein that contains two double-stranded RNA (dsRNA) binding motifs, a nuclear localization motif, and a C-terminal repeat structure suggestive of a protein-protein interaction domain. We present evidence that the HYL1 gene is ABA-regulated and encodes a nuclear dsRNA binding protein. We hypothesize that the HYL1 protein is a regulatory protein functioning at the transcriptional or post-transcriptional level.

  6. Fetal jaw movement affects Ihh signaling in mandibular condylar cartilage development: the possible role of Ihh as mechanotransduction mediator.

    Science.gov (United States)

    Jahan, Esrat; Matsumoto, Akihiro; Rafiq, Ashiq Mahmood; Hashimoto, Ryuju; Inoue, Takayuki; Udagawa, Jun; Sekine, Joji; Otani, Hiroki

    2014-10-01

    Jaw movement is an important mechanical factor for prenatal development of the condylar cartilage of mandible. Fetal jaw movement restriction has been shown to cause deformity of the mandibular condyle. We hypothesized that this treatment affects the expression of mechanosensitive molecules, namely Indian hedgehog (Ihh) and Parathyroid hormone related protein (PTHrP) in the condyle. We restrained jaw movement by suturing the jaw of E15.5 mouse embryos and allowed them to develop until E18.5 using exo utero system, and analyzed them by immunohistochemistry and in situ hybridization methods. Morphological, histomorphometric and immunohistochemical study showed that the mandibular condylar cartilage was reduced and deformed, the volume and total cell numbers in the condylar cartilage were also reduced, and number and/or distribution of 5-bromo-2'-deoxyuridine-positive cells, Ihh-positive cells in the mesenchymal and pre-hypertrophic zones were significantly and correspondingly decreased in the sutured group. Using in situ hybridization, reduced expression of Ihh, PTHrP and their related receptors were observed in condylar cartilage of the sutured embryos. Our results revealed that the altered mechanical stress induced by prenatal jaw movement restriction decreased proliferating cells, the amount of cartilage, and altered expression of the Ihh and PTHrP, suggesting that Ihh act as mechanotransduction mediators in the development of mandibular condylar cartilage. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Structure and barrier properties of human embryonic stem cell-derived retinal pigment epithelial cells are affected by extracellular matrix protein coating.

    Science.gov (United States)

    Sorkio, Anni; Hongisto, Heidi; Kaarniranta, Kai; Uusitalo, Hannu; Juuti-Uusitalo, Kati; Skottman, Heli

    2014-02-01

    Extracellular matrix (ECM) interactions play a vital role in cell morphology, migration, proliferation, and differentiation of cells. We investigated the role of ECM proteins on the structure and function of human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cells during their differentiation and maturation from hESCs into RPE cells in adherent differentiation cultures on several human ECM proteins found in native human Bruch's membrane, namely, collagen I, collagen IV, laminin, fibronectin, and vitronectin, as well as on commercial substrates of xeno-free CELLstart™ and Matrigel™. Cell pigmentation, expression of RPE-specific proteins, fine structure, as well as the production of basal lamina by hESC-RPE on different protein coatings were evaluated after 140 days of differentiation. The integrity of hESC-RPE epithelium and barrier properties on different coatings were investigated by measuring transepithelial resistance. All coatings supported the differentiation of hESC-RPE cells as demonstrated by early onset of cell pigmentation and further maturation to RPE monolayers after enrichment. Mature RPE phenotype was verified by RPE-specific gene and protein expression, correct epithelial polarization, and phagocytic activity. Significant differences were found in the degree of RPE cell pigmentation and tightness of epithelial barrier between different coatings. Further, the thickness of self-assembled basal lamina and secretion of the key ECM proteins found in the basement membrane of the native RPE varied between hESC-RPE cultured on compared protein coatings. In conclusion, this study shows that the cell culture substrate has a major effect on the structure and basal lamina production during the differentiation and maturation of hESC-RPE potentially influencing the success of cell integrations and survival after cell transplantation.

  8. Heterologous minor coat proteins of Citrus tristeza virus strains affect encapsidation, but the coexpression of HSP70h and p61 restores encapsidation to wild-type levels.

    Science.gov (United States)

    Tatineni, Satyanarayana; Gowda, Siddarame; Dawson, William O

    2010-07-05

    The long flexuous bipolar virions of Citrus tristeza virus (CTV), a Closterovirus, are encapsidated with two capsid proteins at opposite ends: the minor coat protein (CPm) encapsidates the 5' 630 nts of the genomic RNA and the major coat protein encapsidates the remainder of the genome. In this study, we found encapsidation of CTV CPm in the absence of other assembly-related proteins is highly specific in contrast to most plant viruses that allow virion assembly by a range of heterologous coat proteins. Heterologous CPms with 95-96% amino acid identity from related strains in CTV-CPm, a replicon with CPm as the only assembly-related ORF, either failed to initiate encapsidation or reduced encapsidation substantially. Substitution of subsets of amino acids revealed that the amino acids that differ between positions 121 and 180 of the VT strain, and 61 and 120 of the T3 strain were involved in specific encapsidation. We further mapped the specific encapsidation to a single amino acid: mutation of methionine(165) to threonine (VT type) or serine(105) to proline (T3 type) in CTV-CPm failed to form nucleocapsids. However, the heterologous CPm in combination with both HSP70h and p61 proteins, but not HSP70h or p61 alone, encapsidated at wild-type levels, suggesting that specific encapsidation by CPm was mitigated by the combination of HSP70h and p61. Thus, in addition to the previously described functions of HSP70h and p61 of greatly enhanced virion formation and restriction of CPm encapsidation to the 5' 630 nts of the genomic RNA, these proteins facilitate encapsidation by heterologous CPms. Published by Elsevier Inc.

  9. Stability of α-tocopherol in freeze-dried sugar-protein-oil emulsion solids as affected by water plasticization and sugar crystallization.

    Science.gov (United States)

    Zhou, Yankun; Roos, Yrjö H

    2012-08-01

    Water plasticization of sugar-protein encapsulants may cause structural changes and decrease the stability of encapsulated compounds during storage. The retention of α-tocopherol in freeze-dried lactose-milk protein-oil, lactose-soy protein-oil, trehalose-milk protein-oil, and trehalose-soy protein-oil systems at various water activities (a(w)) and in the presence of sugar crystallization was studied. Water sorption was determined gravimetrically. Glass transition and sugar crystallization were studied using differential scanning calorimetry and the retention of α-tocopherol spectrophotometrically. The loss of α-tocopherol followed lipid oxidation, but the greatest stability was found at 0 a(w) presumably because of α-tocopherol immobilization at interfaces and consequent reduction in antioxidant activity. A considerable loss of α-tocopherol coincided with sugar crystallization. The results showed that glassy matrices may protect encapsulated α-tocopherol; however, its role as an antioxidant at increasing aw accelerated its loss. Sugar crystallization excluded the oil-containing α-tocopherol from the protecting matrices and exposed it to surroundings, which decreased the stability of α-tocopherol.

  10. Monitoring the progression of calcium and protein solubilisation as affected by calcium chelators during small-scale manufacture of casein-based food matrices.

    Science.gov (United States)

    McIntyre, Irene; O'Sullivan, Michael; O'Riordan, Dolores

    2017-12-15

    Calcium and protein solubilisation during small-scale manufacture of semi-solid casein-based food matrices was investigated and found to be very different in the presence or absence of calcium chelating salts. Calcium concentrations in the dispersed phase increased and calcium-ion activity (ACa++) decreased during manufacture of the matrices containing calcium chelating salts; with ∼23% of total calcium solubilised by the end of manufacture. In the absence of calcium chelating salts, these concentrations were significantly lower at equivalent processing times and remained unchanged as did ACa++, throughout manufacture. The protein content of the dispersed phase was low (≤3% of total protein), but was significantly higher for matrices containing calcium chelating salts. This study elucidates the critical role of calcium chelating salts in modulating casein hydration and dispersion and gives an indication of the levels of soluble calcium and protein required to allow matrix formation during manufacture of casein-based food structures e.g. processed and analogue cheese. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Effect of periodontal treatment on serum C-reactive protein level in obese and normal-weight women affected with chronic periodontitis

    NARCIS (Netherlands)

    Al-Zahrani, M.S.; Alghamdi, H.S.

    2012-01-01

    OBJECTIVE: To investigate the effect of conventional periodontal therapy on serum C-reactive protein (CRP) level and periodontal status in obese and normal-weight chronic periodontitis patients. METHODS: This is a controlled clinical trial conducted at the King Abdulaziz University Faculty of

  12. The evolutionarily conserved protein PHOTOSYNTHESIS AFFECTED MUTANT71 is required for efficient manganese uptake at the thylakoid membrane in Arabidopsis

    DEFF Research Database (Denmark)

    Schneider, Anja; Steinberger, Iris; Herdean, Andrei

    2016-01-01

    In plants, algae, and cyanobacteria, photosystem II (PSII) catalyzes the light-driven oxidation of water. The oxygen-evolving complex of PSII is a Mn4CaO5 cluster embedded in a well-defined protein environment in the thylakoid membrane. However, transport of manganese and calcium into the thylakoid...

  13. A comparative analysis on the physicochemical properties of tick-borne encephalitis virus envelope protein residues that affect its antigenic properties

    Czech Academy of Sciences Publication Activity Database

    Bukin, Y. S.; Dzhioev, Y.; Tkachev, S. E.; Kozlova, I.; Paramonov, A. I.; Růžek, Daniel; Qu, Z.; Zlobin, V. I.

    2017-01-01

    Roč. 238, JUN 15 (2017), s. 124-132 ISSN 0168-1702 Institutional support: RVO:60077344 Keywords : tick-borne encephalitis virus * E protein * physicochemical properties amino acid residue * antigen * antibody Subject RIV: EE - Microbiology, Virology Impact factor: 2.628, year: 2016

  14. Lack of Prion Accumulation in Lymphoid Tissues of Scrapie-affected Sheep with the AA136, QR171 Prion Protein Genotype

    Science.gov (United States)

    Background: Sheep scrapie is a transmissible spongiform encephalopathy which can be transmitted horizontally through the shedding of an infectious conformer (PrP**Sc) of the normal cellular prion protein (PrP**c). Genetics profoundly influence the susceptibility of sheep to scrapie. PrP**c amino-aci...

  15. Mechanical and water barrier properties of isolated soy protein composite edible films as affected by carvacrol and cinnamaldehyde micro and nanoemulsions

    Science.gov (United States)

    Edible films may be used in food packaging, for which they must deliver good barrier and mechanical properties. Films based on proteins have good gas barrier and mechanical properties, but poor water barrier properties. Films made from lipids have good water barrier properties, but poor mechanical p...

  16. Environmental conditions affect semolina quality in durum wheat (Triticum turgidum ssp. durum L.) cultivars with different gluten strength and gluten protein composition.

    Science.gov (United States)

    Fois, Simonetta; Schlichting, Linda; Marchylo, Brian; Dexter, James; Motzo, Rosella; Giunta, Francesco

    2011-11-01

    Sowing time may impact semolina and pasta cooking quality by changing the environmental conditions during grain filling. The effect of an optimum and a delayed sowing time on semolina quality was studied by comparing six cultivars under irrigation, in order to isolate temperature from drought effects. Protein content was higher in the old cultivars and in the late sowings, according to the number of days with temperature between 30 and 40 °C during ripening. Gluten index increased as temperature rose to a threshold of about 30 °C, then decreased under higher temperatures. Mixograph parameters were less sensitive to high temperatures. Gliadin:glutenin correlated with gluten strength. Spaghetti firmness and protein content were positively correlated independently of sowing date. Cultivars Trinakria and Cappelli had the highest spaghetti firmness (900 and 828 g). Late sowings may represent a way of increasing pasta cooking quality whenever they place grain filling under thermal conditions able to increase protein percentage, although the accompanying decrease in yield may represent a drawback in environments prone to drought stress during ripening. The lower protein percentages of modern durum wheat cultivars under conventional sowing times results in a lower pasta cooking quality despite higher gluten strength. Published 2011 by John Wiley & Sons, Ltd.

