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Sample records for intestinal tissue samples

  1. Development of a chromogenic in situ hybridization for Giardia duodenalis and its application in canine, feline, and porcine intestinal tissues samples.

    Science.gov (United States)

    Weissenböck, Herbert; Ondrovics, Martina; Gurtner, Susanne; Schiessl, Peter; Mostegl, Meike M; Richter, Barbara

    2011-05-01

    In the present study, a chromogenic in situ hybridization for the identification of Giardia duodenalis in paraffin-embedded tissue samples was developed. The sensitivity and specificity of the probe was validated by testing it on cultured reference samples of different assemblages of G. duodenalis as well as culture and tissue samples containing other protozoa and infectious agents. The probe gave a positive reaction with the Giardia samples and a negative reaction with all other samples. Further, the probe was used for screening of histological slides of intestine from different animal species (99 canine samples, 85 feline samples, and 202 porcine samples) for the presence of G. duodenalis trophozoites. With this assay, the parasites were detected in samples from 8 dogs (8.08%), 6 cats (7.06%), and zero pigs. The results clearly indicate that the described method is useful for detection of Giardia trophozoites in routinely processed intestinal tissue of different animal species. © 2011 The Author(s)

  2. Detection of Lawsonia intracellularis in formalin-fixed porcine intestinal tissue samples: comparison of immunofluorescence and in-situ hybridization, and evaluation of the effects of controlled autolysis.

    Science.gov (United States)

    Jensen, T K; Boesen, H T; Vigre, H; Boye, M

    2010-01-01

    Two methods, an immunofluorescence assay (IFA; with a Lawsonia intracellularis-specific monoclonal antibody) and fluorescent in-situ hybridization (FISH; with a specific oligonucleotide probe targeting 16S ribosomal RNA of the bacterium), were compared for their ability to detect L. intracellularis (the cause of porcine proliferative enteritis [PE]) in formalin-fixed samples of intestinal tissue. Of 69 intestinal samples with gross lesions of PE, 63 were positive by both FISH and IFA, but six were positive only by IFA. This indicated that the sensitivity of FISH was 91% that of IFA. However, both methods had a specificity of 100%. Fifty normal porcine intestines were negative by both tests. IFA was much less susceptible than FISH to the effects of autolysis. Thus, three of nine samples from pigs with PE were FISH-negative after being kept at 20 degrees C for 4 days, and seven were FISH negative after 2 weeks; after 4 weeks at this temperature, however, six of the nine samples were still IFA positive. After being kept at 4 degrees C for 12 weeks, the majority of samples (> or = 66%) were positive by both methods.

  3. Multispectral tissue characterization for intestinal anastomosis optimization

    Science.gov (United States)

    Cha, Jaepyeong; Shademan, Azad; Le, Hanh N. D.; Decker, Ryan; Kim, Peter C. W.; Kang, Jin U.; Krieger, Axel

    2015-10-01

    Intestinal anastomosis is a surgical procedure that restores bowel continuity after surgical resection to treat intestinal malignancy, inflammation, or obstruction. Despite the routine nature of intestinal anastomosis procedures, the rate of complications is high. Standard visual inspection cannot distinguish the tissue subsurface and small changes in spectral characteristics of the tissue, so existing tissue anastomosis techniques that rely on human vision to guide suturing could lead to problems such as bleeding and leakage from suturing sites. We present a proof-of-concept study using a portable multispectral imaging (MSI) platform for tissue characterization and preoperative surgical planning in intestinal anastomosis. The platform is composed of a fiber ring light-guided MSI system coupled with polarizers and image analysis software. The system is tested on ex vivo porcine intestine tissue, and we demonstrate the feasibility of identifying optimal regions for suture placement.

  4. A new approach to predict human intestinal absorption using porcine intestinal tissue and biorelevant matrices.

    Science.gov (United States)

    Westerhout, Joost; van de Steeg, Evita; Grossouw, Dimitri; Zeijdner, Evelijn E; Krul, Cyrille A M; Verwei, Miriam; Wortelboer, Heleen M

    2014-10-15

    A reliable prediction of the oral bioavailability in humans is crucial and of high interest for pharmaceutical and food industry. The predictive value of currently used in silico methods, in vitro cell lines, ex vivo intestinal tissue and/or in vivo animal studies for human intestinal absorption, however, is often insufficient, especially when food-drug interactions are evaluated. Ideally, for this purpose healthy human intestinal tissue is used, but due to its limited availability there is a need for alternatives. The aim of this study was to evaluate the applicability of healthy porcine intestinal tissue mounted in a newly developed InTESTine™ system to predict human intestinal absorption of compounds with different chemical characteristics, and within biorelevant matrices. To that end, first, a representative set of compounds was chosen of which the apparent permeability (Papp) data in both Caco-2 cells and human intestinal tissue mounted in the Ussing chamber system, and absolute human oral bioavailability were reported. Thereafter, Papp values of the subset were determined in both porcine jejunal tissue and our own Caco-2 cells. In addition, the feasibility of this new approach to study regional differences (duodenum, jejunum, and ileum) in permeability of compounds and to study the effects of luminal factors on permeability was also investigated. For the latter, a comparison was made between the compatibility of porcine intestinal tissue, Caco-2 cells, and Caco-2 cells co-cultured with the mucin producing HT29-MTX cells with biorelevant samples as collected from an in vitro dynamic gastrointestinal model (TIM). The results demonstrated that for the paracellularly transported compounds atenolol, cimetidine, mannitol and ranitidine porcine Papp values are within 3-fold difference of human Papp values, whereas the Caco-2 Papp values are beyond 3-fold difference. Overall, the porcine intestinal tissue Papp values are more comparable to human Papp values (9 out

  5. Detection of Lawsonia intracellularis in formalinfixed Porcine Intestinal Tissue Samples: Comparison of Immunofluorescense and In-situ Hybridization, and Evaluation of the Effectf of Controlled Autolysis

    DEFF Research Database (Denmark)

    Jensen, Tim Kåre; Boesen, H. T.; Vigre, Håkan

    2010-01-01

    methods had a specificity of 100%. Fifty normal porcine intestines were negative by both tests. IFA was much less susceptible than FISH to the effects of autolysis. Thus, three of nine samples from pigs with PE were FISH-negative after being kept at 20°C for 4 days, and seven were FISH negative after 2...

  6. Tissue response after radiation exposure. Intestine

    International Nuclear Information System (INIS)

    Otsuka, Kensuke; Tomita, Masanori; Yamauchi, Motohiro; Iwasaki, Toshiyasu

    2014-01-01

    Gastrointestinal syndrome followed by 'gut death' is due to intestinal disorders. This syndrome is induced by high-dose (>10 Gy) of ionizing radiation. Recovery from the gastrointestinal syndrome would depend on the number of survived clonogens and regeneration capability of crypts. These tissue alterations can be observed by high-dose radiation, however, cellular dynamics in crypts can be affected by low-dose radiation. For example, Potten et al. found that low-dose radiation induce apoptosis of intestinal stem cells, which produce all differentiated function cells. Recently, intestinal stem cells are characterized by molecular markers such as Lgr5. Since intestinal adenomas can be induced by deletion of Apc gene in Lgr5 + stem cells, it is widely recognized that Lgr5 + stem cells are the cell-of-origin of cancer. Duodenal Lgr5 + stem cells are known as radioresistant cells, however, we found that ionizing radiation significantly induces the turnover of colonic Lgr5 + stem cells. Combined with the knowledge of other radioresistant markers, stem-cell dynamics in tissue after irradiation are becoming clear. The present review introduces the history of gastrointestinal syndrome and intestinal stem cells, and discusses those future perspectives. (author)

  7. Intestinal mucosa is a target tissue for pancreatic polypeptide

    International Nuclear Information System (INIS)

    Gilbert, W.R.; Kramer, J.L.; Frank, B.H.; Gingerich, R.L.

    1986-01-01

    Studies were carried out to identify mammalian tissues capable of specifically binding mammalian pancreatic polypeptide (PP). Bovine PP (bPP) radiolabeled with 125 I was purified by HPLC to yield [ 125 I]iodo-(Tyr-27) bPP. The label was injected into three pairs of fasted littermate dogs and allowed to circulate for 5 min. One of the dogs was a control which received an excess of unlabeled porcine PP to provide competition for receptor binding. Unbound bPP was removed by perfusion with Krebs-Ringer bicarbonate and the tissue fixed in situ with Karnovsky's fixative. Tissue samples from various organs were removed, weighed, and counted. The entire gastrointestinal tract demonstrated high levels of 125 I after injection of the labeled peptide. The duodenum, jejunum, ileum, and colon were the only tissues to exhibit specific binding of bPP. These tissues (mucosal and muscle layers) from experimental animals exhibited 31-76% higher binding than the corresponding tissues from the control animals. Sections of the gastrointestinal tract were scraped to separate the mucosal layer from the underlying muscle layer. The mucosal layer of the duodenum, jejunum, and ileum exhibited 145-162% increases in binding compared to the control animals. The muscle layer of these tissues demonstrated no significant increase. These findings demonstrate that mucosal layer of the small intestine is a target tissue for mammalian PP

  8. Distribution of enrofloxacin in intestinal tissue and contents of healthy pigs after oral and intramuscular administrations

    DEFF Research Database (Denmark)

    Wiuff, C.; Lykkesfeldt, J.; Aarestrup, Frank Møller

    2002-01-01

    and colon from pigs killed at 2, 3 and 6 h after dosing. Intramuscular administration resulted in significantly higher concentrations in plasma, intestinal tissue and lymph nodes at 2 h but not at 3 or 6 h compared with p.o. administration. The absorption and distribution phase was longer after oral......The concentration of enrofloxacin in plasma, intestinal tissue, lymph nodes and intestinal contents was investigated in healthy pigs after oral (p.o.) and intramuscular (i.m.) administration of a single dose of 2.5 mg/kg bw. Tissue and content samples were collected from jejunum, ileum, caecum...... administration, and maximum concentrations in tissue and plasma were determined later than after i.m. administration. No difference between route of administration was observed in the intestinal content. Enrofloxacin concentrations in faeces during a 5-day dosing regimen with i.m. and p.o. administration were...

  9. Generation of tissue-engineered small intestine using embryonic stem cell-derived human intestinal organoids.

    Science.gov (United States)

    Finkbeiner, Stacy R; Freeman, Jennifer J; Wieck, Minna M; El-Nachef, Wael; Altheim, Christopher H; Tsai, Yu-Hwai; Huang, Sha; Dyal, Rachel; White, Eric S; Grikscheit, Tracy C; Teitelbaum, Daniel H; Spence, Jason R

    2015-10-12

    Short bowel syndrome (SBS) is characterized by poor nutrient absorption due to a deficit of healthy intestine. Current treatment practices rely on providing supportive medical therapy with parenteral nutrition; while life saving, such interventions are not curative and are still associated with significant co-morbidities. As approaches to lengthen remaining intestinal tissue have been met with only limited success and intestinal transplants have poor survival outcomes, new approaches to treating SBS are necessary. Human intestine derived from embryonic stem cells (hESCs) or induced pluripotent stem cells (iPSCs), called human intestinal organoids (HIOs), have the potential to offer a personalized and scalable source of intestine for regenerative therapies. However, given that HIOs are small three-dimensional structures grown in vitro, methods to generate usable HIO-derived constructs are needed. We investigated the ability of hESCs or HIOs to populate acellular porcine intestinal matrices and artificial polyglycolic/poly L lactic acid (PGA/PLLA) scaffolds, and examined the ability of matrix/scaffolds to thrive when transplanted in vivo. Our results demonstrate that the acellular matrix alone is not sufficient to instruct hESC differentiation towards an endodermal or intestinal fate. We observed that while HIOs reseed acellular porcine matrices in vitro, the HIO-reseeded matrices do not thrive when transplanted in vivo. In contrast, HIO-seeded PGA/PLLA scaffolds thrive in vivo and develop into tissue that looks nearly identical to adult human intestinal tissue. Our results suggest that HIO-seeded PGA/PLLA scaffolds are a promising avenue for developing the mucosal component of tissue engineered human small intestine, which need to be explored further to develop them into fully functional tissue. © 2015. Published by The Company of Biologists Ltd.