  17. Chronic Maternal Low-Protein Diet in Mice Affects Anxiety, Night-Time Energy Expenditure and Sleep Patterns, but Not Circadian Rhythm in Male Offspring.

    Directory of Open Access Journals (Sweden)

    Randy F Crossland

    Full Text Available Offspring of murine dams chronically fed a protein-restricted diet have an increased risk for metabolic and neurobehavioral disorders. Previously we showed that adult offspring, developmentally exposed to a chronic maternal low-protein (MLP diet, had lower body and hind-leg muscle weights and decreased liver enzyme serum levels. We conducted energy expenditure, neurobehavioral and circadian rhythm assays in male offspring to examine mechanisms for the body-weight phenotype and assess neurodevelopmental implications of MLP exposure. C57BL/6J dams were fed a protein restricted (8%protein, MLP or a control protein (20% protein, C diet from four weeks before mating until weaning of offspring. Male offspring were weaned to standard rodent diet (20% protein and single-housed until 8-12 weeks of age. We examined body composition, food intake, energy expenditure, spontaneous rearing activity and sleep patterns and performed behavioral assays for anxiety (open field activity, elevated plus maze [EPM], light/dark exploration, depression (tail suspension and forced swim test, sociability (three-chamber, repetitive (marble burying, learning and memory (fear conditioning, and circadian behavior (wheel-running activity during light-dark and constant dark cycles. We also measured circadian gene expression in hypothalamus and liver at different Zeitgeber times (ZT. Male offspring from separate MLP exposed dams had significantly greater body fat (P = 0.03, less energy expenditure (P = 0.004, less rearing activity (P = 0.04 and a greater number of night-time rest/sleep bouts (P = 0.03 compared to control. MLP offspring displayed greater anxiety-like behavior in the EPM (P<0.01 but had no learning and memory deficit in fear-conditioning assay (P = 0.02. There was an effect of time on Per1, Per 2 and Clock circadian gene expression in the hypothalamus but not on circadian behavior. Thus, transplacental and early developmental exposure of dams to chronic MLP reduces

  18. Testosterone increases the muscle protein synthesis rate but does not affect very-low-density lipoprotein metabolism in obese premenopausal women.

    Science.gov (United States)

    Wang, Xuewen; Smith, Gordon I; Patterson, Bruce W; Reeds, Dominic N; Kampelman, Janine; Magkos, Faidon; Mittendorfer, Bettina

    2012-03-15

    Men and women with hyperandrogenemia have a more proatherogenic plasma lipid profile [e.g., greater triglyceride (TG) and total and low-density lipoprotein-cholesterol and lower high-density lipoprotein-cholesterol concentrations] than healthy premenopausal women. Furthermore, castration of male rats markedly reduces testosterone availability below normal and decreases plasma TG concentration, and testosterone replacement reverses this effect. Testosterone is, therefore, thought to be an important regulator of plasma lipid homeostasis. However, little is known about the effect of testosterone on plasma TG concentration and kinetics. Furthermore, testosterone is a potent skeletal muscle protein anabolic agent in men, but its effect on muscle protein turnover in women is unknown. We measured plasma lipid concentrations, hepatic very low density lipoprotein (VLDL)-TG and VLDL-apolipoprotein B-100 secretion rates, and the muscle protein fractional synthesis rate in 10 obese women before and after trandermal testosterone (1.25 g of 1% AndroGel daily) treatment for 3 wk. Serum total and free testosterone concentrations increased (P testosterone treatment, reaching concentrations that are comparable to those in women with hyperandrogenemia, but lower than the normal range for eugonadal men. Except for a small (∼10%) decrease in plasma high-density lipoprotein particle and cholesterol concentrations (P testosterone therapy had no effect on plasma lipid concentrations, lipoprotein particle sizes, and hepatic VLDL-TG and VLDL-apolipoprotein B-100 secretion rates (all P > 0.05); the muscle protein fractional synthesis rate, however, increased by ∼45% (P testosterone is a potent skeletal muscle protein anabolic agent, but not an important regulator of plasma lipid homeostasis in obese women.

  19. Insights into the functional characteristics of geminivirus rolling-circle replication initiator protein and its interaction with host factors affecting viral DNA replication.

    Science.gov (United States)

    Rizvi, Irum; Choudhury, Nirupam Roy; Tuteja, Narendra

    2015-02-01

    Geminiviruses are DNA viruses that infect several economically important crops, resulting in a reduction in their overall yield. These plant viruses have circular, single-stranded DNA genomes that replicate mainly by a rolling-circle mechanism. Geminivirus infection results in crosstalk between viral and cellular factors to complete the viral life cycle or counteract the infection as part of defense mechanisms of host plants. The geminiviral replication initiator protein Rep is the only essential viral factor required for replication. It is multifunctional and is known to interact with a number of host factors to modulate the cellular environment or to function as a part of the replication machinery. This review provides a holistic view of the research related to the viral Rep protein and various host factors involved in geminiviral DNA replication. Studies on the promiscuous nature of geminiviral satellite DNAs are also reviewed.

  20. Homogenization conditions affect the oxidative stability of fish oil enriched milk emulsions: oxidation linked to changes in protein composition at the oil-water interface

    DEFF Research Database (Denmark)

    Sørensen, Ann-Dorit M.; Baron, Caroline P.; Let, Mette B.

    2007-01-01

    Fish oil was incorporated into milk under different homogenization temperatures (50 and 72 °C) and pressures (5, 15, and 22.5 MPa). Subsequently, the oxidative stability of the milk and changes in the protein composition of the milk fat globule membrane (MFGM) were examined. Results showed...... casein seemed to be present at the oil−water interface with increasing pressure. Overall, the results indicated that a combination of more β-lactoglobulin and less casein at the oil−water interface gave the most stable emulsions with respect to lipid oxidation....... that high pressure and high temperature (72 °C and 22.5 MPa) resulted in less lipid oxidation, whereas low pressure and low temperature (50 °C and 5 MPa) resulted in faster lipid oxidation. Analysis of protein oxidation indicated that especially casein was prone to oxidation. The level of free thiol groups...

  1. Dataset of liver proteins of eu- and hypothyroid rats affected in abundance by any of three factors: in vivo exposure to hexabromocyclododecane (HBCD, thyroid status, gender differences

    Directory of Open Access Journals (Sweden)

    I. Miller

    2016-09-01

    Full Text Available Male Wistar rats with different thyroid status (eu-, hypothyroid were exposed to 0, 3 or 30 mg/kg body weight of the flame retardant HBCD for 7 days and obtained data compared with a previous study in females, “Hexabromocyclododecane (HBCD induced changes in the liver proteome of eu- and hypothyroid female rats” (Miller et al., 2016 [1]. Specifically, proteomic investigation of liver protein patterns obtained by 2D-DIGE was performed and differences between animals groups recorded, based on the factors exposure, thyroid status and gender. All proteins with significantly changed abundance in any of these comparisons were identified by mass spectrometry. General, hormone and proteomic data of both the present and the previous studies are discussed in Miller et al. (2016 [1] and in "Gender specific differences in the liver proteome of rats exposed to hexabromocyclododecane (HBCD" Miller et al. (2016 [2].

  2. Suppression of the lipopolysaccharide-induced expression of MARCKS-related protein (MRP) affects transmigration in activated RAW264.7 cells.

    Science.gov (United States)

    Chun, Kwang-Rok; Bae, Eun Mi; Kim, Jae-Kwan; Suk, Kyoungho; Lee, Won-Ha

    2009-01-01

    The molecular action mechanism of MRP, one of the protein kinase C (PKC) substrates, has been under intense investigation, but reports on its role in macrophage function remain controversial. The treatment of macrophage cell lines with bacterial lipopolysaccharide (LPS) induced a high level of MRP expression suggesting that MRP plays a role in the function of activated macrophages. In order to investigate the role of MRP in activated RAW264.7 cells, we stably transfected MRP-specific shRNA expression constructs and tested for alterations in macrophage-related functions. The down-regulation of MRP expression resulted in a marked reduction in chemotaxis toward MCP-1 or extracellular matrix proteins. Furthermore, pharmacological inhibitors of PKC significantly inhibited the chemotaxis in RAW264.7 cells. These data reveals the pivotal role of MRP in the transmigration of activated RAW264.7 cells.

  3. A single amino acid mutation affects elicitor and expansins-like activities of cerato-platanin, a non-catalytic fungal protein.

    Directory of Open Access Journals (Sweden)

    Simone Luti

    Full Text Available Cerato-platanin (CP is a non-catalytic, cysteine-rich protein, the first member of the cerato-platanin family. It is a single-domain protein with a double Ψ/β barrel domain resembling the D1 domain of plant and bacterial expansins. Similarly to expansins, CP shows a cell wall-loosening activity on cellulose and can be defined as an expanisin-like protein, in spite of the missing D2 domain, normally present in plant expansins. The weakening activity shown on cellulose may facilitate the CP-host interaction, corroborating the role of CP in eliciting plant defence response. Indeed, CP is an elicitor of primary defences acting as a Pathogen-Associated Molecular Patterns (PAMP. So far, structure-function relationship study has been mainly performed on the bacterial BsEXLX1 expansin, probably due to difficulties in expressing plant expansins in heterologous systems. Here, we report a subcloning and purification method of CP in the engineered E. coli SHuffle cells, which proved to be suitable to obtain the properly folded and biologically active protein. The method also enabled the production of the mutant D77A, rationally designed to be inactive. The wild-type and the mutated CP were characterized for cellulose weakening activity and for PAMP activity (i.e. induction of Reactive Oxygen Species synthesis and phytoalexins production. Our analysis reveals that the carboxyl group of D77 is crucial for expansin-like and PAMP activities, thus permitting to establish a correlation between the ability to weaken cellulose and the capacity to induce defence responses in plants. Our results enable the structural and functional characterization of a mono-domain eukaryotic expansin and identify the essential role of a specific aspartic residue in cellulose weakening.

  4. High doses of ursodeoxycholic acid up-regulate the expression of placental breast cancer resistance protein in patients affected by intrahepatic cholestasis of pregnancy.

    Directory of Open Access Journals (Sweden)

    Francesco Azzaroli

    Full Text Available Ursodeoxycholic acid (UDCA administration in intrahepatic cholestasis of pregnancy (ICP induces bile acids (BA efflux from the foetal compartment, but the molecular basis of this transplacental transport is only partially defined.To determine if placental breast cancer resistance protein (BCRP, able to transport BA, is regulated by UDCA in ICP.32 pregnant women with ICP (14 untreated, 34.9±5.17 years; 18 treated with UDCA--25 mg/Kg/day, 32.7±4.62 years, and 12 healthy controls (33.4±3.32 years agreed to participate in the study. Placentas were obtained at delivery and processed for membrane extraction. BCRP protein expression was evaluated by immunoblotting techniques and chemiluminescence quantified with a luminograph measuring emitted photons; mRNA expression with real time PCR. Statistical differences between groups were evaluated by ANOVA with Dunn's Multiple Comparison test.BCRP was expressed only on the apical membrane of the syncytiotrophoblast. A significant difference was observed among the three groups both for mRNA (ANOVA, p = 0.0074 and protein (ANOVA, p<0.0001 expression. BCRP expression was similar in controls and in the untreated ICP group. UDCA induced a significant increase in placental BCRP mRNA and protein expression compared to controls (350.7±106.3 vs 100±18.68% of controls, p<0.05 and 397.8±56.02 vs 100±11.44% of controls, p<0.001, respectively and untreated ICP (90.29±17.59% of controls, p<0.05 and 155.0±13.87%, p<0.01.Our results confirm that BCRP is expressed only on the apical membrane of the syncytiotrophoblast and show that ICP treatment with high dose UDCA significantly upregulates placental BCRP expression favouring BA efflux from the foetal compartment.

  5. High doses of ursodeoxycholic acid up-regulate the expression of placental breast cancer resistance protein in patients affected by intrahepatic cholestasis of pregnancy.

    Science.gov (United States)

    Azzaroli, Francesco; Raspanti, Maria Elena; Simoni, Patrizia; Montagnani, Marco; Lisotti, Andrea; Cecinato, Paolo; Arena, Rosario; Simonazzi, Giuliana; Farina, Antonio; Rizzo, Nicola; Mazzella, Giuseppe

    2013-01-01

    Ursodeoxycholic acid (UDCA) administration in intrahepatic cholestasis of pregnancy (ICP) induces bile acids (BA) efflux from the foetal compartment, but the molecular basis of this transplacental transport is only partially defined. To determine if placental breast cancer resistance protein (BCRP), able to transport BA, is regulated by UDCA in ICP. 32 pregnant women with ICP (14 untreated, 34.9±5.17 years; 18 treated with UDCA--25 mg/Kg/day, 32.7±4.62 years,) and 12 healthy controls (33.4±3.32 years) agreed to participate in the study. Placentas were obtained at delivery and processed for membrane extraction. BCRP protein expression was evaluated by immunoblotting techniques and chemiluminescence quantified with a luminograph measuring emitted photons; mRNA expression with real time PCR. Statistical differences between groups were evaluated by ANOVA with Dunn's Multiple Comparison test. BCRP was expressed only on the apical membrane of the syncytiotrophoblast. A significant difference was observed among the three groups both for mRNA (ANOVA, p = 0.0074) and protein (ANOVA, p<0.0001) expression. BCRP expression was similar in controls and in the untreated ICP group. UDCA induced a significant increase in placental BCRP mRNA and protein expression compared to controls (350.7±106.3 vs 100±18.68% of controls, p<0.05 and 397.8±56.02 vs 100±11.44% of controls, p<0.001, respectively) and untreated ICP (90.29±17.59% of controls, p<0.05 and 155.0±13.87%, p<0.01). Our results confirm that BCRP is expressed only on the apical membrane of the syncytiotrophoblast and show that ICP treatment with high dose UDCA significantly upregulates placental BCRP expression favouring BA efflux from the foetal compartment.

  6. Isolation of a protein-containing cell surface component from Streptococcus sanguis which affects its adherence to saliva-coated hydroxyapatite.

    OpenAIRE

    Liljemark, W. F.; Bloomquist, C G

    1981-01-01

    The isolation and partial characterization of a protein-containing cell surface component from Streptococcus sanguis which blocks the adherence of this microbe to saliva-coated hydroxyapatite are described. Several methods of extraction were attempted. Sonication of whole cells and cell walls proved to be the most successful and yielded biologically active adherence-blocking components. The adherence-blocking ability of these components was effective in intraspecies blocking experiments. The ...

  7. BAY 81-8973, a full-length recombinant factor VIII: Human heat shock protein 70 improves the manufacturing process without affecting clinical safety.

    Science.gov (United States)

    Maas Enriquez, Monika; Thrift, John; Garger, Stephen; Katterle, Yvonne

    2016-11-01

    BAY 81-8973 is a full-length, unmodified recombinant human factor VIII (FVIII) approved for the treatment of hemophilia A. BAY 81-8973 has the same amino acid sequence as the currently marketed sucrose-formulated recombinant FVIII (rFVIII-FS) product and is produced using additional advanced manufacturing technologies. One of the key manufacturing advances for BAY 81-8973 is introduction of the gene for human heat shock protein 70 (HSP70) into the rFVIII-FS cell line. HSP70 facilitates proper folding of proteins, enhances cell survival by inhibiting apoptosis, and potentially impacts rFVIII glycosylation. HSP70 expression in the BAY 81-8973 cell line along with other manufacturing advances resulted in a higher-producing cell line and improvements in the pharmacokinetics of the final product as determined in clinical studies. HSP70 protein is not detected in the harvest or in the final BAY 81-8973 product. However, because this is a new process, clinical trial safety assessments included monitoring for anti-HSP70 antibodies. Most patients, across all age groups, had low levels of anti-HSP70 antibodies before exposure to the investigational product. During BAY 81-8973 treatment, 5% of patients had sporadic increases in anti-HSP70 antibody levels above a predefined threshold (cutoff value, 239 ng/mL). No clinical symptoms related to anti-HSP70 antibody development occurred. In conclusion, addition of HSP70 to the BAY 81-8973 cell line is an innovative technology for manufacturing rFVIII aimed at improving protein folding and expression. Improved pharmacokinetics and no effect on safety of BAY 81-8973 were observed in clinical trials in patients with hemophilia A. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  8. The Ketogenic Diet Alters the Hypoxic Response and Affects Expression of Proteins Associated with Angiogenesis, Invasive Potential and Vascular Permeability in a Mouse Glioma Model.

    Directory of Open Access Journals (Sweden)

    Eric C Woolf

    Full Text Available The successful treatment of malignant gliomas remains a challenge despite the current standard of care, which consists of surgery, radiation and temozolomide. Advances in the survival of brain cancer patients require the design of new therapeutic approaches that take advantage of common phenotypes such as the altered metabolism found in cancer cells. It has therefore been postulated that the high-fat, low-carbohydrate, adequate protein ketogenic diet (KD may be useful in the treatment of brain tumors. We have demonstrated that the KD enhances survival and potentiates standard therapy in a mouse model of malignant glioma, yet the mechanisms are not fully understood.To explore the effects of the KD on various aspects of tumor growth and progression, we used the immunocompetent, syngeneic GL261-Luc2 mouse model of malignant glioma.Tumors from animals maintained on KD showed reduced expression of the hypoxia marker carbonic anhydrase 9, hypoxia inducible factor 1-alpha, and decreased activation of nuclear factor kappa B. Additionally, tumors from animals maintained on KD had reduced tumor microvasculature and decreased expression of vascular endothelial growth factor receptor 2, matrix metalloproteinase-2 and vimentin. Peritumoral edema was significantly reduced in animals fed the KD and protein analyses showed altered expression of zona occludens-1 and aquaporin-4.The KD directly or indirectly alters the expression of several proteins involved in malignant progression and may be a useful tool for the treatment of gliomas.

  9. The Ketogenic Diet Alters the Hypoxic Response and Affects Expression of Proteins Associated with Angiogenesis, Invasive Potential and Vascular Permeability in a Mouse Glioma Model.

    Science.gov (United States)

    Woolf, Eric C; Curley, Kara L; Liu, Qingwei; Turner, Gregory H; Charlton, Julie A; Preul, Mark C; Scheck, Adrienne C

    2015-01-01

    The successful treatment of malignant gliomas remains a challenge despite the current standard of care, which consists of surgery, radiation and temozolomide. Advances in the survival of brain cancer patients require the design of new therapeutic approaches that take advantage of common phenotypes such as the altered metabolism found in cancer cells. It has therefore been postulated that the high-fat, low-carbohydrate, adequate protein ketogenic diet (KD) may be useful in the treatment of brain tumors. We have demonstrated that the KD enhances survival and potentiates standard therapy in a mouse model of malignant glioma, yet the mechanisms are not fully understood. To explore the effects of the KD on various aspects of tumor growth and progression, we used the immunocompetent, syngeneic GL261-Luc2 mouse model of malignant glioma. Tumors from animals maintained on KD showed reduced expression of the hypoxia marker carbonic anhydrase 9, hypoxia inducible factor 1-alpha, and decreased activation of nuclear factor kappa B. Additionally, tumors from animals maintained on KD had reduced tumor microvasculature and decreased expression of vascular endothelial growth factor receptor 2, matrix metalloproteinase-2 and vimentin. Peritumoral edema was significantly reduced in animals fed the KD and protein analyses showed altered expression of zona occludens-1 and aquaporin-4. The KD directly or indirectly alters the expression of several proteins involved in malignant progression and may be a useful tool for the treatment of gliomas.

  10. Residues at a Single Site Differentiate Animal Cryptochromes from Cyclobutane Pyrimidine Dimer Photolyases by Affecting the Proteins' Preferences for Reduced FAD.

    Science.gov (United States)

    Xu, Lei; Wen, Bin; Wang, Yuan; Tian, Changqing; Wu, Mingcai; Zhu, Guoping

    2017-06-19

    Cryptochromes (CRYs) and photolyases belong to the cryptochrome/photolyase family (CPF). Reduced FAD is essential for photolyases to photorepair UV-induced cyclobutane pyrimidine dimers (CPDs) or 6-4 photoproducts in DNA. In Drosophila CRY (dCRY, a type I animal CRY), FAD is converted to the anionic radical but not to the reduced state upon illumination, which might induce a conformational change in the protein to relay the light signal downstream. To explore the foundation of these differences, multiple sequence alignment of 650 CPF protein sequences was performed. We identified a site facing FAD (Ala377 in Escherichia coli CPD photolyase and Val415 in dCRY), hereafter referred to as "site 377", that was distinctly conserved across these sequences: CPD photolyases often had Ala, Ser, or Asn at this site, whereas animal CRYs had Ile, Leu, or Val. The binding affinity for reduced FAD, but not the photorepair activity of E. coli photolyase, was dramatically impaired when replacing Ala377 with any of the three CRY residues. Conversely, in V415S and V415N mutants of dCRY, FAD was photoreduced to its fully reduced state after prolonged illumination, and light-dependent conformational changes of these mutants were severely inhibited. We speculate that the residues at site 377 play a key role in the different preferences of CPF proteins for reduced FAD, which differentiate animal CRYs from CPD photolyases. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Decorin protein core affects the global gene expression profile of the tumor microenvironment in a triple-negative orthotopic breast carcinoma xenograft model.

    Directory of Open Access Journals (Sweden)

    Simone Buraschi

    Full Text Available Decorin, a member of the small leucine-rich proteoglycan gene family, exists and functions wholly within the tumor microenvironment to suppress tumorigenesis by directly targeting and antagonizing multiple receptor tyrosine kinases, such as the EGFR and Met. This leads to potent and sustained signal attenuation, growth arrest, and angiostasis. We thus sought to evaluate the tumoricidal benefits of systemic decorin on a triple-negative orthotopic breast carcinoma xenograft model. To this end, we employed a novel high-density mixed expression array capable of differentiating and simultaneously measuring gene signatures of both Mus musculus (stromal and Homo sapiens (epithelial tissue origins. We found that decorin protein core modulated the differential expression of 374 genes within the stromal compartment of the tumor xenograft. Further, our top gene ontology classes strongly suggests an unexpected and preferential role for decorin protein core to inhibit genes necessary for immunomodulatory responses while simultaneously inducing expression of those possessing cellular adhesion and tumor suppressive gene properties. Rigorous verification of the top scoring candidates led to the discovery of three genes heretofore unlinked to malignant breast cancer that were reproducibly found to be induced in several models of tumor stroma. Collectively, our data provide highly novel and unexpected stromal gene signatures as a direct function of systemic administration of decorin protein core and reveals a fundamental basis of action for decorin to modulate the tumor stroma as a biological mechanism for the ascribed anti-tumorigenic properties.