  10. Generation of tissue-engineered small intestine using embryonic stem cell-derived human intestinal organoids

    Directory of Open Access Journals (Sweden)

    Stacy R. Finkbeiner

    2015-11-01

    Full Text Available Short bowel syndrome (SBS is characterized by poor nutrient absorption due to a deficit of healthy intestine. Current treatment practices rely on providing supportive medical therapy with parenteral nutrition; while life saving, such interventions are not curative and are still associated with significant co-morbidities. As approaches to lengthen remaining intestinal tissue have been met with only limited success and intestinal transplants have poor survival outcomes, new approaches to treating SBS are necessary. Human intestine derived from embryonic stem cells (hESCs or induced pluripotent stem cells (iPSCs, called human intestinal organoids (HIOs, have the potential to offer a personalized and scalable source of intestine for regenerative therapies. However, given that HIOs are small three-dimensional structures grown in vitro, methods to generate usable HIO-derived constructs are needed. We investigated the ability of hESCs or HIOs to populate acellular porcine intestinal matrices and artificial polyglycolic/poly L lactic acid (PGA/PLLA scaffolds, and examined the ability of matrix/scaffolds to thrive when transplanted in vivo. Our results demonstrate that the acellular matrix alone is not sufficient to instruct hESC differentiation towards an endodermal or intestinal fate. We observed that while HIOs reseed acellular porcine matrices in vitro, the HIO-reseeded matrices do not thrive when transplanted in vivo. In contrast, HIO-seeded PGA/PLLA scaffolds thrive in vivo and develop into tissue that looks nearly identical to adult human intestinal tissue. Our results suggest that HIO-seeded PGA/PLLA scaffolds are a promising avenue for developing the mucosal component of tissue engineered human small intestine, which need to be explored further to develop them into fully functional tissue.

  11. Microbiome Heterogeneity Characterizing Intestinal Tissue and Inflammatory Bowel Disease Phenotype.

    Science.gov (United States)

    Tyler, Andrea D; Kirsch, Richard; Milgrom, Raquel; Stempak, Joanne M; Kabakchiev, Boyko; Silverberg, Mark S

    2016-04-01

    Inflammatory bowel disease has been associated with differential abundance of numerous organisms when compared to healthy controls (HCs); however, few studies have investigated variability in the microbiome across intestinal locations and how this variability might be related to disease location and phenotype. In this study, we have analyzed the microbiome of a large cohort of individuals recruited at Mount Sinai Hospital in Toronto, Canada. Biopsies were taken from subjects with Crohn's disease, ulcerative colitis, and HC, and also individuals having undergone ileal pouch-anal anastomosis for treatment of ulcerative colitis or familial adenomatous polyposis. Microbial 16S rRNA was sequenced using the Illumina MiSeq platform. We observed a great deal of variability in the microbiome characterizing different sampling locations. Samples from pouch and afferent limb were comparable in microbial composition. When comparing sigmoid and terminal ileum samples, more differences were observed. The greatest number of differentially abundant microbes was observed when comparing either pouch or afferent limb samples to sigmoid or terminal ileum. Despite these differences, we were able to observe modest microbial variability between inflammatory bowel disease phenotypes and HCs, even when controlling for sampling location and additional experimental factors. Most detected associations were observed between HCs and Crohn's disease, with decreases in specific genera in the families Ruminococcaceae and Lachnospiraceae characterizing tissue samples from individuals with Crohn's disease. This study highlights important considerations when analyzing the composition of the microbiome and also provides useful insight into differences in the microbiome characterizing these seemingly related phenotypes.

  12. Distribution of enrofloxacin in intestinal tissue and contents of healthy pigs after oral and intramuscular administrations

    DEFF Research Database (Denmark)

    Wiuff, C.; Lykkesfeldt, J.; Aarestrup, Frank Møller

    2002-01-01

    The concentration of enrofloxacin in plasma, intestinal tissue, lymph nodes and intestinal contents was investigated in healthy pigs after oral (p.o.) and intramuscular (i.m.) administration of a single dose of 2.5 mg/kg bw. Tissue and content samples were collected from jejunum, ileum, caecum...... and colon from pigs killed at 2, 3 and 6 h after dosing. Intramuscular administration resulted in significantly higher concentrations in plasma, intestinal tissue and lymph nodes at 2 h but not at 3 or 6 h compared with p.o. administration. The absorption and distribution phase was longer after oral....... On the basis of these results it was concluded that in order to ensure an immediate high concentration of enrofloxacin, and thereby avoid an initial selection for resistant mutants, the intramuscular route seems to be preferable to the oral route....

  13. Lactobacillus rhamnosus GG supernatant upregulates serotonin transporter expression in intestinal epithelial cells and mice intestinal tissues.

    Science.gov (United States)

    Wang, Y M; Ge, X Z; Wang, W Q; Wang, T; Cao, H L; Wang, B L; Wang, B M

    2015-09-01

    The role that probiotics play in relieving irritable bowel syndrome (IBS) has been demonstrated; however, the mechanism by which IBS is affected remains unclear. In this study, serotonin transporter (SERT) mRNA and serotonin transporter protein (SERT-P) levels in HT-29, Caco-2 cells, and mice intestinal tissues were examined after treatment with Lactobacillus rhamnosus GG supernatant (LGG-s). HT-29 and Caco-2 cells were treated with different concentrations of LGG-s for 12 and 24 h and C57BL/6 mice received supplements of different concentrations for 4 weeks. SERT mRNA and SERT-P levels were detected by real-time PCR and Western blotting. SERT mRNA and SERT-P levels in HT-29 and Caco-2 cells were higher than those in the control 24 h after treatment. Undiluted LGG-s upregulated SERT mRNA levels by 9.4-fold in the first week, which dropped in the second week. The double-diluted LGG-s upregulated SERT mRNA by 2.07-fold in the first week; levels dropped to 1.75-fold within the second week and under base expression levels by the third week, while they again climbed to 1.56-fold in the fourth week. The triple-diluted LGG-s could not upregulate SERT mRNA expression until the end of the fourth week. The SERT-P levels in the double-diluted LGG-s group were higher than that in the control but fluctuated with time. SERT-P levels in the triple-diluted LGG-s were higher than that in the control in the last 2 weeks and increased with time. LGG-s can upregulate SERT mRNA and SERT-P levels in intestinal epithelial cells and mice intestinal tissues. © 2015 John Wiley & Sons Ltd.

  14. A Sensitive Medium-Throughput Method to Predict Intestinal Absorption in Humans Using Rat Intestinal Tissue Segments.

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    Da Silva, Laís Cristina; Da Silva, Taynara Lourenço; Antunes, Alisson Henrique; Rezende, Kênnia Rocha

    2015-09-01

    A range of in vitro, ex vivo, and in vivo approaches are currently used for drug development. Highly predictive human intestinal absorption models remain lagging behind the times because of numerous variables concerning permeability through gastrointestinal tract in humans. However, there is a clear need for a drug permeability model early in the drug development process that can balance the requirements for high throughput and effective predictive potential. The present study developed a medium throughput screening Snapwell (MTS-Snapwell) ex vivo model to provide an alternative method to classify drug permeability. Rat small intestine tissue segments were mounted in commercial Snapwell™ inserts. Unidirectional drug transport (A-B) was measured by collecting samples at different time points. Viability of intestinal tissue segments was measured by examining transepithelial electric resistance (TEER) and phenol red and caffeine transport. As a result, the apparent permeability (Papp; ×10(-6) cm/s) was determined for atenolol (10.7 ± 1.2), caffeine (17.6 ± 3.1), cimetidine (6.9 ± 0.1), metoprolol (12.6 ± 0.7), theophylline (15.3 ± 1.6) and, ranitidine (3.8 ± 0.4). All drugs were classified in high/low permeability according to Biopharmaceutics Classification System showing high correlation with human data (r = 0.89). These findings showed a high correlation with human data (r = 0.89), suggesting that this model has potential predictive capacity for paracellular and transcellular passively absorbed molecules. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

  15. SEM investigation of heart tissue samples

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    Saunders, R; Amoroso, M [Physics Department, University of the West Indies, St. Augustine, Trinidad and Tobago, West Indies (Trinidad and Tobago)

    2010-07-01

    We used the scanning electron microscope to examine the cardiac tissue of a cow (Bos taurus), a pig (Sus scrofa), and a human (Homo sapiens). 1mm{sup 3} blocks of left ventricular tissue were prepared for SEM scanning by fixing in 96% ethanol followed by critical point drying (cryofixation), then sputter-coating with gold. The typical ridged structure of the myofibrils was observed for all the species. In addition crystal like structures were found in one of the samples of the heart tissue of the pig. These structures were investigated further using an EDVAC x-ray analysis attachment to the SEM. Elemental x-ray analysis showed highest peaks occurred for gold, followed by carbon, oxygen, magnesium and potassium. As the samples were coated with gold for conductivity, this highest peak is expected. Much lower peaks at carbon, oxygen, magnesium and potassium suggest that a cystallized salt such as a carbonate was present in the tissue before sacrifice.

  16. SEM investigation of heart tissue samples

    Science.gov (United States)

    Saunders, R.; Amoroso, M.

    2010-07-01

    We used the scanning electron microscope to examine the cardiac tissue of a cow (Bos taurus), a pig (Sus scrofa), and a human (Homo sapiens). 1mm3 blocks of left ventricular tissue were prepared for SEM scanning by fixing in 96% ethanol followed by critical point drying (cryofixation), then sputter-coating with gold. The typical ridged structure of the myofibrils was observed for all the species. In addition crystal like structures were found in one of the samples of the heart tissue of the pig. These structures were investigated further using an EDVAC x-ray analysis attachment to the SEM. Elemental x-ray analysis showed highest peaks occurred for gold, followed by carbon, oxygen, magnesium and potassium. As the samples were coated with gold for conductivity, this highest peak is expected. Much lower peaks at carbon, oxygen, magnesium and potassium suggest that a cystallized salt such as a carbonate was present in the tissue before sacrifice.