  12. NS1 Protein Amino Acid Changes D189N and V194I Affect Interferon Responses, Thermosensitivity, and Virulence of Circulating H3N2 Human Influenza A Viruses.

    Science.gov (United States)

    Nogales, Aitor; Martinez-Sobrido, Luis; Topham, David J; DeDiego, Marta L

    2017-03-01

    Influenza virus NS1 protein is a nonstructural, multifunctional protein that counteracts host innate immune responses, modulating virus pathogenesis. NS1 protein variability in subjects infected with H3N2 influenza A viruses (IAVs) during the 2010/2011 season was analyzed, and amino acid changes in residues 86, 189, and 194 were found. The consequences of these mutations for the NS1-mediated inhibition of IFN responses and the pathogenesis of the virus were evaluated, showing that NS1 mutations D189N and V194I impaired the ability of the NS1 protein to inhibit general gene expression, most probably because these mutations decreased the binding of NS1 to the cleavage and polyadenylation specificity factor 30 (CPSF30). A recombinant A/Puerto Rico/8/34 (PR8) H1N1 virus encoding the H3N2 NS1-D189N protein was slightly attenuated, whereas the virus encoding the H3N2 NS1-V194I protein was further attenuated in mice. The higher attenuation of this virus could not be explained by differences in the ability of the two NS1 proteins to counteract host innate immune responses, indicating that another factor must be responsible. In fact, we showed that the virus encoding the H3N2 NS1-V194I protein demonstrated a temperature-sensitive (ts) phenotype, providing a most likely explanation for the stronger attenuation observed. As far as we know, this is the first description of a mutation in NS1 residue 194 conferring a ts phenotype. These studies are relevant in order to identify new residues important for NS1 functions and in human influenza virus surveillance to assess mutations affecting the pathogenicity of circulating viruses. IMPORTANCE Influenza viral infections represent a serious public health problem, with influenza virus causing a contagious respiratory disease that is most effectively prevented through vaccination. The multifunctional nonstructural protein 1 (NS1) is the main viral factor counteracting the host antiviral response. Therefore, influenza virus

  13. TGF-β1 signaling pathway serves a role in HepG2 cell regulation by affecting the protein expression of PCNA, gankyrin, p115, XIAP and survivin

    Science.gov (United States)

    Wang, Xin-Hong; Chen, Zhi-Guo; Xu, Rui-Ling; Lv, Cheng-Qian; Liu, Jing; Du, Bing

    2017-01-01

    The transforming growth factor-β (TGF-β) signaling pathway serves a key role in the pathogenesis of liver cancer. To investigate the association between TGF-β1 and the following proteins: Proliferating cell nuclear antigen (PCNA), gankyrin, general vesicular transport factor p115 (p115), X-linked inhibitor of apoptosis protein (XIAP) and survivin, HepG2 liver cancer cells were transfected with small interfering RNA (siRNA) directed against TGF-β1, or were treated with exogenous TGF-β1. TGF-β1 protein expression levels were assessed at 72 and 96 h using western blotting, cell growth was evaluated using a Cell Counting kit-8 assay, and flow cytometry was used to examine cell cycle distribution and apoptosis. In addition, PCNA, gankyrin, p115, XIAP and survivin protein levels were evaluated using western blotting. TGF-β1 protein expression levels were decreased at 72 and 96 h following siRNA transfection, indicating that the siRNA against TGF-β1 was effective. In the TGF-β1-knockdown group, the HepG2 cells exhibited G1 or S-phase cell cycle arrest; therefore, the number of G2-phase cells was decreased, cell growth was inhibited and apoptotic peaks were observed. By contrast, no significant alteration in cell cycle distribution or apoptosis was observed in the cells treated with exogenous TGF-β1. In the exogenous TGF-β1 group, PCNA and XIAP protein expression levels were increased, whereas gankyrin, p115 and survivin protein expression was observed to be dependent on the duration of treatment. By contrast, PCNA, gankyrin, XIAP and survivin protein expression decreased following TGF-β1 knockdown; however, p115 protein expression increased. In conclusion, the TGF-β1 signaling pathway may affect cell growth, cell cycle distribution and apoptosis through the regulation of PCNA, gankyrin, p115, XIAP and survivin protein expression in liver cancer. The results of the present study may improve the current understanding of the role of the TGF-β signaling pathway

  14. Distribution of non-collagenous dentin matrix proteins and proteoglycans, and their relation to calcium accumulation in bisphosphonate-affected rat incisors.

    Science.gov (United States)

    Ohma, N; Takagi, Y; Takano, Y

    2000-06-01

    It has been reported that multiple injections of 1-hydroxyethylidene- 1,1-bisphosphonate (HEBP) to rats prevent mineralization of incisor dentin, thereby revealing high concentrations of calcium in the non-mineralized matrix of circumpulpal dentin. To identify the molecules responsible for calcium accumulation in circumpulpal dentin matrix, rats were injected daily with HEBP (8 mg P/kg) for 7 d, and the incisors processed for various histochemical and immunohistochemical staining of non-collagenous matrices of dentin. Cuprolinic blue reactions for proteoglycans (PGs) were equally distributed in non-mineralized matrix of mantle and circumpulpal dentin layers. Dentin sialoprotein (DSP) and osteopontin (OPN) immunoreactions were found in non-mineralized circumpulpal dentin matrix, but not in mantle dentin. In normal incisors, however, predentin matrix showing significant DSP immunoreactivity was negative for Ca-GBHA reactions. HEBP-affected, non-mineralized OPN immunopositive bone matrix was also non-reactive for calcium. From these observations, neither PGs, OPN nor DSP appear to be responsible for calcium accumulation in HEBP-affected circumpulpal dentin. Stains-all reactive component, possibly dentin phosphoprotein (DPP), only showed the same distribution as that of Ca-GBHA in both HEBP-affected and normal dentin matrix, implicating a possible contribution of DPP to calcium accumulation in circumpulpal dentin and, hence, to appositional mineralization of dentin.

  15. The UL24 protein of herpes simplex virus 1 affects the sub-cellular distribution of viral glycoproteins involved in fusion

    Energy Technology Data Exchange (ETDEWEB)

    Ben Abdeljelil, Nawel; Rochette, Pierre-Alexandre; Pearson, Angela, E-mail: angela.pearson@iaf.inrs.ca

    2013-09-15

    Mutations in UL24 of herpes simplex virus type 1 can lead to a syncytial phenotype. We hypothesized that UL24 affects the sub-cellular distribution of viral glycoproteins involved in fusion. In non-immortalized human foreskin fibroblasts (HFFs) we detected viral glycoproteins B (gB), gD, gH and gL present in extended blotches throughout the cytoplasm with limited nuclear membrane staining; however, in HFFs infected with a UL24-deficient virus (UL24X), staining for the viral glycoproteins appeared as long, thin streaks running across the cell. Interestingly, there was a decrease in co-localized staining of gB and gD with F-actin at late times in UL24X-infected HFFs. Treatment with chemical agents that perturbed the actin cytoskeleton hindered the formation of UL24X-induced syncytia in these cells. These data support a model whereby the UL24 syncytial phenotype results from a mislocalization of viral glycoproteins late in infection. - Highlights: • UL24 affects the sub-cellular distribution of viral glycoproteins required for fusion. • Sub-cellular distribution of viral glycoproteins varies in cell-type dependent manner. • Drugs targeting actin microfilaments affect formation of UL24-related syncytia in HFFs.

  16. Overexpression of retinal degeneration slow (RDS protein adversely affects rods in the rd7 model of enhanced S-cone syndrome.

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    Dibyendu Chakraborty

    Full Text Available The nuclear receptor NR2E3 promotes expression of rod photoreceptor genes while repressing cone genes. Mice lacking NR2E3 (Nr2e3(rd7/rd7 referred to here as rd7 are a model for enhanced S-cone syndrome, a disease associated with increased sensitivity to blue light and night blindness. Rd7 retinas have reduced levels of the outer segment (OS structural protein retinal degeneration slow (RDS. We test the hypothesis that increasing RDS levels would improve the Rd7 phenotype. Transgenic mice over-expressing normal mouse peripherin/RDS (NMP in rods and cones were crossed onto the rd7 background. Disease phenotypes were assessed in NMP/rd7 eyes and compared to wild-type (WT and rd7 eyes at postnatal day 30. NMP/rd7 retinas expressed total RDS (transgenic and endogenous message at WT levels, and NMP protein was correctly localized to the OS. NMP/rd7 retinas have shorter OSs compared to rd7 and WT and significantly reduced number of rosettes. NMP/rd7 mice also exhibited significant deficits in scotopic ERG amplitudes compared to rd7 while photopic amplitudes remained unaffected. Protein levels of rhodopsin, RDS, and the RDS homologue ROM-1 were significantly reduced in the NMP/rd7 retinas compared to rd7. We show that correcting the levels of RDS gene expression does not improve the phenotype of the rd7 suggesting that RDS deficiency is not responsible for the defect in this model. We suggest that the specific rod defect in the NMP/rd7 is likely associated with ongoing problems in the rd7 that are related to the expression of cone genes in rod cells, a characteristic of the model.

  17. C4b-binding protein is present in affected areas of myocardial infarction during the acute inflammatory phase and covers a larger area than C3.

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    Leendert A Trouw

    Full Text Available BACKGROUND: During myocardial infarction reduced blood flow in the heart muscle results in cell death. These dying/dead cells have been reported to bind several plasma proteins such as IgM and C-reactive protein (CRP. In the present study we investigated whether fluid-phase complement inhibitor C4b-binding protein (C4BP would also bind to the infarcted heart tissue. METHODS AND FINDINGS: Initial studies using immunohistochemistry on tissue arrays for several cardiovascular disorders indicated that C4BP can be found in heart tissue in several cardiac diseases but that it is most abundantly found in acute myocardial infarction (AMI. This condition was studied in more detail by analyzing the time window and extent of C4BP positivity. The binding of C4BP correlates to the same locations as C3b, a marker known to correlate to the patterns of IgM and CRP staining. Based on criteria that describe the time after infarction we were able to pinpoint that C4BP binding is a relatively early marker of tissue damage in myocardial infarction with a peak of binding between 12 hours and 5 days subsequent to AMI, the phase in which infiltration of neutrophilic granulocytes in the heart is the most extensive. CONCLUSIONS: C4BP, an important fluid-phase inhibitor of the classical and lectin pathway of complement activation binds to jeopardized cardiomyocytes early after AMI and co-localizes to other well known markers such as C3b.

  18. Consuming High-Protein Soy Snacks Affects Appetite Control, Satiety, and Diet Quality in Young People and Influences Select Aspects of Mood and Cognition.

    Science.gov (United States)

    Leidy, Heather J; Todd, Chelsie B; Zino, Adam Z; Immel, Jordan E; Mukherjea, Ratna; Shafer, Rebecca S; Ortinau, Laura C; Braun, Michelle

    2015-07-01

    Data concerning the effects of afternoon snacking on ingestive behavior, mood, and cognition are limited. The purpose of this study was to compare 1088 kJ of high-protein (HP) or high-fat (HF) afternoon snacks vs. no snacking on appetite, food intake, mood, and cognition in adolescents. Thirty-one healthy adolescents (age: 17 ± 1 y) consumed the following afternoon snacks (in randomized order) for 3 d: HP snack (26 g of protein/6 g of fat per 27 g of carbohydrates), HF snack (4 g of protein/12 g of fat per 32 g of carbohydrates), and no snack (NoS). On day 4 of each treatment, the participants completed an 8-h testing day containing pre- and postsnack appetite questionnaires, food cue-stimulated functional MRI brain scans, mood, cognitive function, and eating initiation. Ad libitum dinner and evening snacks were provided and assessed. HP, but not HF, delayed eating initiation vs. NoS (P brain regions controlling food motivation/reward vs. NoS (P sugar evening snacks than NoS (P < 0.01) and HF (P = 0.09). Although no treatment effects were detected for mood and cognition, HP tended to reduce confusion-bewilderment (P = 0.07) and increase cognitive flexibility (P = 0.09), whereas NoS reduced tension-anxiety (P < 0.05) and vigor-activity (P < 0.05). Afternoon snacking, particularly on HP soy foods, improves appetite, satiety, and diet quality in adolescents, while beneficially influencing aspects of mood and cognition. This trial was registered at clinicaltrials.gov as NCT01781286. © 2015 American Society for Nutrition.

  19. Co-Ingestion of Whey Protein with a Carbohydrate-Rich Breakfast Does Not Affect Glycemia, Insulinemia or Subjective Appetite Following a Subsequent Meal in Healthy Males

    Directory of Open Access Journals (Sweden)

    Dean M. Allerton

    2016-02-01

    Full Text Available We aimed to assess postprandial metabolic and appetite responses to a mixed-macronutrient lunch following prior addition of whey protein to a carbohydrate-rich breakfast. Ten healthy males (age: 24 ± 1 years; body mass index (BMI: 24.5 ± 0.7 kg/m2 completed three trials in a non-isocaloric, crossover design. A carbohydrate-rich breakfast (93 g carbohydrate; 1799 kJ was consumed with (CHO + WP or without (CHO 20 g whey protein isolate (373 kJ, or breakfast was omitted (NB. At 180 min, participants consumed a mixed-macronutrient lunch meal. Venous blood was sampled at 15 min intervals following each meal and every 30 min thereafter, while subjective appetite sensations were collected every 30 min throughout. Post-breakfast insulinemia was greater after CHO + WP (time-averaged area under the curve (AUC0––180 min: 193.1 ± 26.3 pmol/L, compared to CHO (154.7 ± 18.5 pmol/L and NB (46.1 ± 8.0 pmol/L; p < 0.05, with no difference in post-breakfast (0–180 min glycemia (CHO + WP, 3.8 ± 0.2 mmol/L; CHO, 4.2 ± 0.2 mmol/L; NB, 4.2 ± 0.1 mmol/L; p = 0.247. There were no post-lunch (0–180 min effects of condition on glycemia (p = 0.492, insulinemia (p = 0.338 or subjective appetite (p > 0.05. Adding whey protein to a carbohydrate-rich breakfast enhanced the acute postprandial insulin response, without influencing metabolic or appetite responses following a subsequent mixed-macronutrient meal.

  20. Cellular Prion Protein Combined with Galectin-3 and -6 Affects the Infectivity Titer of an Endogenous Retrovirus Assayed in Hippocampal Neuronal Cells

    OpenAIRE

    Kim, Boe-Hyun; Shin, Hae-Young; Goto, Joy J.; Carp, Richard I.; Choi, Eun-Kyoung; Kim, Yong-Sun

    2016-01-01

    Prion diseases are infectious and fatal neurodegenerative diseases which require the cellular prion protein, PrPC, for development of diseases. The current study shows that the PrPC augments infectivity and plaque formation of a mouse endogenous retrovirus, MuLV. We have established four neuronal cell lines expressing mouse PrPC, PrP+/+; two express wild type PrPC (MoPrP wild ) and the other two express mutant PrPC (MoPrP mut ). Infection of neuronal cells from various PrP+/+ and PrP-/- (MoPr...

  1. Sophorolipid Butyl Ester Diacetate Does Not Affect Macrophage Polarization but Enhances Astrocytic Glial Fibrillary Acidic Protein Expression at Micromolar Concentrations in Vitro.

    Science.gov (United States)

    Ziemba, Alexis M; Gottipati, Manoj K; Totsingan, Filbert; Hanes, Cheryl M; Gross, Richard A; Lennartz, Michelle R; Gilbert, Ryan J

    2017-04-19

    Peritoneal macrophages (PMACs) and spinal cord astrocytes were exposed to varying concentrations of soluble sophorolipid butyl ester diacetate (SLBEDA) in vitro. Macrophages and astrocytes demonstrated no decrease in viability in response to SLBEDA. Studying pro- and anti-inflammatory genes, PMACs did not show a shift toward a pro-inflammatory phenotype. However, at higher concentrations (3 and 30 μM), astrocytes showed an increase in their expression of glial acidic fibrillary protein. This novel category of compounds poses low risk to PMAC and astrocyte viability; however, the effect on PMAC polarization and astrocyte reactivity requires more elucidation.

  2. Conserved retinoblastoma protein-binding motif in human cytomegalovirus UL97 kinase minimally impacts viral replication but affects susceptibility to maribavir

    Directory of Open Access Journals (Sweden)

    Chou Sunwen

    2009-01-01

    Full Text Available Abstract The UL97 kinase has been shown to phosphorylate and inactivate the retinoblastoma protein (Rb and has three consensus Rb-binding motifs that might contribute to this activity. Recombinant viruses containing mutations in the Rb-binding motifs generally replicated well in human foreskin fibroblasts with only a slight delay in replication kinetics. Their susceptibility to the specific UL97 kinase inhibitor, maribavir, was also examined. Mutation of the amino terminal motif, which is involved in the inactivation of Rb, also renders the virus hypersensitive to the drug and suggests that the motif may play a role in its mechanism of action.

  3. The outer membrane protein Omp35 affects the reduction of Fe(III, nitrate, and fumarate by Shewanella oneidensis MR-1

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    Myers Charles R

    2004-06-01

    Full Text Available Abstract Background Shewanella oneidensis MR-1 uses several electron acceptors to support anaerobic respiration including insoluble species such as iron(III and manganese(IV oxides, and soluble species such as nitrate, fumarate, dimethylsulfoxide and many others. MR-1 has complex branched electron transport chains that include components in the cytoplasmic membrane, periplasm, and outer membrane (OM. Previous studies have implicated a role for anaerobically upregulated OM electron transport components in the use of insoluble electron acceptors, and have suggested that other OM components may also contribute to insoluble electron acceptor use. In this study, the role for an anaerobically upregulated 35-kDa OM protein (Omp35 in the use of anaerobic electron acceptors was explored. Results Omp35 was purified from the OM of anaerobically grown cells, the gene encoding Omp35 was identified, and an omp35 null mutant (OMP35-1 was isolated and characterized. Although OMP35-1 grew on all electron acceptors tested, a significant lag was seen when grown on fumarate, nitrate, and Fe(III. Complementation studies confirmed that the phenotype of OMP35-1 was due to the loss of Omp35. Despite its requirement for wild-type rates of electron acceptor use, analysis of Omp35 protein and predicted sequence did not identify any electron transport moieties or predicted motifs. OMP35-1 had normal levels and distribution of known electron transport components including quinones, cytochromes, and fumarate reductase. Omp35 is related to putative porins from MR-1 and S. frigidimarina as well as to the PorA porin from Neisseria meningitidis. Subcellular fraction analysis confirmed that Omp35 is an OM protein. The seven-fold anaerobic upregulation of Omp35 is mediated post-transcriptionally. Conclusion Omp35 is a putative porin in the OM of MR-1 that is markedly upregulated anaerobically by a post-transcriptional mechanism. Omp35 is required for normal rates of growth on Fe

  4. Epidermal Growth Factor Cytoplasmic Domain Affects ErbB Protein Degradation by the Lysosomal and Ubiquitin-Proteasome Pathway in Human Cancer Cells

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    Aleksandra Glogowska

    2012-05-01

    Full Text Available The cytoplasmic domains of EGF-like ligands, including EGF cytoplasmic domain (EGFcyt, have important biological functions. Using specific constructs and peptides of human EGF cytoplasmic domain, we demonstrate that EGFcyt facilitates lysosomal and proteasomal protein degradation, and this coincided with growth inhibition of human thyroid and glioma carcinoma cells. EGFcyt and exon 22–23-encoded peptide (EGF22.23 enhanced procathepsin B (procathB expression and procathB-mediated lysosomal degradation of EGFR/ErbB1 as determined by inhibitors for procathB and the lysosomal ATPase inhibitor BafA1. Presence of mbEGFctF, EGFcyt, EGF22.23, and exon 23-encoded peptides suppressed the expression of the deubiqitinating enzyme ubiquitin C-terminal hydrolase-L1 (UCH-L1. This coincided with hyperubiquitination of total cellular proteins and ErbB1/2 and reduced proteasome activity. Upon small interfering RNA-mediated silencing of endogenously expressed UCH-L1, a similar hyperubiquitinylation phenotype, reduced ErbB1/2 content, and attenuated growth was observed. The exon 23-encoded peptide region of EGFcyt was important for these biologic actions. Structural homology modeling of human EGFcyt showed that this molecular region formed an exposed surface loop. Peptides derived from this EGFcyt loop structure may aid in the design of novel peptide therapeutics aimed at inhibiting growth of cancer cells.

  5. The composition of readily available carbon sources produced by fermentation of fish faeces is affected by dietary protein:energy ratios

    DEFF Research Database (Denmark)

    Letelier-Gordo, Carlos Octavio; Larsen, Bodil Katrine; Dalsgaard, Johanne

    2017-01-01

    , 17, 19, 21 and 23 g/MJ) to rainbow trout (Oncorhynchus mykiss) on the production of volatile fatty acids (VFAs) and ethanol during 7 days fermentation of the produced fish faeces. The total yields of VFAs and ethanol obtained (expressed as chemical oxygen demand (COD)) ranged between 0.21–0.24 g...... of acetic and valeric acid. Changing the diet composition thus affects the composition of readily available carbon that can be derived from the faeces. This can be applied to enhance on-farm single sludge denitrification and reduce the need for adding external carbon sources such as e.g. methanol....

  6. Higher Total Protein Intake and Change in Total Protein Intake Affect Body Composition but Not Metabolic Syndrome Indexes in Middle-Aged Overweight and Obese Adults Who Perform Resistance and Aerobic Exercise for 36 Weeks.

    Science.gov (United States)

    Campbell, Wayne W; Kim, Jung Eun; Amankwaah, Akua F; Gordon, Susannah L; Weinheimer-Haus, Eileen M

    2015-09-01

    Studies assessing the effects of protein supplementation on changes in body composition (BC) and health rarely consider the impact of total protein intake (TPro) or the change in TPro (CTPro) from participants' usual diets. This secondary data analysis assessed the impact of TPro and CTPro on changes in BC and metabolic syndrome (MetS) indexes in overweight and obese middle-aged adults who participated in an exercise training program. Men and women [n = 117; age: 50 ± 0.7 y, body mass index (BMI; in kg/m(2)): 30.1 ± 0.3; means ± SEs] performed resistance exercise 2 d/wk and aerobic exercise 1 d/wk and consumed an unrestricted diet along with 200-kcal supplements (0, 10, 20, or 30 g whey protein) twice daily for 36 wk. Protein intake was assessed via 4-d food records. Multiple linear regression model and stratified analysis were applied for data analyses. Among all subjects, TPro and CTPro were inversely associated (P changes in body mass, fat mass (FM), and BMI. Changes in BC were different (P changes in FM, %FM, and %LM. The gain in LM was not different among groups. In addition, MetS indexes were not influenced by TPro and CTPro. In conjunction with exercise training, higher TPro promoted positive changes in BC but not in MetS indexes in overweight and obese middle-aged adults. Changes in TPro from before to during the intervention also influenced BC responses and should be considered in future research when different TPro is achieved via diet or supplements. This trial was registered at clinicaltrials.gov as NCT00812409. © 2015 American Society for Nutrition.