  17. Identification and quantitive analysis of calcium phosphate microparticles in intestinal tissue by nuclear microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Gomez-Morilla, Inmaculada [Advanced Technology Institute, University of Surrey, Guildford GU2 7XH (United Kingdom)]. E-mail: i.gomez-morilla@surrey.ac.uk; Thoree, Vinay [Gastrointestinal Laboratory, Rayne Institute, St. Thomas' Hospital, London SE1 7EH (United Kingdom); Powell, Jonathan J. [MRC Human Nutrition Research, Elsie Widdowson Laboratory, Fulbourn Road, Cambridge CB1 9NL (United Kingdom); Kirkby, Karen J. [Advanced Technology Institute, University of Surrey, Guildford GU2 7XH (United Kingdom); Grime, Geoffrey W. [Department of Physics, University of Surrey, GU2 7XH (United Kingdom)

    2006-08-15

    Microscopic particles (0.5-2 {mu}m diameter), rich in calcium and phosphorus, are found in the lumen of the mid-distal gut of all mammals investigated, including humans, and these may play a role in immuno-surveillance and immune regulation of antigens from food and symbiotic bacteria that are contained in the gut. Whether these particles can cross in to tissue of the intestinal mucosa is unclear. If so, characterising their morphology and chemical composition is an important task in elucidating their function. The analysis of calcium phosphate in biological tissues has been approached in several ways including optical microscopy, scanning electron microscopy and, most recently in this work, with nuclear microscopy. In this paper, we describe the use of microPIXE and microRBS to locate these particles and to determine, accurately, the ratio of phosphorus to calcium using the information on sample thickness obtained from RBS to allow the PIXE ratios to be corrected. A commercial sample of hydroxy apatite was used to demonstrate accuracy and precision of the technique. Then, in a pilot study on intestinal tissue of mice, we demonstrated the presence of calcium phosphate microparticles, consistent with confocal microscopy observations, and we identified the average molar P:Ca molar ratio as 1.0. Further work will confirm the exact chemical speciation of these particles and will examine the influence of differing calcium containing diets on the formation of these microparticles.

  18. Detection of Tritrichomonas foetus and Pentatrichomonas hominis in intestinal tissue specimens of cats by chromogenic in situ hybridization.

    Science.gov (United States)

    Mostegl, Meike M; Wetscher, Andreas; Richter, Barbara; Nedorost, Nora; Dinhopl, Nora; Weissenböck, Herbert

    2012-02-10

    In this retrospective study 102 cats were analyzed for the presence of trichomonads in intestinal tissue sections using chromogenic in situ hybridization (CISH). Two intestinal trichomonad species are described in cats: Pentatrichomonas hominis and Tritrichomonas foetus. While P. hominis is considered a mere commensal, T. foetus has been found to be the causative agent of feline large-bowel diarrhea. For the detection of both agents within intestinal tissue CISH assays using three different probes were performed. In the first CISH run a probe specific for all relevant members of the order Trichomonadida (OT probe) was used. In a second CISH run all positive samples were further examined on three consecutive tissue sections using the OT probe, a probe specific for the family of Tritrichomonadidae (Tritri probe) and a newly designed probe specifically detecting P. hominis (Penta hom probe). In total, four of the 102 cats were found to be positive with the OT probe. Thereof, one cat gave a positive reaction with the P. hominis probe and three cats were positive with the T. foetus probe. All Trichomonas-positive cats were pure-bred and between 8 and 32 weeks of age. In one cat positive for T. foetus large amounts of parasites were found in the gut lumen and invading the intestinal mucosa. The species of the detected trichomonads were confirmed by polymerase chain reaction and nucleotide sequencing of a part of the 18S ribosomal RNA gene. In this study, the usefulness of CISH to detect intestinal trichomonads within feline tissue samples was shown. Additionally, the specific detection of P. hominis using CISH was established. Generally, it was shown that CISH is well suited for detection and differentiation of trichomonosis in retrospective studies using tissue samples. Copyright © 2011 Elsevier B.V. All rights reserved.

  19. Hydrolysis of Pyrethroids by Human and Rat Tissues: Examination of Intestinal, Liver and Serum Carboxylesterases

    Science.gov (United States)

    Crow, J. Allen; Borazjani, Abdolsamad; Potter, Philip M.; Ross, Matthew K.

    2009-01-01

    Hydrolytic metabolism of pyrethroid insecticides in humans is one of the major catabolic pathways that clear these compounds from the body. Rodent models are often used to determine the disposition and clearance rates of these esterified compounds. In this study the distribution and activities of esterases that catalyze pyrethroid metabolism have been investigated in vitro using several human and rat tissues, including small intestine, liver, and serum. The major esterase in human intestine is carboxylesterase 2 (hCE2). We found that the pyrethroid trans-permethrin is effectively hydrolyzed by a sample of pooled human intestinal microsomes (5 individuals), while deltamethrin and bioresmethrin are not. This result correlates well with the substrate specificity of recombinant hCE2 enzyme. In contrast, a sample of pooled rat intestinal microsomes (5 animals) hydrolyze trans-permethrin 4.5-fold slower than the sample of human intestinal microsomes. Furthermore, it is demonstrated that pooled samples of cytosol from human or rat liver are ~2-fold less hydrolytically active (normalized per mg protein) than the corresponding microsomal fraction toward pyrethroid substrates; however, the cytosolic fractions do have significant amounts (~40%) of the total esteratic activity. Moreover, a 6-fold interindividual variation in carboxylesterase 1 protein expression in human hepatic cytosols was observed. Human serum was shown to lack pyrethroid hydrolytic activity, but rat serum has hydrolytic activity that is attributed to a single CE isozyme. We purified the serum CE enzyme to homogeneity to determine its contribution to pyrethroid metabolism in the rat. Both trans-permethrin and bioresmethrin were effectively cleaved by this serum CE, but deltamethrin, esfenvalerate, alpha-cypermethrin, and cis-permethrin were slowly hydrolyzed. Lastly, two model lipase enzymes were examined for their ability to hydrolyze pyrethroids. However, no hydrolysis products could be detected. Together

  20. Hydrolysis of pyrethroids by human and rat tissues: Examination of intestinal, liver and serum carboxylesterases

    International Nuclear Information System (INIS)

    Crow, J. Allen; Borazjani, Abdolsamad; Potter, Philip M.; Ross, Matthew K.

    2007-01-01

    Hydrolytic metabolism of pyrethroid insecticides in humans is one of the major catabolic pathways that clear these compounds from the body. Rodent models are often used to determine the disposition and clearance rates of these esterified compounds. In this study the distribution and activities of esterases that catalyze pyrethroid metabolism have been investigated in vitro using several human and rat tissues, including small intestine, liver and serum. The major esterase in human intestine is carboxylesterase 2 (hCE2). We found that the pyrethroid trans-permethrin is effectively hydrolyzed by a sample of pooled human intestinal microsomes (5 individuals), while deltamethrin and bioresmethrin are not. This result correlates well with the substrate specificity of recombinant hCE2 enzyme. In contrast, a sample of pooled rat intestinal microsomes (5 animals) hydrolyze trans-permethrin 4.5-fold slower than the sample of human intestinal microsomes. Furthermore, it is demonstrated that pooled samples of cytosol from human or rat liver are ∼ 2-fold less hydrolytically active (normalized per mg protein) than the corresponding microsomal fraction toward pyrethroid substrates; however, the cytosolic fractions do have significant amounts (∼ 40%) of the total esteratic activity. Moreover, a 6-fold interindividual variation in carboxylesterase 1 protein expression in human hepatic cytosols was observed. Human serum was shown to lack pyrethroid hydrolytic activity, but rat serum has hydrolytic activity that is attributed to a single CE isozyme. We purified the serum CE enzyme to homogeneity to determine its contribution to pyrethroid metabolism in the rat. Both trans-permethrin and bioresmethrin were effectively cleaved by this serum CE, but deltamethrin, esfenvalerate, alpha-cypermethrin and cis-permethrin were slowly hydrolyzed. Lastly, two model lipase enzymes were examined for their ability to hydrolyze pyrethroids. However, no hydrolysis products could be detected

  1. Morphological abnormalities and apoptosis in lamellar tissue of equines after intestinal obstruction and treatment with hydrocortisone

    Directory of Open Access Journals (Sweden)

    L.M Laskoski

    2010-12-01

    Full Text Available Four experimental groups of equines were used in order to study morphological abnormalities and apoptosis in lamellar tissue. Group Cg (control was composed of animals without any surgical procedure; group Ig (instrumented, animals that underwent enterotomy; group Tg (treated, animals that were subjected to intestinal obstruction and were treated with hydrocortisone; and group Ug (untreated, animals that were subjected to intestinal obstruction without treatment. The lamellar tissue was analyzed regarding the presence of tissue abnormalities and apoptosis. No morphological abnormalities were observed in animals of surgical groups, and no difference in apoptosis was observed between groups. It was concluded that intestinal obstruction allowed laminitis to develop, probably by systemic activation, and that the maneuvers performed in the enterotomy aggravated the process. Hydrocortisone did not aggravate the lesions of the lamellar tissue

  2. Microbial Population Differentials between Mucosal and Submucosal Intestinal Tissues in Advanced Crohn's Disease of the Ileum.

    Directory of Open Access Journals (Sweden)

    Rodrick J Chiodini

    Full Text Available Since Crohn's disease is a transmural disease, we hypothesized that examination of deep submucosal tissues directly involved in the inflammatory disease process may provide unique insights into bacterial populations transgressing intestinal barriers and bacterial populations more representative of the causes and agents of the disease. We performed deep 16s microbiota sequencing on isolated ilea mucosal and submucosal tissues on 20 patients with Crohn's disease and 15 non-inflammatory bowel disease controls with a depth of coverage averaging 81,500 sequences in each of the 70 DNA samples yielding an overall resolution down to 0.0001% of the bacterial population. Of the 4,802,328 total sequences generated, 98.9% or 4,749,183 sequences aligned with the Kingdom Bacteria that clustered into 8545 unique sequences with <3% divergence or operational taxonomic units enabling the identification of 401 genera and 698 tentative bacterial species. There were significant differences in all taxonomic levels between the submucosal microbiota in Crohn's disease compared to controls, including organisms of the Order Desulfovibrionales that were present within the submucosal tissues of most Crohn's disease patients but absent in the control group. A variety of organisms of the Phylum Firmicutes were increased in the subjacent submucosa as compared to the parallel mucosal tissue including Ruminococcus spp., Oscillospira spp., Pseudobutyrivibrio spp., and Tumebacillus spp. In addition, Propionibacterium spp. and Cloacibacterium spp. were increased as well as large increases in Proteobacteria including Parasutterella spp. and Methylobacterium spp. This is the first study to examine the microbial populations within submucosal tissues of patients with Crohn's disease and to compare microbial communities found deep within the submucosal tissues with those present on mucosal surfaces. Our data demonstrate the existence of a distinct submucosal microbiome and ecosystem

  3. Prediction of drug intestinal absorption in human using the Ussing chamber system: A comparison of intestinal tissues from animals and humans.

    Science.gov (United States)

    Miyake, Masateru; Koga, Toshihisa; Kondo, Satoshi; Yoda, Noriaki; Emoto, Chie; Mukai, Tadashi; Toguchi, Hajime

    2017-01-01

    An adequate evaluation system for drug intestinal absorption is essential in the pharmaceutical industry. Previously, we established a novel prediction system of drug intestinal absorption in humans, using the mini-Ussing chamber equipped with human intestinal tissues. In this system, the TI value was defined as the sum of drug amounts transported to the basal-side component (X corr ) and drug amounts accumulated in the tissue (T corr ), which are normalized by AUC of a drug in the apical compartment, as an index for drug absorption. In order to apply this system to the screening assay, it is important to understand the differences between animal and human tissues in the intestinal absorption of drugs. In this study, the transport index (TI) values of three drugs, with different levels of membrane permeability, were determined to evaluate the rank order of drug absorbability in intestinal tissues from rats, dogs, and monkeys. The TI values in small intestinal tissues in rats and dogs showed a good correlation with those in humans. On the other hand, the correlation of TI values in monkeys was lower compared to rats and dogs. The rank order of the correlation coefficient between human and investigated animal tissues was as follows: dog (r 2 =0.978), rat (r 2 =0.955), and monkey (r 2 =0.620). TI values in large intestinal tissues from rats (r 2 =0.929) and dogs (r 2 =0.808) also showed a good correlation. The obtained TI values in small intestinal tissues in rats and dogs were well correlated with the fraction of drug absorbed (F a ) in humans. From these results, the mini-Ussing chamber, equipped with intestinal tissues in rats and dogs, would be useful as a screening tool in the drug discovery stage. In addition, the obtained TI values can be used for the prediction of the F a in humans. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Identification and characterization of novel gut-associated lymphoid tissues in rat small intestine.