  7. A single amino acid substitution of the human immunodeficiency virus type 1 capsid protein affects viral sensitivity to TRIM5α

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    Shioda Tatsuo

    2010-07-01

    Full Text Available Abstract Background Human immunodeficiency virus type 1 (HIV-1 productively infects only humans and chimpanzees but not Old World monkeys, such as rhesus and cynomolgus (CM monkeys. To establish a monkey model of HIV-1/AIDS, several HIV-1 derivatives have been constructed. We previously reported that efficient replication of HIV-1 in CM cells was achieved after we replaced the loop between α-helices 6 and 7 (L6/7 of the capsid protein (CA with that of SIVmac239 in addition to the loop between α-helices 4 and 5 (L4/5 and vif. This virus (NL-4/5S6/7SvifS was supposed to escape from host restriction factors cyclophilin A, CM TRIM5α, and APOBEC3G. However, the replicative capability of NL-4/5S6/7SvifS in human cells was severely impaired. Results By long-term cultivation of human CEMss cells infected with NL-4/5S6/7SvifS, we succeeded in rescuing the impaired replicative capability of the virus in human cells. Sequence analysis of the CA region of the adapted virus revealed a G-to-E substitution at the 116th position of the CA (G116E. Introduction of this substitution into the molecular DNA clone of NL-4/5S6/7SvifS indeed improved the virus' replicative capability in human cells. Although the G116E substitution occurred during long-term cultivation of human cells infected with NL-4/5S6/7SvifS, the viruses with G116E unexpectedly became resistant to CM, but not human TRIM5α-mediated restriction. The 3-D model showed that position 116 is located in the 6th helix near L4/5 and L6/7 and is apparently exposed to the protein surface. The amino acid substitution at the 116th position caused a change in the structure of the protein surface because of the replacement of G (which has no side chain with E (which has a long negatively charged side chain. Conclusions We succeeded in rescuing the impaired replicative capability of NL-4/5S6/7SvifS and report a mutation that improved the replicative capability of the virus. Unexpectedly, HIV-1 with this

  8. The O-mannosylation and production of recombinant APA (45/47 KDa) protein from Mycobacterium tuberculosis in Streptomyces lividans is affected by culture conditions in shake flasks.

    Science.gov (United States)

    Gamboa-Suasnavart, Ramsés A; Valdez-Cruz, Norma A; Cordova-Dávalos, Laura E; Martínez-Sotelo, José A; Servín-González, Luis; Espitia, Clara; Trujillo-Roldán, Mauricio A

    2011-12-20

    The Ala-Pro-rich O-glycoprotein known as the 45/47 kDa or APA antigen from Mycobacterium tuberculosis is an immunodominant adhesin restricted to mycobacterium genus and has been proposed as an alternative candidate to generate a new vaccine against tuberculosis or for diagnosis kits. In this work, the recombinant O-glycoprotein APA was produced by the non-pathogenic filamentous bacteria Streptomyces lividans, evaluating three different culture conditions. This strain is known for its ability to produce heterologous proteins in a shorter time compared to M. tuberculosis. Three different shake flask geometries were used to provide different shear and oxygenation conditions; and the impact of those conditions on the morphology of S. lividans and the production of rAPA was characterized and evaluated. Small unbranched free filaments and mycelial clumps were found in baffled and coiled shake flasks, but one order of magnitude larger pellets were found in conventional shake flasks. The production of rAPA is around 3 times higher in small mycelia than in larger pellets, most probably due to difficulties in mass transfer inside pellets. Moreover, there are four putative sites of O-mannosylation in native APA, one of which is located at the carboxy-terminal region. The carbohydrate composition of this site was determined for rAPA by mass spectrometry analysis, and was found to contain different glycoforms depending on culture conditions. Up to two mannoses residues were found in cultures carried out in conventional shake flasks, and up to five mannoses residues were determined in coiled and baffled shake flasks. The shear and/or oxygenation parameters determine the bacterial morphology, the productivity, and the O-mannosylation of rAPA in S. lividans. As demonstrated here, culture conditions have to be carefully controlled in order to obtain recombinant O-glycosylated proteins with similar "quality" in bacteria, particularly, if the protein activity depends on the

  9. The O-mannosylation and production of recombinant APA (45/47 KDa protein from Mycobacterium tuberculosis in Streptomyces lividans is affected by culture conditions in shake flasks

    Directory of Open Access Journals (Sweden)

    Gamboa-Suasnavart Ramsés A

    2011-12-01

    Full Text Available Abstract Background The Ala-Pro-rich O-glycoprotein known as the 45/47 kDa or APA antigen from Mycobacterium tuberculosis is an immunodominant adhesin restricted to mycobacterium genus and has been proposed as an alternative candidate to generate a new vaccine against tuberculosis or for diagnosis kits. In this work, the recombinant O-glycoprotein APA was produced by the non-pathogenic filamentous bacteria Streptomyces lividans, evaluating three different culture conditions. This strain is known for its ability to produce heterologous proteins in a shorter time compared to M. tuberculosis. Results Three different shake flask geometries were used to provide different shear and oxygenation conditions; and the impact of those conditions on the morphology of S. lividans and the production of rAPA was characterized and evaluated. Small unbranched free filaments and mycelial clumps were found in baffled and coiled shake flasks, but one order of magnitude larger pellets were found in conventional shake flasks. The production of rAPA is around 3 times higher in small mycelia than in larger pellets, most probably due to difficulties in mass transfer inside pellets. Moreover, there are four putative sites of O-mannosylation in native APA, one of which is located at the carboxy-terminal region. The carbohydrate composition of this site was determined for rAPA by mass spectrometry analysis, and was found to contain different glycoforms depending on culture conditions. Up to two mannoses residues were found in cultures carried out in conventional shake flasks, and up to five mannoses residues were determined in coiled and baffled shake flasks. Conclusions The shear and/or oxygenation parameters determine the bacterial morphology, the productivity, and the O-mannosylation of rAPA in S. lividans. As demonstrated here, culture conditions have to be carefully controlled in order to obtain recombinant O-glycosylated proteins with similar "quality" in bacteria

  10. A Costa Rican family affected with Charcot-Marie-Tooth disease due to the myelin protein zero (MPZ p.Thr124Met mutation shares the Belgian haplotype

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    Alejandro Leal

    2014-12-01

    Full Text Available The p.Thr124Met mutation in the myelin protein zero (MPZ causes the Charcot-Marie-Tooth disease type 2J, a peripheral neuropathy with additional symptoms as pupillary alterations and deafness. It was observed in several families around the world originating e. g. from Germany, Belgium, Japan, Italy and North America. Here we report Central American patients originating from a family in Costa Rica carrying this mutation. Clinical, electrophysiological and molecular analysis of patients and controls were performed, including gene and linked markers´ sequencing. Carriers share almost the entire haplotype with two non related Belgian CMT patients. As a result of the haplotype analysis, based on ten markers (seven SNPs, two microsatellites and an intronic polyA stretch, the founder effect hypothesis for this allele migration is suggestive. Rev. Biol. Trop. 62 (4: 1285-1293. Epub 2014 December 01.

  11. Isolation of a protein-containing cell surface component from Streptococcus sanguis which affects its adherence to saliva-coated hydroxyapatite.

    Science.gov (United States)

    Liljemark, W F; Bloomquist, C G

    1981-01-01

    The isolation and partial characterization of a protein-containing cell surface component from Streptococcus sanguis which blocks the adherence of this microbe to saliva-coated hydroxyapatite are described. Several methods of extraction were attempted. Sonication of whole cells and cell walls proved to be the most successful and yielded biologically active adherence-blocking components. The adherence-blocking ability of these components was effective in intraspecies blocking experiments. The extract obtained from cell walls of S. sanguis was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and shown to contain one major and two to three minor bands when stained with Coomassie blue. The molecular weight of the major band was estimated to be 70,000 to 90,000. Gel filtration of the sonified cell wall extract on 10% agarose yielded two active adherence-blocking peaks, the void volume and a second peak. Images PMID:6273317

  12. The junctional SR protein JP-45 affects the functional expression of the voltage-dependent Ca2+ channel Cav1.1.

    Science.gov (United States)

    Anderson, Ayuk A; Altafaj, Xavier; Zheng, Zhenlin; Wang, Zhong-Min; Delbono, Osvaldo; Ronjat, Michel; Treves, Susan; Zorzato, Francesco

    2006-05-15

    JP-45, an integral protein of the junctional face membrane of the skeletal muscle sarcoplasmic reticulum (SR), colocalizes with its Ca2+ -release channel (the ryanodine receptor), and interacts with calsequestrin and the skeletal-muscle dihydropyridine receptor Cav1. We have identified the domains of JP-45 and the Cav1.1 involved in this interaction, and investigated the functional effect of JP-45. The cytoplasmic domain of JP-45, comprising residues 1-80, interacts with Cav1.1. JP-45 interacts with two distinct and functionally relevant domains of Cav1.1, the I-II loop and the C-terminal region. Interaction between JP-45 and the I-II loop occurs through the alpha-interacting domain in the I-II loop. beta1a, a Cav1 subunit, also interacts with the cytosolic domain of JP-45, and its presence drastically reduces the interaction between JP-45 and the I-II loop. The functional effect of JP-45 on Cav1.1 activity was assessed by investigating charge movement in differentiated C2C12 myotubes after overexpression or depletion of JP-45. Overexpression of JP-45 decreased peak charge-movement and shifted VQ1/2 to a more negative potential (-10 mV). JP-45 depletion decreased both the content of Cav1.1 and peak charge-movements. Our data demonstrate that JP-45 is an important protein for functional expression of voltage-dependent Ca2+ channels.

  13. The ESAT-6 Protein of Mycobacterium tuberculosis Interacts with Beta-2-Microglobulin (β2M) Affecting Antigen Presentation Function of Macrophage

    Science.gov (United States)

    Parveen, Nazia; Jha, Vishwanath; Valluri, Vijaya Lakshmi; Ghosh, Sudip; Mukhopadhyay, Sangita

    2014-01-01

    ESAT-6, an abundantly secreted protein of Mycobacterium tuberculosis (M. tuberculosis) is an important virulence factor, inactivation of which leads to reduced virulence of M. tuberculosis. ESAT-6 alone, or in complex with its chaperone CFP-10 (ESAT-6:CFP-10), is known to modulate host immune responses; however, the detailed mechanisms are not well understood. The structure of ESAT-6 or ESAT-6:CFP-10 complex does not suggest presence of enzymatic or DNA-binding activities. Therefore, we hypothesized that the crucial role played by ESAT-6 in the virulence of mycobacteria could be due to its interaction with some host cellular factors. Using a yeast two-hybrid screening, we identified that ESAT-6 interacts with the host protein beta-2-microglobulin (β2M), which was further confirmed by other assays, like GST pull down, co-immunoprecipitation and surface plasmon resonance. The C-terminal six amino acid residues (90–95) of ESAT-6 were found to be essential for this interaction. ESAT-6, in complex with CFP-10, also interacts with β2M. We found that ESAT-6/ESAT-6:CFP-10 can enter into the endoplasmic reticulum where it sequesters β2M to inhibit cell surface expression of MHC-I-β2M complexes, resulting in downregulation of class I-mediated antigen presentation. Interestingly, the ESAT-6:β2M complex could be detected in pleural biopsies of individuals suffering from pleural tuberculosis. Our data highlight a novel mechanism by which M. tuberculosis may undermine the host adaptive immune responses to establish a successful infection. Identification of such novel interactions may help us in designing small molecule inhibitors as well as effective vaccine design against tuberculosis. PMID:25356553

  14. Affect Regulation

    DEFF Research Database (Denmark)

    Pedersen, Signe Holm; Poulsen, Stig Bernt; Lunn, Susanne

    2014-01-01

    Gergely and colleagues’ state that their Social Biofeedback Theory of Parental Affect Mirroring” can be seen as a kind of operationalization of the classical psychoanalytic concepts of holding, containing and mirroring. This article examines to what extent the social biofeedback theory of parental...... affect mirroring may be understood as a specification of these concepts. It is argued that despite similarities at a descriptive level the concepts are embedded in theories with different ideas of subjectivity. Hence an understanding of the concept of affect regulation as a concretizisation...... and specification of the classical concepts dilutes the complexity of both the concept of affect regulation and of the classical concepts....

  15. Pre-Slaughter Stress Affects Ryanodine Receptor Protein Gene Expression and the Water-Holding Capacity in Fillets of the Nile Tilapia.

    Directory of Open Access Journals (Sweden)

    Elenice S R Goes

    Full Text Available Current study evaluated the effect of pre-slaughter stress on serum cortisol levels, pH, colorimetry, water-holding capacity (WHC and gene expression of ryanodine receptors (RyR1 and RyR3 in the Nile tilapia. A 3x4 factorial scheme experiment was conducted comprising three densities (100, 200, 400 kg/m³ with four transportation times (60, 120, 180, and 240 minutes.Transportation times alone reduced cortisol levels up to 180 minutes, followed by increased WHC and mRNA expression, RyR1 and RyR3 (200 kg/m³ density. No effect of density x transportation time interacted on the evaluated parameters. Results provided the first evidence that pre-slaughter stress affected ryanodine gene expression receptors and, consequently, the water-holding capacity in tilapia fillets.

  16. All-trans retinoic acid inhibits KIT activity and induces apoptosis in gastrointestinal stromal tumor GIST-T1 cell line by affecting on the expression of survivin and Bax protein

    Directory of Open Access Journals (Sweden)

    Taguchi Takahiro

    2010-12-01

    Full Text Available Abstract Background Imatinib, a selective tyrosine kinase inhibitor, has been used as a standard first-line therapy for irresectable and metastasized gastrointestinal stromal tumor (GIST patients. Unfortunately, most patients responding to imatinib will eventually exhibit imatinib-resistance, the cause of which is not fully understood. The serious clinical problem of imatinib-resistance demands alternative therapeutic strategy. This study was conducted to investigate the effect of all-trans retinoic acid (ATRA on GIST cell lines. Methods Cell proliferation was determined by trypan blue dye exclusion test. Western blot analysis was performed to test the expression of activated KIT, its downstream proteins, and apoptosis associated proteins. The cytotoxic interactions of imatinib with ATRA were evaluated using the isobologram of Steel and Peckham. Results and conclusion In this work, for the first time we have demonstrated that ATRA affected on cell proliferation of GIST-T1 and GIST-882 cell line through inhibition of cell growth in a dose dependent manner and induced apoptosis. High dose of ATRA induced morphologic change in GIST-T1 cells, rounded-up cells, and activated the caspase-3 protein. In further examination, we found that the ATRA-induced apoptosis in GIST-T1 cells was accompanied by the down-regulated expression of survivin and up-regulated expression of Bax protein. Moreover, ATRA suppressed the activity of KIT protein in GIST-T1 cells and its downstream signal, AKT activity, but not MAPK activity. We also have demonstrated that combination of ATRA with imatinib showed additive effect by isobologram, suggesting that the combination of ATRA and imatinib may be a novel potential therapeutic option for GIST treatment. Furthermore, the scracht assay result suggested that ATRA was a potential reagent to prevent the invasion or metastasis of GIST cells.

  17. Proanthocyanidins from the American Cranberry (Vaccinium macrocarpon Induce Cell Cycle Alterations in DU145 Human Prostate Cancer Cells in Vitro by Affecting the Expression of Cell Cycle-Associated Proteins

    Directory of Open Access Journals (Sweden)

    Joseph Kim

    2014-04-01

    Full Text Available Background: Prostate cancer is one of the most common cancers in the world. There are genetic and environmental factors that can potentially impact the development and progression of many types of cancer, including prostate cancer. As a consequence of environmental factors, such as diet having a potential effect on the development of prostate cancer, considerable interest in the possible health benefits associated with the inclusion and consumption of certain foods in the diet exists. Context and purpose of this study: This study describes the effects of a proanthocyanidinenriched fraction (PACs isolated from the American cranberry (Vaccinium macrocarpon on the behaviour of androgen-refractory (insensitive DU145 human prostate cancer cells in vitro. Results: Following treatment of DU145 human prostate cancer cells with 25 µg/mL of PACs for six hours, PACs significantly decreased the cellular viability of DU145 cells. PACs treatment (25 µg/mL for 6 hours of DU145 cells increased the proportion of cells in the G2-M phase of the cell cycle and decreased the proportion of cells in the G1 phase of the cell cycle. These alterations were associated with changes in cell cycle regulatory proteins and other cell cycle associated proteins. PACs increased the expression of cyclin E, cyclin D1, CDK2 and CDK4, and decreased the expression of cyclin A and cyclin B1. The protein expression level of p27 increased, and the protein expression levels of p16INK4a, p21, and pRBp107 decreased in response to PACs treatment. The protein expression level of pRBp130 was unchanged in Functional Foods in Health and Disease 2014; 4(2:130- 146 Page 131 of 146 response to PACs treatment. Conclusions: These findings demonstrate that proanthocyanidins from the American cranberry can affect the behaviour of human prostate cancer cells in vitro and further support the potential health benefits associated with cranberries.

  18. Maternal low-protein diet affects epigenetic regulation of hepatic mitochondrial DNA transcription in a sex-specific manner in newborn piglets associated with GR binding to its promoter.

    Directory of Open Access Journals (Sweden)

    Yimin Jia

    Full Text Available Mitochondrial oxidative phosphorylation (OXPHOS plays an important role in energy homeostasis by controlling electron transfer and ATP generation. Maternal malnutrition during pregnancy affects mitochondrial (mt DNA-encoded OXPHOS activity in offspring, yet it is unknown whether epigenetic mechanism is involved in the transcriptional regulation of mtDNA-encoded OXPHOS genes. In this study, 14 primiparous purebred Meishan sows were fed either standard- (SP, 12% crude protein or low-protein (LP; 6% crude protein diets throughout gestation, and the hepatic expression and transcriptional regulation of mtDNA-encoded OXPHOS genes were analyzed in newborn piglets. Maternal low protein diet decreased hepatic mtDNA copy number in males, but not in females. LP male piglets had significantly higher hepatic AMP concentration and low energy charge, which was accompanied by enhanced mRNA expression of NADH dehydrogenase subunits 6, cytochrome c oxidase subunit 1, 2, 3 and cytochrome b, as well as increased cytochrome c oxidase enzyme activity. In contrast, LP female piglets showed significantly lower hepatic AMP concentrations and higher energy charge with no alterations in OXPHOS gene expression. Moreover, LP males demonstrated higher glucocorticoid receptor (GR binding to the mtDNA promoter compared with SP males, which was accompanied by lower cytosine methylation and hydroxymethylation on mtDNA promoter. Interestingly, opposite changes were seen in females, which showed diminished GR binding and enriched cytosine methylation and hydroxymethylation on mtDNA promoter. These results suggest that maternal low protein diet during pregnancy causes sex-dependent epigenetic alterations in mtDNA-encoded OXPHOS gene expression, possibly GR is involved in mtDNA transcription regulation.

  19. Multidrug resistance-associated protein 2 (MRP2/ABCC2) haplotypes significantly affect the pharmacokinetics of tacrolimus in kidney transplant recipients

    Science.gov (United States)

    Ogasawara, Ken; Chitnis, Shripad D.; Gohh, Reginald Y.; Christians, Uwe; Akhlaghi, Fatemeh

    2013-01-01

    BACKGROUND AND OBJECTIVE Tacrolimus is an immunosuppressive drug used for the prevention of the allograft rejection in the kidney allograft recipients. It exhibits a narrow therapeutic index and a large pharmacokinetic variability. Tacrolimus is mainly metabolized by cytochrome P450 (CYP) 3A4 and 3A5, and effluxed via ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp), encoded by ABCB1 gene. The influence of CYP3A5*3 on the pharmacokinetics of tacrolimus has been well characterized. On the other hand, the contribution of polymorphisms in other genes is controversial. In addition, the involvement of other efflux transporter than P-gp in tacrolimus disposition is uncertain. The present study was designed to investigate the effects of genetic polymorphisms of CYP3As and efflux transporters on the pharmacokinetics of tacrolimus. SUBJECTS AND METHODS A total of 500 blood concentrations of tacrolimus from 102 adult stable kidney transplant recipients were included in the analyses. Genetic polymorphisms in CYP3A4 and CYP3A5 genes as well as the genes of efflux transporters including P-gp (ABCB1), multidrug resistance-associated protein (MRP2/ABCC2) and breast cancer resistance protein (BCRP/ABCG2) were genotyped. For ABCC2 gene, haplotypes were determined as follows: H1 (wild type), H2 (1249G>A), H9 (3972C>T) and H12 (−24C>T and 3972C>T). Population pharmacokinetic analysis was performed using nonlinear mixed effects modeling. RESULTS Analyses revealed that CYP3A5 expressers (CYP3A5*1 carriers) and MRP2 high activity group (ABCC2 H2/H2 and H1/H2) decreased the dose-normalized trough concentration of tacrolimus by 2.3-fold (ptacrolimus was best described using a two-compartment model with first order absorption and an absorption lag time. In the population pharmacokinetic analysis, CYP3A5 expressers and MRP2 high activity groups were identified as the significant covariates for tacrolimus apparent clearance expressed as 20.7 × (Age/50)−0.78 × 2

  20. Cardiovascular risk prediction by N-terminal pro brain natriuretic peptide and high sensitivity C-reactive protein is affected by age and sex

    DEFF Research Database (Denmark)

    Olsen, M.H.; Hansen, T.W.; Christensen, M.K.