    Science.gov (United States)

    Hitotsumatsu, Osamu; Hamada, Hiromasa; Naganuma, Makoto; Inoue, Nagamu; Ishii, Hiromasa; Hibi, Toshifumi; Ishikawa, Hiromichi

    2005-10-01

    The crypt lamina propria of the mouse small intestine has been shown to harbor multiple tiny clusters filled with c-kit- and interleukin 7 receptor (IL-7R)-positive lympho-hemopoietic cells (cryptopatches; CPs). However, it has remained an open question whether similar lymphoid tissue are present in the gastrointesitinal tract in other animals. In the present study, we investigated whether the small intestine of rats harbored lymphoid tissues similar to mouse CPs. Immunohistochemical and flow cytometric analyses were carried out using various antibodies, including those to c-kit and IL-7R molecules. Lymphocyte-filled villi (LFVs), populated predominantly with c-kit- and IL-7 receptor (IL-7R)-positive cells and less with T cell receptor (TCR)-alphabeta T cells were found throughout the small intestine of young adult rats. Although LFVs were absent from fetal rat intestine, they were first detected at around 2 weeks after birth. Notably, in most LFVs that settled in the antimesenteric wall of the small intestine in young adult rats, immunoglobulin M-positive B cells were also detectable at the bottom of the LFVs. In aged rats, lymphocytes in some LFVs displayed a different phenotype, comprising a large B-cell area that included a germinal center. Thus, these clusters represent the first description of isolated lymphoid follicles (ILFs) in the rat small intestine. The present study provides the first evidence for c-kit- and IL-7R-positive lymphocyte clusters in the rat small intestine. Our data also indicating that LFVs and ILFs may constitute novel organized gut-associated lymphoid tissues in lamina propria of the rat small intestine.

  5. Activation of intestinal epithelial Stat3 orchestrates tissue defense during gastrointestinal infection.

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    Nadine Wittkopf

    Full Text Available Gastrointestinal infections with EHEC and EPEC are responsible for outbreaks of diarrheal diseases and represent a global health problem. Innate first-line-defense mechanisms such as production of mucus and antimicrobial peptides by intestinal epithelial cells are of utmost importance for host control of gastrointestinal infections. For the first time, we directly demonstrate a critical role for Stat3 activation in intestinal epithelial cells upon infection of mice with Citrobacter rodentium - a murine pathogen that mimics human infections with attaching and effacing Escherichia coli. C. rodentium induced transcription of IL-6 and IL-22 in gut samples of mice and was associated with activation of the transcription factor Stat3 in intestinal epithelial cells. C. rodentium infection induced expression of several antimicrobial peptides such as RegIIIγ and Pla2g2a in the intestine which was critically dependent on Stat3 activation. Consequently, mice with specific deletion of Stat3 in intestinal epithelial cells showed increased susceptibility to C. rodentium infection as indicated by high bacterial load, severe gut inflammation, pronounced intestinal epithelial cell death and dissemination of bacteria to distant organs. Together, our data implicate an essential role for Stat3 activation in intestinal epithelial cells during C. rodentium infection. Stat3 concerts the host response to bacterial infection by controlling bacterial growth and suppression of apoptosis to maintain intestinal epithelial barrier function.

  6. Auricular Tissue Engineering Using Osteogenic Differentiation of Adipose Stem Cells with Small Intestine Submucosa.

    Science.gov (United States)

    Lin, Chih-Hsun; Yang, I-Chen; Tsai, Chi-Han; Fang, Hsu-Wei; Ma, Hsu

    2017-08-01

    Ear reconstruction remains a challenge for plastic surgeons. A tissue-engineering approach could provide another route for obtaining shape maintenance in neoauricular tissue. The authors designed a novel tissue-engineering auricular construct by culturing human adipose stem cells, which differentiated into osteocytes but not chondrocytes, in small intestine submucosa scaffolds. The authors evaluated cell growth potential and mechanical properties. An ear-shaped construct was created in vitro and then implanted in the backs of nude mice. The histology, cellularity, neovascularization, mechanical properties, and ear shape maintenance were investigated. In vitro, human adipose stem cells could be successfully seeded in the small intestine submucosa and differentiated toward osteogenesis. The ear-shaped human adipose stem cell/small intestine submucosa construct could maintain its shape in vivo up to 1 year. Alizarin Red S staining confirmed osteogenic differentiation. CD31 stain showed prominent angiogenesis in the human adipose stem cell/small intestine submucosa construct at 6 months and persistence up to 1 year. h-MHC stain revealed the maintenance of cellularity at 6 months and persistence up to 1 year. The mechanical properties were similar to those of native ear cartilage. The authors' study found that the combination of human adipose stem cells and small intestine submucosa could provide a more durable ear-shaped construct in vivo. The mechanical properties, shape, and cellularity were maintained in the constructs for up to 12 months. Therapeutic, V.

  7. Evaluation of small intestine grafts decellularization methods for corneal tissue engineering.

    Directory of Open Access Journals (Sweden)

    Ana Celeste Oliveira

    Full Text Available Advances in the development of cornea substitutes by tissue engineering techniques have focused on the use of decellularized tissue scaffolds. In this work, we evaluated different chemical and physical decellularization methods on small intestine tissues to determine the most appropriate decellularization protocols for corneal applications. Our results revealed that the most efficient decellularization agents were the SDS and triton X-100 detergents, which were able to efficiently remove most cell nuclei and residual DNA. Histological and histochemical analyses revealed that collagen fibers were preserved upon decellularization with triton X-100, NaCl and sonication, whereas reticular fibers were properly preserved by decellularization with UV exposure. Extracellular matrix glycoproteins were preserved after decellularization with SDS, triton X-100 and sonication, whereas proteoglycans were not affected by any of the decellularization protocols. Tissue transparency was significantly higher than control non-decellularized tissues for all protocols, although the best light transmittance results were found in tissues decellularized with SDS and triton X-100. In conclusion, our results suggest that decellularized intestinal grafts could be used as biological scaffolds for cornea tissue engineering. Decellularization with triton X-100 was able to efficiently remove all cells from the tissues while preserving tissue structure and most fibrillar and non-fibrillar extracellular matrix components, suggesting that this specific decellularization agent could be safely used for efficient decellularization of SI tissues for cornea TE applications.

  8. NOD-Like Receptors in Intestinal Homeostasis and Epithelial Tissue Repair

    Science.gov (United States)

    Parlato, Marianna; Yeretssian, Garabet

    2014-01-01

    The intestinal epithelium constitutes a dynamic physical barrier segregating the luminal content from the underlying mucosal tissue. Following injury, the epithelial integrity is restored by rapid migration of intestinal epithelial cells (IECs) across the denuded area in a process known as wound healing. Hence, through a sequence of events involving restitution, proliferation and differentiation of IECs the gap is resealed and homeostasis reestablished. Relapsing damage followed by healing of the inflamed mucosa is a hallmark of several intestinal disorders including inflammatory bowel diseases (IBD). While several regulatory peptides, growth factors and cytokines stimulate restitution of the epithelial layer after injury, recent evidence in the field underscores the contribution of innate immunity in controlling this process. In particular, nucleotide-binding and oligomerization domain-like receptors (NLRs) play critical roles in sensing the commensal microbiota, maintaining homeostasis, and regulating intestinal inflammation. Here, we review the process of intestinal epithelial tissue repair and we specifically focus on the impact of NLR-mediated signaling mechanisms involved in governing epithelial wound healing during disease. PMID:24886810

  9. Tissue distribution of aryl hydrocarbon receptor in the intestine: Implication of putative roles in tumor suppression

    International Nuclear Information System (INIS)

    Ikuta, Togo; Kurosumi, Masafumi; Yatsuoka, Toshimasa; Nishimura, Yoji

    2016-01-01

    Intestinal homeostasis is maintained by complex interactions between intestinal microorganisms and the gut immune system. Dysregulation of gut immunity may lead to inflammatory disorders and tumorigenesis. We previously have shown the tumor suppressive effects of aryl hydrocarbon receptor (AhR) in intestinal carcinogenesis. In the present study, we investigated AhR distribution in the mouse and human intestine by histochemical analysis. In the normal intestine, AhR was mainly localized in the stroma containing immune cells in the lamina propria and lymphoid follicles. On the other hand, in the tumor tissue from human colon cancer and that developed in Apc Min/+ mice, AhR expression was elevated. AhR immunostaining was found in both stromal and tumor cells. Although AhR was localized in the cytoplasm of tumor cells in most cases, nuclear AhR was also observed in some. AhR knockdown using siRNA resulted in significant promotion of cell growth in colon cancer cell lines. Furthermore, AhR activation by AhR ligands supplemented in culture medium suppressed cell growth. Our study results suggest that tumor suppressive roles of AhR are estimated in two distinct ways: in normal tissue, AhR is associated with tumor prevention by regulating gut immunity, whereas in tumor cells, it is involved in growth suppression. - Highlights: • In the normal intestine, AhR was mainly localized in stroma containing immune cells. • In the tumor tissue, AhR expression was found in both stromal and tumor cells. • AhR knockdown promoted cell growth in colon cancer cell lines.

  10. Tissue distribution of aryl hydrocarbon receptor in the intestine: Implication of putative roles in tumor suppression

    Energy Technology Data Exchange (ETDEWEB)

    Ikuta, Togo, E-mail: togo@cancer-c.pref.saitama.jp [Department of Cancer Prevention, Research Institute for Clinical Oncology, Saitama Cancer Center, 818 Komuro, Ina-machi, Kitaadachi-gun, Saitama 362-0806 (Japan); Kurosumi, Masafumi, E-mail: mkurosumi@cancer-c.pref.saitama.jp [Division of Pathology, Saitama Cancer Center, 780 Komuro, Ina-machi, Kitaadachi-gun, Saitama 362-0806 (Japan); Yatsuoka, Toshimasa, E-mail: yatsuoka-gi@umin.ac.jp [Division of Gastroenterological Surgery, Saitama Cancer Center, 780 Komuro, Ina-machi, Kitaadachi-gun, Saitama 362-0806 (Japan); Nishimura, Yoji, E-mail: yojinish@cancr-c.pref.saitama.jp [Division of Gastroenterological Surgery, Saitama Cancer Center, 780 Komuro, Ina-machi, Kitaadachi-gun, Saitama 362-0806 (Japan)

    2016-05-01

    Intestinal homeostasis is maintained by complex interactions between intestinal microorganisms and the gut immune system. Dysregulation of gut immunity may lead to inflammatory disorders and tumorigenesis. We previously have shown the tumor suppressive effects of aryl hydrocarbon receptor (AhR) in intestinal carcinogenesis. In the present study, we investigated AhR distribution in the mouse and human intestine by histochemical analysis. In the normal intestine, AhR was mainly localized in the stroma containing immune cells in the lamina propria and lymphoid follicles. On the other hand, in the tumor tissue from human colon cancer and that developed in Apc{sup Min/+}mice, AhR expression was elevated. AhR immunostaining was found in both stromal and tumor cells. Although AhR was localized in the cytoplasm of tumor cells in most cases, nuclear AhR was also observed in some. AhR knockdown using siRNA resulted in significant promotion of cell growth in colon cancer cell lines. Furthermore, AhR activation by AhR ligands supplemented in culture medium suppressed cell growth. Our study results suggest that tumor suppressive roles of AhR are estimated in two distinct ways: in normal tissue, AhR is associated with tumor prevention by regulating gut immunity, whereas in tumor cells, it is involved in growth suppression. - Highlights: • In the normal intestine, AhR was mainly localized in stroma containing immune cells. • In the tumor tissue, AhR expression was found in both stromal and tumor cells. • AhR knockdown promoted cell growth in colon cancer cell lines.

  11. Development of Chitosan Scaffolds with Enhanced Mechanical Properties for Intestinal Tissue Engineering Applications.

    Science.gov (United States)

    Zakhem, Elie; Bitar, Khalil N

    2015-10-13

    Massive resections of segments of the gastrointestinal (GI) tract lead to intestinal discontinuity. Functional tubular replacements are needed. Different scaffolds were designed for intestinal tissue engineering application. However, none of the studies have evaluated the mechanical properties of the scaffolds. We have previously shown the biocompatibility of chitosan as a natural material in intestinal tissue engineering. Our scaffolds demonstrated weak mechanical properties. In this study, we enhanced the mechanical strength of the scaffolds with the use of chitosan fibers. Chitosan fibers were circumferentially-aligned around the tubular chitosan scaffolds either from the luminal side or from the outer side or both. Tensile strength, tensile strain, and Young's modulus were significantly increased in the scaffolds with fibers when compared with scaffolds without fibers. Burst pressure was also increased. The biocompatibility of the scaffolds was maintained as demonstrated by the adhesion of smooth muscle cells around the different kinds of scaffolds. The chitosan scaffolds with fibers provided a better candidate for intestinal tissue engineering. The novelty of this study was in the design of the fibers in a specific alignment and their incorporation within the scaffolds.

  12. Transcriptomic Analysis of Intestinal Tissues from Two 90-Day Feeding Studies in Rats Using Genetically Modified MON810 Maize Varieties

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    Jutta Sharbati

    2017-12-01

    Full Text Available Background: Global as well as specific expression profiles of selected rat tissues were characterized to assess the safety of genetically modified (GM maize MON810 containing the insecticidal protein Cry1Ab. Gene expression was evaluated by use of Next Generation Sequencing (NGS as well as RT-qPCR within rat intestinal tissues based on mandatory 90-day rodent feeding studies. In parallel to two 90-day feeding studies, the transcriptional response of rat tissues was assessed as another endpoint to enhance the mechanistic interpretation of GM feeding studies and/or to facilitate the generation of a targeted hypothesis. Rats received diets containing 33% GM maize (MON810 or near-isogenic control maize. As a site of massive exposure to ingested feed the transcriptomic response of ileal and colonic tissue was profiled via RT-qPCR arrays targeting apoptosis, DNA-damage/repair, unfolded protein response (UPR. For global RNA profiling of rat ileal tissue, we applied NGS.Results: No biological response to the GM-diet was observed in male and in female rat tissues. Transcriptome wide analysis of gene expression by RNA-seq confirmed these findings. Nevertheless, gene ontology (GO analysis clearly associated a set of distinctly regulated transcripts with circadian rhythms. We confirmed differential expression of circadian clock genes using RT-qPCR and immunoassays for selected factors, thereby indicating physiological effects caused by the time point of sampling.Conclusion: Prediction of potential unintended effects of GM-food/feed by transcriptome based profiling of intestinal tissue presents a novel approach to complement classical toxicological testing procedures. Including the detection of alterations in signaling pathways in toxicity testing procedures may enhance the confidence in outcomes of toxicological trials. In this study, no significant GM-related changes in intestinal expression profiles were found in rats fed GM-maize MON810. Relevant

  13. Transcriptomic Analysis of Intestinal Tissues from Two 90-Day Feeding Studies in Rats Using Genetically Modified MON810 Maize Varieties.

    Science.gov (United States)

    Sharbati, Jutta; Bohmer, Marc; Bohmer, Nils; Keller, Andreas; Backes, Christina; Franke, Andre; Steinberg, Pablo; Zeljenková, Dagmar; Einspanier, Ralf

    2017-01-01

    Background: Global as well as specific expression profiles of selected rat tissues were characterized to assess the safety of genetically modified (GM) maize MON810 containing the insecticidal protein Cry1Ab. Gene expression was evaluated by use of Next Generation Sequencing (NGS) as well as RT-qPCR within rat intestinal tissues based on mandatory 90-day rodent feeding studies. In parallel to two 90-day feeding studies, the transcriptional response of rat tissues was assessed as another endpoint to enhance the mechanistic interpretation of GM feeding studies and/or to facilitate the generation of a targeted hypothesis. Rats received diets containing 33% GM maize (MON810) or near-isogenic control maize. As a site of massive exposure to ingested feed the transcriptomic response of ileal and colonic tissue was profiled via RT-qPCR arrays targeting apoptosis, DNA-damage/repair, unfolded protein response (UPR). For global RNA profiling of rat ileal tissue, we applied NGS. Results: No biological response to the GM-diet was observed in male and in female rat tissues. Transcriptome wide analysis of gene expression by RNA-seq confirmed these findings. Nevertheless, gene ontology (GO) analysis clearly associated a set of distinctly regulated transcripts with circadian rhythms. We confirmed differential expression of circadian clock genes using RT-qPCR and immunoassays for selected factors, thereby indicating physiological effects caused by the time point of sampling. Conclusion: Prediction of potential unintended effects of GM-food/feed by transcriptome based profiling of intestinal tissue presents a novel approach to complement classical toxicological testing procedures. Including the detection of alterations in signaling pathways in toxicity testing procedures may enhance the confidence in outcomes of toxicological trials. In this study, no significant GM-related changes in intestinal expression profiles were found in rats fed GM-maize MON810. Relevant alterations of

  14. Mdm4 loss in the intestinal epithelium leads to compartmentalized cell death but no tissue abnormalities

    Science.gov (United States)

    Valentin-Vega, Yasmine A.; Box, Neil; Terzian, Tamara; Lozano, Guillermina

    2014-01-01

    Mdm4 is a critical inhibitor of the p53 tumor suppressor. Mdm4 null mice die early during embryogenesis due to increased p53 activity. In this study, we explore the role that Mdm4 plays in the intestinal epithelium by crossing mice carrying the Mdm4 floxed allele to mice with the Villin Cre transgene. Our data show that loss of Mdm4 (Mdm4intΔ) in this tissue resulted in viable animals with no obvious morphological abnormalities. However, these mutants displayed increased p53 levels and apoptosis exclusively in the proliferative compartment of the intestinal epithelium. This phenotype was completely rescued in a p53 null background. Notably, the observed compartmentalized apoptosis in proliferative intestinal epithelial cells was not due to restricted Mdm4 expression in this region. Thus, in this specific cellular context, p53 is negatively regulated by Mdm4 exclusively in highly proliferative cells. PMID:19371999

  15. LPS-induced genes in intestinal tissue of the sea cucumber Holothuria glaberrima.

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    Francisco Ramírez-Gómez

    2009-07-01

    Full Text Available Metazoan immunity is mainly associated with specialized cells that are directly involved with the immune response. Nevertheless, both in vertebrates and invertebrates other organs might respond to immune activation and participate either directly or indirectly in the ongoing immune process. However, most of what is known about invertebrate immunity has been restricted to immune effector cells and little information is available on the immune responses of other tissues or organs. We now focus on the immune reactions of the intestinal tissue of an echinoderm. Our study employs a non-conventional model, the echinoderm Holothuria glaberrima, to identify intestinal molecules expressed after an immune challenge presented by an intra-coelomic injection of lipopolysaccharides (LPS. The expression profiles of intestinal genes expressed differentially between LPS-injected animals and control sea water-injected animals were determined using a custom-made Agilent microarray with 7209 sea cucumber intestinal ESTs. Fifty (50 unique sequences were found to be differentially expressed in the intestine of LPS-treated sea cucumbers. Seven (7 of these sequences represented homologues of known proteins, while the remaining (43 had no significant similarity with any protein, EST or RNA database. The known sequences corresponded to cytoskeletal proteins (Actin and alpha-actinin, metabolic enzymes (GAPDH, Ahcy and Gnmt, metal ion transport/metabolism (major yolk protein and defense/recognition (fibrinogen-like protein. The expression pattern of 11 genes was validated using semi-quantitative RT-PCR. Nine of these corroborated the microarray results and the remaining two showed a similar trend but without statistical significance. Our results show some of the molecular events by which the holothurian intestine responds to an immune challenge and provide important information to the study of the evolution of the immune response.

  16. Stomach, intestine, and colon tissue discriminators for wireless capsule endoscopy images

    Science.gov (United States)

    Berens, Jeff; Mackiewicz, Michal; Bell, Duncan

    2005-04-01

    Wireless Capsule Endoscopy (WCE) is a new colour imaging technology that enables close examination of the interior of the entire small intestine. Typically, the WCE operates for ~8 hours and captures ~40,000 useful images. The images are viewed as a video sequence, which generally takes a doctor over an hour to analyse. In order to activate certain key features of the software provided with the capsule, it is necessary to locate and annotate the boundaries between certain gastrointestinal (GI) tract regions (stomach, intestine and colon) in the footage. In this paper we propose a method of automatically discriminating stomach, intestine and colon tissue in order to significantly reduce the video assessment time. We use hue saturation chromaticity histograms which are compressed using a hybrid transform, incorporating the Discrete Cosine Transform (DCT) and Principal Component Analysis (PCA). The performance of two classifiers is compared: k-nearest neighbour (kNN) and Support Vector Classifier (SVC). After training the classifier, we applied a narrowing step algorithm to converge to the points in the video where the capsule firstly passes through the pylorus (the valve between the stomach and the intestine) and later the ileocaecal valve (IV, the valve between the intestine and colon). We present experimental results that demonstrate the effectiveness of this method.

  17. Human Intestinal Tissue with Adult Stem Cell Properties Derived from Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Ryan Forster

    2014-06-01

    Full Text Available Genetically engineered human pluripotent stem cells (hPSCs have been proposed as a source for transplantation therapies and are rapidly becoming valuable tools for human disease modeling. However, many applications are limited due to the lack of robust differentiation paradigms that allow for the isolation of defined functional tissues. Here, using an endogenous LGR5-GFP reporter, we derived adult stem cells from hPSCs that gave rise to functional human intestinal tissue comprising all major cell types of the intestine. Histological and functional analyses revealed that such human organoid cultures could be derived with high purity and with a composition and morphology similar to those of cultures obtained from human biopsies. Importantly, hPSC-derived organoids responded to the canonical signaling pathways that control self-renewal and differentiation in the adult human intestinal stem cell compartment. This adult stem cell system provides a platform for studying human intestinal disease in vitro using genetically engineered hPSCs.

  18. Gut-associated lymphoid tissue, T cell trafficking, and chronic intestinal inflammation.

    Science.gov (United States)

    Koboziev, Iurii; Karlsson, Fridrik; Grisham, Matthew B

    2010-10-01

    The etiologies of the inflammatory bowel diseases (IBD; Crohn's disease, ulcerative colitis) have not been fully elucidated. However, there is very good evidence implicating T cell and T cell trafficking to the gut and its associated lymphoid tissue as important components in disease pathogenesis. The objective of this review is to provide an overview of the mechanisms involved in naive and effector T cell trafficking to the gut-associated lymphoid tissue (GALT; Peyer's patches, isolated lymphoid follicles), mesenteric lymph nodes and intestine in response to commensal enteric antigens under physiological conditions as well as during the induction of chronic gut inflammation. In addition, recent data suggests that the GALT may not be required for enteric antigen-driven intestinal inflammation in certain mouse models of IBD. These new data suggest a possible paradigm shift in our understanding of how and where naive T cells become activated to yield disease-producing effector cells. © 2010 New York Academy of Sciences.