    2008-01-01

    BACKGROUND: Previous studies have shown that the urine albumin/creatinine ratio (UACR), high sensitivity C-reactive protein (hsCRP) and N-terminal pro brain natriuretic peptide (Nt-proBNP) predict cardiovascular events in a general population aged 41, 51, 61 or 71 years. This study investigated...... factors, UACR, hsCRP and Nt-proBNP. The composite cardiovascular endpoint (CEP) of cardiovascular death and non-fatal stroke or myocardial infarction was assessed after 9.5 years. RESULTS: In Cox regression analyses predicting CEP, the effects of log(hsCRP) and log(Nt-proBNP) were modulated by sex (P ....05) and age (P effect of logUACR was not significantly modulated by age or sex. Log(hsCRP)/SD did not predict CEP in women, but did predict CEP in 41 plus 51-year-old men [hazard ratio (HR) 1.71; 95% confidence interval, 1.1-2.6; P

  1. Experimental exposure to cadmium affects metallothionein-like protein levels but not survival and growth in wolf spiders from polluted and reference populations

    Energy Technology Data Exchange (ETDEWEB)

    Eraly, Debbie, E-mail: debbie.eraly@ugent.b [Terrestrial Ecology Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent (Belgium); Hendrickx, Frederik, E-mail: frederik.hendrickx@naturalsciences.b [Terrestrial Ecology Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent (Belgium); Royal Belgian Institute of Natural Sciences, Department of Entomology, Vautierstraat 29, 1000 Brussels (Belgium); Bervoets, Lieven, E-mail: lieven.bervoets@ua.ac.b [Ecophysiology, Biochemistry and Toxicology Group, Department of Biology, University of Antwerp, Groenenborgerlaan 171, 2020 Antwerp (Belgium); Lens, Luc, E-mail: luc.lens@ugent.b [Terrestrial Ecology Unit, Department of Biology, Ghent University, K.L. Ledeganckstraat 35, 9000 Ghent (Belgium)

    2010-06-15

    Both local adaptation and acclimation in tolerance mechanisms may allow populations to persist under metal pollution. However, both mechanisms are presumed to incur (energetic) costs and to trade-off with other life-history traits. To test this hypothesis, we exposed Pardosa saltans (Lycosidae) spiderlings originating from metal-polluted and unpolluted sites to a controlled cadmium (Cd) treatment, and compared contents of metal-binding metallothionein-like proteins (MTLPs), internal metal concentrations, and individual survival and growth rates with a reference treatment. While increased MTLP concentrations in offspring originating from both polluted and unpolluted populations upon exposure indicates a plastic tolerance mechanism, survival and growth rates remain largely unaffected, independent of the population of origin. However, MTLP and Cd concentrations were not significantly correlated. We suggest that MTLP production may be an important mechanism enabling P. saltans populations to persist in ecosystems polluted with heavy metals above a certain level. - Spiders from metal-polluted and unpolluted populations show a similar increase in MTLP production when exposed to Cd, with unaffected growth and survival.

  2. Does estrogen affect the development of abnormal vascular function in offspring of rats fed a low-protein diet in pregnancy?

    Science.gov (United States)

    Musha, Yuka; Itoh, Shigeru; Hanson, Mark A; Kinoshita, Katsuyuki

    2006-06-01

    It is established that there are gender-related differences in the effects on offspring blood pressure induced by maternal protein restriction in animal studies. Since such effects may depend on estrogen levels, we hypothesized that lower estrogen would induce an earlier onset of hypertension caused by maternal under-nutrition. Wistar rats were fed a diet containing either 18% (C) or 9% (R) casein throughout pregnancy. Half of the offspring in both C and R groups were ovariectomized on day 50 (CX, RX), and the other half underwent a sham operation (CO, RO). On d 175, offspring were killed for small artery reactivity and histologic investigation. Birth weight and later growth were not significantly different between C and R. RX had higher systolic blood pressure than CX on d125, but no difference was seen between RO and CO. On d 175, systolic blood pressure was higher in R than in C, whether or not ovariectomized. Dilator responses to acetylcholine and bradykinin in small mesenteric arteries were significantly attenuated in RX, although responses to SNP and isoprenaline showed no attenuation in R. The ratio of coronary peri-vascular fibrosis to total vascular area was higher in R, and the fibrosis became prominent in ovariectomized rats. These findings suggest that estrogen plays an important role in limiting the elevation of offspring blood pressure induced by maternal under-nutrition, possibly via BK-mediated mechanisms. The processes may underlie gender and life course patterns of hypertension and also the developmental origins of this disease.

  3. Changes in dietary zinc and copper affect zinc-status indicators of postmenopausal women, notably, extracellular superoxide dismutase and amyloid precursor proteins.

    Science.gov (United States)

    Davis, C D; Milne, D B; Nielsen, F H

    2000-03-01

    Zinc is an essential trace element for human health and well-being; however, methods currently available for the assessment of zinc status in humans are unsatisfactory. The objective was to critically evaluate the use of various indicators of zinc status in humans in a controlled metabolic ward study. Indicators of zinc status were measured in 25 healthy postmenopausal women aged 64.9 +/- 6.7 y. After a 10-d equilibration period, volunteers consumed a diet with either a low (1 mg/d; n = 12) or a high (3 mg/d; n = 13) copper content based on a total energy content of 8.4 MJ. They received the same amount of copper throughout the study. Both groups were fed the basal diet (3 mg Zn/d) with no zinc supplement for one 90-d period, and the diet supplemented with 50 mg Zn/d for another 90-d period. Zinc supplementation significantly increased (P < 0.0001) extracellular but not erythrocyte superoxide dismutase activity. This increase was more apparent when subjects were fed the low-copper diet. Zinc supplementation in combination with the low-copper diet significantly decreased (P < 0.01) amyloid precursor protein expression in platelets. Other indicators of zinc status that were significantly elevated after zinc supplementation were as follows: plasma zinc and free thyroxine concentrations and mononuclear 5'-nucleotidase activity. The measurement of serum extracellular superoxide dismutase activity may be useful as a marker for the functional assessment of zinc status in humans.

  4. Mutations in the capsid protein of Brome mosaic virus affecting encapsidation eliminate vesicle induction in planta: implications for virus cell-to-cell spread.

    Science.gov (United States)

    Bamunusinghe, Devinka; Chaturvedi, Sonali; Seo, Jang-Kyun; Rao, A L N

    2013-08-01

    Positive-strand RNA viruses are known to rearrange the endomembrane network to make it more conducive for replication, maturation, or egress. Our previous transmission electron microscopic (TEM) analysis showed that ectopic expression of wild-type (wt) capsid protein (CP) of Brome mosaic virus (BMV) has an intrinsic property of modifying the endoplasmic reticulum (ER) to induce vesicles similar to those present in wt BMV infection. In this study, we evaluated the functional significance of CP-mediated vesicle induction to the BMV infection cycle in planta. Consequently, the cytopathologic changes induced by wt CP or its mutants defective in virion assembly due to mutations engineered in either N- or C-proximal domains were comparatively analyzed by TEM in two susceptible (Nicotiana benthamiana and Chenopodium quinoa) and one nonhost (N. clevelandii) plant species. The results showed that in susceptible hosts, CP-mediated ER-derived vesicle induction is contingent on the expression of encapsidation-competent CP. In contrast, unlike in N. benthamiana and C. quinoa, transient expression of wt CP in nonhost N. clevelandii plants eliminated vesicle induction. Additionally, comparative source-to-sink analysis of virus spread in leaves of N. benthamiana and N. clevelandii coexpressing wt BMV and Cucumber mosaic virus (CMV) showed that despite trans-encapsidation, CMV failed to complement the defective cell-to-cell movement of BMV. The significance and relation of CP-mediated vesicle induction to virus cell-to-cell movement are discussed.

  5. Odorant-binding proteins OBP57d and OBP57e affect taste perception and host-plant preference in Drosophila sechellia.

    Directory of Open Access Journals (Sweden)

    Takashi Matsuo

    2007-05-01

    Full Text Available Despite its morphological similarity to the other species in the Drosophila melanogaster species complex, D. sechellia has evolved distinct physiological and behavioral adaptations to its host plant Morinda citrifolia, commonly known as Tahitian Noni. The odor of the ripe fruit of M. citrifolia originates from hexanoic and octanoic acid. D. sechellia is attracted to these two fatty acids, whereas the other species in the complex are repelled. Here, using interspecies hybrids between D. melanogaster deficiency mutants and D. sechellia, we showed that the Odorant-binding protein 57e (Obp57e gene is involved in the behavioral difference between the species. D. melanogaster knock-out flies for Obp57e and Obp57d showed altered behavioral responses to hexanoic acid and octanoic acid. Furthermore, the introduction of Obp57d and Obp57e from D. simulans and D. sechellia shifted the oviposition site preference of D. melanogaster Obp57d/e(KO flies to that of the original species, confirming the contribution of these genes to D. sechellia's specialization to M. citrifolia. Our finding of the genes involved in host-plant determination may lead to further understanding of mechanisms underlying taste perception, evolution of plant-herbivore interactions, and speciation.

  6. Increased ID4 expression, accompanied by mutant p53 accumulation and loss of BRCA1/2 proteins in triple-negative breast cancer, adversely affects survival.

    Science.gov (United States)

    Thike, Aye A; Tan, Puay H; Ikeda, Murasaki; Iqbal, Jabed

    2016-04-01

    Breast cancer 1 (BRCA1) expression is down-regulated in a significant proportion of non-hereditary breast cancers, in the absence of any mutation. This phenomenon is more pronounced in oestrogen (ER)-negative tumours. Recent studies have suggested that inhibitor of DNA binding 4 (ID4), as well as p53, participate in the transcriptional regulation of BRCA1. Immunohistochemical expression of ID4, BRCA1, BRCA2 and p53 in 699 women with triple-negative breast cancer was investigated using tissue microarrays. The prognostic role of these biomarkers was also evaluated. Survival outcomes were estimated with the Kaplan-Meier method and compared between groups with log-rank statistics. Loss of BRCA1 and BRCA2 expression and overexpression of ID4 and p53 was observed in 75%, 90%, 95% and 66% of tumours, respectively. ID4 expression was increased in higher tumour grade (P P P = 0.037) and p53 accumulation (P triple-negative breast cancers (P = 0.041) and basal-like triple-negative breast cancers (P = 0.026). There is frequent ID4 expression and concomitant loss of BRCA proteins in triple-negative breast cancer. We hypothesize that strong ID4 expression could be useful as a prognostic marker in triple-negative breast cancer, predicting early tumour recurrence. © 2015 John Wiley & Sons Ltd.

  7. Analysis of SAT type foot-and-mouth disease virus capsid proteins and the identification of putative amino acid residues affecting virus stability.

    Science.gov (United States)

    Maree, Francois F; Blignaut, Belinda; de Beer, Tjaart A P; Rieder, Elizabeth

    2013-01-01

    Foot-and-mouth disease virus (FMDV) initiates infection by adhering to integrin receptors on target cells, followed by cell entry and disassembly of the virion through acidification within endosomes. Mild heating of the virions also leads to irreversible dissociation into pentamers, a characteristic linked to reduced vaccine efficacy. In this study, the structural stability of intra- and inter-serotype chimeric SAT2 and SAT3 virus particles to various conditions including low pH, mild temperatures or high ionic strength, was compared. Our results demonstrated that while both the SAT2 and SAT3 infectious capsids displayed different sensitivities in a series of low pH buffers, their stability profiles were comparable at high temperatures or high ionic strength conditions. Recombinant vSAT2 and intra-serotype chimeric viruses were used to map the amino acid differences in the capsid proteins of viruses with disparate low pH stabilities. Four His residues at the inter-pentamer interface were identified that change protonation states at pH 6.0. Of these, the H145 of VP3 appears to be involved in interactions with A141 in VP3 and K63 in VP2, and may be involved in orientating H142 of VP3 for interaction at the inter-pentamer interfaces.

  8. The Hydroxyl at Position C1 of Genipin Is the Active Inhibitory Group that Affects Mitochondrial Uncoupling Protein 2 in Panc-1 Cells.

    Directory of Open Access Journals (Sweden)

    Yang Yang

    Full Text Available Genipin (GNP effectively inhibits uncoupling protein 2 (UCP2, which regulates the leakage of protons across the inner mitochondrial membrane. UCP2 inhibition may induce pancreatic adenocarcinoma cell death by increasing reactive oxygen species (ROS levels. In this study, the hydroxyls at positions C10 (10-OH and C1 (1-OH of GNP were hypothesized to be the active groups that cause these inhibitory effects. Four GNP derivatives in which the hydroxyl at position C10 or C1 was replaced with other chemical groups were synthesized and isolated. Differences in the inhibitory ef