  19. The Gut-Associated Lymphoid Tissues in the Small Intestine, Not the Large Intestine, Play a Major Role in Oral Prion Disease Pathogenesis.

    Science.gov (United States)

    Donaldson, David S; Else, Kathryn J; Mabbott, Neil A

    2015-09-01

    Prion diseases are infectious neurodegenerative disorders characterized by accumulations of abnormally folded cellular prion protein in affected tissues. Many natural prion diseases are acquired orally, and following exposure, the early replication of some prion isolates upon follicular dendritic cells (FDC) within gut-associated lymphoid tissues (GALT) is important for the efficient spread of disease to the brain (neuroinvasion). Prion detection within large intestinal GALT biopsy specimens has been used to estimate human and animal disease prevalence. However, the relative contributions of the small and large intestinal GALT to oral prion pathogenesis were unknown. To address this issue, we created mice that specifically lacked FDC-containing GALT only in the small intestine. Our data show that oral prion disease susceptibility was dramatically reduced in mice lacking small intestinal GALT. Although these mice had FDC-containing GALT throughout their large intestines, these tissues were not early sites of prion accumulation or neuroinvasion. We also determined whether pathology specifically within the large intestine might influence prion pathogenesis. Congruent infection with the nematode parasite Trichuris muris in the large intestine around the time of oral prion exposure did not affect disease pathogenesis. Together, these data demonstrate that the small intestinal GALT are the major early sites of prion accumulation and neuroinvasion after oral exposure. This has important implications for our understanding of the factors that influence the risk of infection and the preclinical diagnosis of disease. Many natural prion diseases are acquired orally. After exposure, the accumulation of some prion diseases in the gut-associated lymphoid tissues (GALT) is important for efficient spread of disease to the brain. However, the relative contributions of GALT in the small and large intestines to oral prion pathogenesis were unknown. We show that the small intestinal

  20. Intestinal tissue transglutaminase in coeliac disease of children and adults: ultrastructural localization and variation in expression.

    Science.gov (United States)

    Skovbjerg, H; Hansen, G H; Niels-Christiansen, L L; Anthonsen, D; Ascher, H; Midhagen, G; Hallert, C; Norén, O; Sjöström, H

    2004-12-01

    Tissue transglutaminase is the main antigen for the anti-endomysial antibodies used for diagnosis of coeliac disease and can with some specificity in vitro deamidate gliadins generating potent epitopes. The intestinal levels and the ultrastructural localization of tissue transglutaminase in normal and affected persons were investigated to provide further information on its role in this disease. Intestinal biopsies were taken from normal and coeliac children and adults. The level of transglutaminase was analysed by means of a quantitative enzymatic assay and its ultrastructural localization by immunogold electronmicroscopy using a monoclonal antibody against tissue transglutaminase. In relation to normal individuals, the enzymatic activity of tissue transglutaminase in adult coeliac patients was increased. The enzyme was found in the enterocytes and in increased amount just beneath the enterocytes, where cytosolic and nuclear labelling of distinct elongated cells was seen in addition to extracellular labelling close to collagen fibrils. In children, the enzymatic activity and the immunogold labelling could not be shown to be related to disease. In all cases the enzyme activity was EDTA-sensitive. The increased amount of tissue transglutaminase activity in coeliac adults was shown to be due to the appearance of the enzyme in enterocytes and increased expression in the lamina propria. No evidence was found to support the idea of a changed localization or changed amounts as primary elements in coeliac disease pathogenesis, nor for the involvement of non-calcium dependent microbial transglutaminases.

  1. Zearalenone in the Intestinal Tissues of Immature Gilts Exposed per os to Mycotoxins

    Science.gov (United States)

    Zielonka, Łukasz; Waśkiewicz, Agnieszka; Beszterda, Monika; Kostecki, Marian; Dąbrowski, Michał; Obremski, Kazimierz; Goliński, Piotr; Gajęcki, Maciej

    2015-01-01

    Zearalenone and its metabolites, α-zearalenol and β-zearalenol, demonstrate estradiol-like activity and disrupt physiological functions in animals. This article evaluates the carryover of zearalenone and its selected metabolites from the digesta to intestinal walls (along the entire intestines) in pre-pubertal gilts exposed to low doses of zearalenone over long periods of time. The term “carryover” describes the transfer of mycotoxins from feed to edible tissues, and it was used to assess the risk of mycotoxin exposure for consumers. The experimental gilts with body weight of up to 25 kg were per os administered zearalenone at a daily dose of 40 μg/kg BW (Group E, n = 18) or placebo (Group C, n = 21) over a period of 42 days. In the first weeks of exposure, the highest values of the carryover factor were noted in the duodenum and the jejunum. In animals receiving pure zearalenone, the presence of metabolites was not determined in intestinal tissues. In the last three weeks of the experiment, very high values of the carryover factor were observed in the duodenum and the descending colon. The results of the study indicate that in animals exposed to subclinical doses of zearalenone, the carryover factor could be determined by the distribution and expression of estrogen receptor beta. PMID:26295259

  2. Zearalenone in the Intestinal Tissues of Immature Gilts Exposed per os to Mycotoxins

    Directory of Open Access Journals (Sweden)

    Łukasz Zielonka

    2015-08-01

    Full Text Available Zearalenone and its metabolites, α-zearalenol and β-zearalenol, demonstrate estradiol-like activity and disrupt physiological functions in animals. This article evaluates the carryover of zearalenone and its selected metabolites from the digesta to intestinal walls (along the entire intestines in pre-pubertal gilts exposed to low doses of zearalenone over long periods of time. The term “carryover” describes the transfer of mycotoxins from feed to edible tissues, and it was used to assess the risk of mycotoxin exposure for consumers. The experimental gilts with body weight of up to 25 kg were per os administered zearalenone at a daily dose of 40 μg/kg BW (Group E, n = 18 or placebo (Group C, n = 21 over a period of 42 days. In the first weeks of exposure, the highest values of the carryover factor were noted in the duodenum and the jejunum. In animals receiving pure zearalenone, the presence of metabolites was not determined in intestinal tissues. In the last three weeks of the experiment, very high values of the carryover factor were observed in the duodenum and the descending colon. The results of the study indicate that in animals exposed to subclinical doses of zearalenone, the carryover factor could be determined by the distribution and expression of estrogen receptor beta.

  3. A 3D intestinal tissue model supports Clostridioides difficile germination, colonization, toxin production and epithelial damage.

    Science.gov (United States)

    Shaban, Lamyaa; Chen, Ying; Fasciano, Alyssa C; Lin, Yinan; Kaplan, David L; Kumamoto, Carol A; Mecsas, Joan

    2018-04-01

    Endospore-forming Clostridioides difficile is a causative agent of antibiotic-induced diarrhea, a major nosocomial infection. Studies of its interactions with mammalian tissues have been hampered by the fact that C. difficile requires anaerobic conditions to survive after spore germination. We recently developed a bioengineered 3D human intestinal tissue model and found that low O 2 conditions are produced in the lumen of these tissues. Here, we compared the ability of C. difficile spores to germinate, produce toxin and cause tissue damage in our bioengineered 3D tissue model versus in a 2D transwell model in which human cells form a polarized monolayer. 3D tissue models or 2D polarized monolayers on transwell filters were challenged with the non-toxin producing C. difficile CCUG 37787 serotype X (ATCC 43603) and the toxin producing UK1 C. difficile spores in the presence of the germinant, taurocholate. Spores germinated in both the 3D tissue model as well as the 2D transwell system, however toxin activity was significantly higher in the 3D tissue models compared to the 2D transwells. Moreover, the epithelium damage in the 3D tissue model was significantly more severe than in 2D transwells and damage correlated significantly with the level of toxin activity detected but not with the amount of germinated spores. Combined, these results show that the bioengineered 3D tissue model provides a powerful system with which to study early events leading to toxin production and tissue damage of C. difficile with mammalian cells under anaerobic conditions. Furthermore, these systems may be useful for examining the effects of microbiota, novel drugs and other potential therapeutics directed towards C. difficile infections. Copyright © 2018 Elsevier Ltd. All rights reserved.

  4. Ex vivo permeability experiments in excised rat intestinal tissue and in vitro solubility measurements in aspirated human intestinal fluids support age-dependent oral drug absorption.

    Science.gov (United States)

    Annaert, Pieter; Brouwers, Joachim; Bijnens, Ann; Lammert, Frank; Tack, Jan; Augustijns, Patrick

    2010-01-31

    The possible influence of advanced age on intestinal drug absorption was investigated by determining the effects of aging on (i) solubility of model drugs in human intestinal fluids (HIF) obtained from two age groups (18-25 years; 62-72 years); and (ii) transepithelial permeation of model drugs across intestinal tissue excised from young, adult and old rats. Average equilibrium solubility values for 10 poorly soluble compounds in HIF aspirated from both age groups showed high interindividual variability, but did not reveal significant differences. Characterization of the HIF from both age groups demonstrated comparable pH profiles, while concentrations of individual bile salts showed pronounced variability between individuals, however without statistical differences between age groups. Transepithelial permeation of the transcellular probe metoprolol was significantly increased in old rats (38 weeks) compared to the younger age groups, while the modulatory role of P-glycoprotein in transepithelial talinolol transport was observed in adult and old rats but not in young rats. In conclusion, age-dependent permeability of intestinal tissue (rather than age-dependent luminal drug solubility) may contribute to altered intestinal drug absorption in older patients compared to young adults. Copyright 2009 Elsevier B.V. All rights reserved.

  5. Depth of Bacterial Invasion in Resected Intestinal Tissue Predicts Mortality in Surgical Necrotizing Enterocolitis

    Science.gov (United States)

    Remon, Juan I.; Amin, Sachin C.; Mehendale, Sangeeta R.; Rao, Rakesh; Luciano, Angel A.; Garzon, Steven A.; Maheshwari, Akhil

    2015-01-01

    Objective Up to a third of all infants who develop necrotizing enterocolitis (NEC) require surgical resection of necrotic bowel. We hypothesized that the histopathological findings in surgically-resected bowel can predict the clinical outcome of these infants. Study design We reviewed the medical records and archived pathology specimens from all patients who underwent bowel resection/autopsy for NEC at a regional referral center over a 10-year period. Pathology specimens were graded for the depth and severity of necrosis, inflammation, bacteria invasion, and pneumatosis, and histopathological findings were correlated with clinical outcomes. Results We performed clinico-pathological analysis on 33 infants with confirmed NEC, of which 18 (54.5%) died. Depth of bacterial invasion in resected intestinal tissue predicted death from NEC (odds ratio 5.39 per unit change in the depth of bacterial invasion, 95% confidence interval 1.33-21.73). The presence of transmural necrosis and bacteria in the surgical margins of resected bowel was also associated with increased mortality. Conclusions Depth of bacterial invasion in resected intestinal tissue predicts mortality in surgical NEC. PMID:25950918

  6. In vitro enteroid-derived three-dimensional tissue model of human small intestinal epithelium with innate immune responses.

    Directory of Open Access Journals (Sweden)

    Ying Chen

    Full Text Available There is a need for functional in vitro 3D human intestine systems that can bridge the gap between conventional cell culture studies and human trials. The successful engineering in vitro of human intestinal tissues relies on the use of the appropriate cell sources, biomimetic scaffolds, and 3D culture conditions to support vital organ functions. We previously established a compartmentalized scaffold consisting of a hollow space within a porous bulk matrix, in which a functional and physiologically relevant intestinal epithelium system was generated using intestinal cell lines. In this study, we adopt the 3D scaffold system for the cultivation of stem cell-derived human small intestinal enteriods (HIEs to engineer an in vitro 3D model of a nonstransformed human small intestinal epithelium. Characterization of tissue properties revealed a mature HIE-derived epithelium displaying four major terminally differentiated epithelial cell types (enterocytes, Goblet cells, Paneth cells, enteroendocrine cells, with tight junction formation, microvilli polarization, digestive enzyme secretion, and low oxygen tension in the lumen. Moreover, the tissue model demonstrates significant antibacterial responses to E. coli infection, as evidenced by the significant upregulation of genes involved in the innate immune response. Importantly, many of these genes are activated in human patients with inflammatory bowel disease (IBD, implicating the potential application of the 3D stem-cell derived epithelium for the in vitro study of host-microbe-pathogen interplay and IBD pathogenesis.

  7. In vitro enteroid-derived three-dimensional tissue model of human small intestinal epithelium with innate immune responses.

    Science.gov (United States)

    Chen, Ying; Zhou, Wenda; Roh, Terrence; Estes, Mary K; Kaplan, David L

    2017-01-01

    There is a need for functional in vitro 3D human intestine systems that can bridge the gap between conventional cell culture studies and human trials. The successful engineering in vitro of human intestinal tissues relies on the use of the appropriate cell sources, biomimetic scaffolds, and 3D culture conditions to support vital organ functions. We previously established a compartmentalized scaffold consisting of a hollow space within a porous bulk matrix, in which a functional and physiologically relevant intestinal epithelium system was generated using intestinal cell lines. In this study, we adopt the 3D scaffold system for the cultivation of stem cell-derived human small intestinal enteriods (HIEs) to engineer an in vitro 3D model of a nonstransformed human small intestinal epithelium. Characterization of tissue properties revealed a mature HIE-derived epithelium displaying four major terminally differentiated epithelial cell types (enterocytes, Goblet cells, Paneth cells, enteroendocrine cells), with tight junction formation, microvilli polarization, digestive enzyme secretion, and low oxygen tension in the lumen. Moreover, the tissue model demonstrates significant antibacterial responses to E. coli infection, as evidenced by the significant upregulation of genes involved in the innate immune response. Importantly, many of these genes are activated in human patients with inflammatory bowel disease (IBD), implicating the potential application of the 3D stem-cell derived epithelium for the in vitro study of host-microbe-pathogen interplay and IBD pathogenesis.

  8. Fluoroscence in situ hybridization of chicken intestinal samples with bacterial rRNA targeted oligonucleotide probes

    DEFF Research Database (Denmark)

    Olsen, Katja Nyholm; Francesch, M.; Christensen, Henrik

    2006-01-01

    The objective was to develop a fast and accurate molecular method for the quantification of the intestinal flora in chickens by rRNA fluorescence in situ hybridization (FISH). Seven weeks old conventionally reared Lohmann hens were used to set up the method. To sample ileal intestinal content......, the distal part from Meckels diverticulum to the ileo-caecal junction was removed. Fixation was performed in ethanol and phosphate buffered saline. After washing by centrifugation, the sample was resuspended in pre-heated hybridization buffer with oligonucleotide probe labelled with Cy3 (10ng/µl). The cells...... were hybridized for 24-72h, centrifuged, washed with pre-heated hybridization buffer, centrifuged and resuspended in Millipore quality water before filtration onto a 0.22 µm black polycarbonate filter. The probes used in this study were, LGC354A, LGC354B, LGC354C, Strc493, Bacto1080, Sal3, Chis150, EUB...

  9. Preparation of tissue samples for X-ray fluorescence microscopy

    International Nuclear Information System (INIS)

    Chwiej, Joanna; Szczerbowska-Boruchowska, Magdalena; Lankosz, Marek; Wojcik, Slawomir; Falkenberg, Gerald; Stegowski, Zdzislaw; Setkowicz, Zuzanna

    2005-01-01

    As is well-known, trace elements, especially metals, play an important role in the pathogenesis of many disorders. The topographic and quantitative elemental analysis of pathologically changed tissues may shed some new light on processes leading to the degeneration of cells in the case of selected diseases. An ideal and powerful tool for such purpose is the Synchrotron Microbeam X-ray Fluorescence technique. It enables the carrying out of investigations of the elemental composition of tissues even at the single cell level. The tissue samples for histopathological investigations are routinely fixed and embedded in paraffin. The authors try to verify the usefulness of such prepared tissue sections for elemental analysis with the use of X-ray fluorescence microscopy. Studies were performed on rat brain samples. Changes in elemental composition caused by fixation in formalin or paraformaldehyde and embedding in paraffin were examined. Measurements were carried out at the bending magnet beamline L of the Hamburger Synchrotronstrahlungslabor HASYLAB in Hamburg. The decrease in mass per unit area of K, Br and the increase in P, S, Fe, Cu and Zn in the tissue were observed as a result of the fixation. For the samples embedded in paraffin, a lower level of most elements was observed. Additionally, for these samples, changes in the composition of some elements were not uniform for different analyzed areas of rat brain

  10. Small bowel tissue engineering using small intestinal submucosa as a scaffold.

    Science.gov (United States)

    Chen, M K; Badylak, S F

    2001-08-01

    Small intestinal submucosa (SIS) is an extracellular matrix used in tissue engineering studies to create de novo abdominal wall, urinary bladder, tendons, blood vessels, and dura mater. The purpose of this study is to evaluate the feasibility of using SIS as a scaffold for small bowel regeneration in an in situ xenograft model. Twenty-three dogs had a partial defect created on the small bowel wall which was repaired with a SIS patch. Four dogs underwent small bowel resection with placement of an interposed tube of SIS. The animals were followed 2 weeks to 1 year. Three of the 23 dogs with SIS placed as a patch died shortly after surgery due to leakage from the site. The other 20 dogs survived up to time of elective necropsy with no evidence of intestinal dysfunction. At necropsy, the bowel circumference in the patched area had no stenosis. Histological evaluation showed the presence of a mucosal epithelial layer, varying amount of smooth muscle, sheets of collagen, and a serosal covering. Architecturally, the layers were not well organized in the submucosal region. An abundance of inflammatory cells was present in the early postoperative period but receded with time. All 4 dogs with a tubular segment of SIS interposed had significant problems. One had partial obstruction at 1 month, and 3 died in the early postoperative period due to leakage. This preliminary study suggests that SIS patches can be used for small bowel regeneration. Tubular segmental replacement is not feasible at this time. Copyright 2001 Academic Press.

  11. Pentoxifylline and prostaglandin E1 action on ischemia and reperfusion of small intestine tissue in rats. An immunohistochemical study.

    Science.gov (United States)

    Brasileiro, José Lacerda; Ramalho, Rondon Tosta; Aydos, Ricardo Dutra; Silva, Iandara Schettert; Takita, Luis Carlos; Marks, Guido; Assis, Peterson Vieira de

    2015-02-01

    To investigate the action of pentoxifylline (PTX) and prostaglandin E1 (PGE1) on ischemia and reperfusion of small intestine tissue in rats, using immunohistochemical analysis. Thirty-five Wistar rats were distributed as follows: group A (n=10): subjected to intestinal ischemia and reperfusion for 60 min, with no drugs; group B (n=10): PTX given during tissue ischemia and reperfusion; group C (n=10): PGE1 given during tissue ischemia and reperfusion; group D (n=5): sham. A segment of the small intestine was excised from each euthanized animal and subjected to immunohistochemical examination. Mean number of cells expressing anti-FAS ligand in the crypts was highest in Group A (78.9 ± 17.3), followed by groups B (16.7 ± 2.8), C (11.3 ± 1.8), and D (2.5 ± 0.9), with very significant differences between groups (pprostaglandin E1 proved beneficial during tissue reperfusion. The immunohistochemical results demonstrated a decrease in apoptotic cells, while protecting other intestinal epithelium cells against death after reperfusion, allowing these cells to renew the epithelial tissue.

  12. Blood transfusion in preterm infants improves intestinal tissue oxygenation without alteration in blood flow.

    Science.gov (United States)

    Banerjee, J; Leung, T S; Aladangady, N

    2016-11-01

    The objective of the study was to investigate the splanchnic blood flow velocity and oximetry response to blood transfusion in preterm infants according to postnatal age. Preterm infants receiving blood transfusion were recruited to three groups: 1-7 (group 1; n = 20), 8-28 (group 2; n = 21) and ≥29 days of life (group 3; n = 18). Superior mesenteric artery (SMA) peak systolic (PSV) and diastolic velocities were measured 30-60 min pre- and post-transfusion using Doppler ultrasound scan. Splanchnic tissue haemoglobin index (sTHI), tissue oxygenation index (sTOI) and fractional tissue oxygen extraction (sFTOE) were measured from 15-20 min before to post-transfusion using near-infrared spectroscopy. The mean pretransfusion Hb in group 1, 2 and 3 was 11, 10 and 9 g/dl, respectively. The mean (SD) pretransfusion SMA PSV in group 1, 2 and 3 was 0·63 (0·32), 0·81 (0·33) and 0·97 (0·40) m/s, respectively, and this did not change significantly following transfusion. The mean (SD) pretransfusion sTOI in group 1, 2 and 3 was 36·7 (19·3), 44·6 (10·4) and 41·3 (10·4)%, respectively. The sTHI and sTOI increased (P transfusion in all groups. On multivariate analysis, changes in SMA PSV and sTOI following blood transfusion were not associated with PDA, feeding, pretransfusion Hb and mean blood pressure. Pretransfusion baseline splanchnic tissue oximetry and blood flow velocity varied with postnatal age. Blood transfusion improved intestinal tissue oxygenation without altering mesenteric blood flow velocity irrespective of postnatal ages. © 2016 International Society of Blood Transfusion.

  13. Murine Neonates Infected with Yersinia enterocolitica Develop Rapid and Robust Proinflammatory Responses in Intestinal Lymphoid Tissues

    Science.gov (United States)

    Siefker, David T.; Echeverry, Andrea; Brambilla, Roberta; Fukata, Masayuki; Schesser, Kurt

    2014-01-01

    Neonatal animals are generally very susceptible to infection with bacterial pathogens. However, we recently reported that neonatal mice are highly resistant to orogastric infection with Yersinia enterocolitica. Here, we show that proinflammatory responses greatly exceeding those in adults arise very rapidly in the mesenteric lymph nodes (MLN) of neonates. High-level induction of proinflammatory gene expression occurred in the neonatal MLN as early as 18 h postinfection. Marked innate phagocyte recruitment was subsequently detected at 24 h postinfection. Enzyme-linked immunosorbent spot assay (ELISPOT) analyses indicated that enhanced inflammation in neonatal MLN is contributed to, in part, by an increased frequency of proinflammatory cytokine-secreting cells. Moreover, both CD11b+ and CD11b− cell populations appeared to play a role in proinflammatory gene expression. The level of inflammation in neonatal MLN was also dependent on key bacterial components. Y. enterocolitica lacking the virulence plasmid failed to induce innate phagocyte recruitment. In contrast, tumor necrosis factor alpha (TNF-α) protein expression and neutrophil recruitment were strikingly higher in neonatal MLN after infection with a yopP-deficient strain than with wild-type Y. enterocolitica, whereas only modest increases occurred in adults. This hyperinflammatory response was associated with greater colonization of the spleen and higher mortality in neonates, while there was no difference in mortality among adults. This model highlights the dynamic levels of inflammation in the intestinal lymphoid tissues and reveals the protective (wild-type strain) versus harmful (yopP-deficient strain) consequences of inflammation in neonates. Moreover, these results reveal that the neonatal intestinal lymphoid tissues have great potential to rapidly mobilize innate components in response to infection with bacterial enteropathogens. PMID:24478090

  14. Intestinal tissues induce an SNP mutation in Pseudomonas aeruginosa that enhances its virulence: possible role in anastomotic leak.

    Directory of Open Access Journals (Sweden)

    Andrea D Olivas

    Full Text Available The most feared complication following intestinal resection is anastomotic leakage. In high risk areas (esophagus/rectum where neoadjuvant chemoradiation is used, the incidence of anastomotic leaks remains unacceptably high (≈ 10% even when performed by specialist surgeons in high volume centers. The aims of this study were to test the hypothesis that anastomotic leakage develops when pathogens colonizing anastomotic sites become in vivo transformed to express a tissue destroying phenotype. We developed a novel model of anastomotic leak in which rats were exposed to pre-operative radiation as in cancer surgery, underwent distal colon resection and then were intestinally inoculated with Pseudomonas aeruginosa, a common colonizer of the radiated intestine. Results demonstrated that intestinal tissues exposed to preoperative radiation developed a significant incidence of anastomotic leak (>60%; p<0.01 when colonized by P. aeruginosa compared to radiated tissues alone (0%. Phenotype analysis comparing the original inoculating strain (MPAO1- termed P1 and the strain retrieved from leaking anastomotic tissues (termed P2 demonstrated that P2 was altered in pyocyanin production and displayed enhanced collagenase activity, high swarming motility, and a destructive phenotype against cultured intestinal epithelial cells (i.e. apoptosis, barrier function, cytolysis. Comparative genotype analysis between P1 and P2 revealed a single nucleotide polymorphism (SNP mutation in the mexT gene that led to a stop codon resulting in a non-functional truncated protein. Replacement of the mutated mexT gene in P2 with mexT from the original parental strain P1 led to reversion of P2 to the P1 phenotype. No spontaneous transformation was detected during 20 passages in TSB media. Use of a novel virulence suppressing compound PEG/Pi prevented P. aeruginosa transformation to the tissue destructive phenotype and prevented anastomotic leak in rats. This work demonstrates that

  15. Intestinal tissues induce an SNP mutation in Pseudomonas aeruginosa that enhances its virulence: possible role in anastomotic leak.

    Science.gov (United States)

    Olivas, Andrea D; Shogan, Benjamin D; Valuckaite, Vesta; Zaborin, Alexander; Belogortseva, Natalya; Musch, Mark; Meyer, Folker; Trimble, William L; An, Gary; Gilbert, Jack; Zaborina, Olga; Alverdy, John C

    2012-01-01

    The most feared complication following intestinal resection is anastomotic leakage. In high risk areas (esophagus/rectum) where neoadjuvant chemoradiation is used, the incidence of anastomotic leaks remains unacceptably high (≈ 10%) even when performed by specialist surgeons in high volume centers. The aims of this study were to test the hypothesis that anastomotic leakage develops when pathogens colonizing anastomotic sites become in vivo transformed to express a tissue destroying phenotype. We developed a novel model of anastomotic leak in which rats were exposed to pre-operative radiation as in cancer surgery, underwent distal colon resection and then were intestinally inoculated with Pseudomonas aeruginosa, a common colonizer of the radiated intestine. Results demonstrated that intestinal tissues exposed to preoperative radiation developed a significant incidence of anastomotic leak (>60%; pSNP) mutation in the mexT gene that led to a stop codon resulting in a non-functional truncated protein. Replacement of the mutated mexT gene in P2 with mexT from the original parental strain P1 led to reversion of P2 to the P1 phenotype. No spontaneous transformation was detected during 20 passages in TSB media. Use of a novel virulence suppressing compound PEG/Pi prevented P. aeruginosa transformation to the tissue destructive phenotype and prevented anastomotic leak in rats. This work demonstrates that in vivo transformation of microbial pathogens to a tissue destroying phenotype may have important implications in the pathogenesis of anastomotic leak.

  16. Colorectal tissue engineering: A comparative study between porcine small intestinal submucosa (SIS) and chitosan hydrogel patches.

    Science.gov (United States)

    Denost, Quentin; Adam, Jean-Philippe; Pontallier, Arnaud; Montembault, Alexandra; Bareille, Reine; Siadous, Robin; Delmond, Samantha; Rullier, Eric; David, Laurent; Bordenave, Laurence

    2015-12-01

    Tissue engineering may provide new operative tools for colorectal surgery in elective indications. The aim of this study was to define a suitable bioscaffold for colorectal tissue engineering. We compared 2 bioscaffolds with in vitro and in vivo experiments: porcine small intestinal submucosa (SIS) versus chitosan hydrogel matrix. We assessed nontoxicity of the scaffold in vitro by using human adipose-derived stem cells (hADSC). In vivo, a 1 × 2-cm colonic wall defect was created in 16 rabbits. Animals were divided randomly into 2 groups according to the graft used, SIS or chitosan hydrogel. Graft area was explanted at 4 and 8 weeks. The end points of in vivo experiments were technical feasibility, behavior of the scaffold, in situ putative inflammatory effect, and the quality of tissue regeneration, in particular smooth muscle layer regeneration. In vitro, hADSC attachment and proliferation occurred on both scaffolds without a substantial difference. After proliferation, hADSCs kept their mesenchymal stem cell characteristics. In vivo, one animal died in each group. Eight weeks after implantation, the chitosan scaffold allowed better wound healing compared with the SIS scaffold, with more effective control of inflammatory activity and an integral regeneration of the colonic wall including the smooth muscle cell layer. The outcomes of in vitro experiments did not differ greatly between the 2 groups. Macroscopic and histologic findings, however, revealed better wound healing of the colonic wall in the chitosan group suggesting that the chitosan hydrogel could serve as a better scaffold for colorectal tissue engineering. Copyright © 2015 Elsevier Inc. All rights reserved.

  17. Influence of mesenchymal stem cells on stomach tissue engineering using small intestinal submucosa.

    Science.gov (United States)

    Nakatsu, Hiroki; Ueno, Tomio; Oga, Atsunori; Nakao, Mitsuhiro; Nishimura, Taku; Kobayashi, Sei; Oka, Masaaki

    2015-03-01

    Small intestinal submucosa (SIS) is a biodegradable collagen-rich matrix containing functional growth factors. We have previously reported encouraging outcomes for regeneration of an artificial defect in the rodent stomach using SIS grafts, although the muscular layer was diminutive. In this study, we investigated the feasibility of SIS in conjunction with mesenchymal stem cells (MSCs) for regeneration of the gastrointestinal tract. MSCs from the bone marrow of green fluorescence protein (GFP)-transgenic Sprague-Dawley (SD) rats were isolated and expanded ex vivo. A 1 cm whole-layer stomach defect in SD rats was repaired using: a plain SIS graft without MSCs (group 1, control); a plain SIS graft followed by intravenous injection of MSCs (group 2); a SIS graft co-cultured with MSCs (group 3); or a SIS sandwich containing an MSC sheet (group 4). Pharmacological, electrophysiological and immunohistochemical examination was performed to evaluate the regenerated stomach tissue. Contractility in response to a muscarinic receptor agonist, a nitric oxide precursor or electrical field stimulation was observed in all groups. SIS grafts seeded with MSCs (groups 3 and 4) appeared to support improved regeneration compared with SIS grafts not seeded with MSCs (groups 1 and 2), by enabling the development of well-structured smooth muscle layers of significantly increased length. GFP expression was detected in the regenerated interstitial tissue, with fibroblast-like cells in the seeded-SIS groups. SIS potently induced pharmacological and electrophysiological regeneration of the digestive tract, and seeded MSCs provided an enriched environment that supported tissue regeneration by the SIS graft in the engineered stomach. © 2013 The Authors. Journal of Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

  18. Type 3 innate lymphoid cells maintain intestinal epithelial stem cells after tissue damage

    NARCIS (Netherlands)

    P. Aparicio-Domingo (Patricia); M. Romera-Hernandez (Monica); J.J. Karrich (Julien J.); F.H.J. Cornelissen (Ferry); N. Papazian (Natalie); D.J. Lindenbergh-Kortleve (Dicky); J.A. Butler (James A.); L. Boon (Louis); M. Coles (Mark); J.N. Samsom (Janneke); T. Cupedo (Tom)

    2015-01-01

    textabstractDisruption of the intestinal epithelial barrier allows bacterial translocation and predisposes to destructive inflammation. To ensure proper barrier composition, crypt-residing stem cells continuously proliferate and replenish all intestinal epithelial cells within days. As a consequence

  19. Fluoroscence in situ hybridization of chicken intestinal samples with bacterial rRNA targeted oligonucleotide probes

    DEFF Research Database (Denmark)

    Olsen, Katja Nyholm; Francesch, M.; Christensen, Henrik

    2006-01-01

    The objective was to develop a fast and accurate molecular method for the quantification of the intestinal flora in chickens by rRNA fluorescence in situ hybridization (FISH). Seven weeks old conventionally reared Lohmann hens were used to set up the method. To sample ileal intestinal content...... were hybridized for 24-72h, centrifuged, washed with pre-heated hybridization buffer, centrifuged and resuspended in Millipore quality water before filtration onto a 0.22 µm black polycarbonate filter. The probes used in this study were, LGC354A, LGC354B, LGC354C, Strc493, Bacto1080, Sal3, Chis150, EUB...... were counted by the EUB338 probe. Three weeks old male broiler Ross 308 chickens were used to investigate the bacterial composition of the intestine. The birds received a wheat-barley diet. Counts with the EUB338 probe were 1.97x108(std 1.45x108) The means of counts obtained with probes targeting the r...

  20. Effect of ochratoxin A on the intestinal mucosa and mucosa-associated lymphoid tissues in broiler chickens.

    Science.gov (United States)

    Solcan, Carmen; Pavel, Geta; Floristean, Viorel Cezar; Chiriac, Ioan Sorin Beschea; Şlencu, Bogdan Gabriel; Solcan, Gheorghe

    2015-03-01

    The immunotoxic effect of ochratoxin A (OTA) on the intestinal mucosa-associated lymphoid tissue and its cytotoxic action on the intestinal epithelium were studied in broiler chickens experimentally treated with the toxin. From the 7th day of life, 80 male broiler chickens (Ross 308) were randomly divided into four groups of 20 birds each. The three experimental groups (E1-3) were treated with OTA for 28 days (E1: 50 μg/kg body weight [bw]/day; E2: 20 μg/kg bw/day; E3: 1 μg/kg bw/day) and the fourth group served as control. Histological examination of the intestinal mucosa and immunohistochemical staining for identification of CD4+, CD8+, TCR1 and TCR2 lymphocytes in the duodenum, jejunum and ileocaecal junction were performed, and CD4+/CD8+ and TCR1/TCR2 ratios were calculated. OTA toxicity resulted in decreased body weight gain, poorer feed conversion ratio, lower leukocyte and lymphocyte count, and altered intestinal mucosa architecture. After 14 days of exposure to OTA, immunohistochemistry showed a significant reduction of the lymphocyte population in the intestinal epithelium and the lamina propria. After 28 days of exposure, an increase in the CD4+ and CD8+ values in both the duodenum and jejunum of chickens in Groups E1 and E2 was observed, but the TCR1 and TCR2 lymphocyte counts showed a significant reduction. No significant changes were observed in Group E3. The results indicate that OTA induced a decrease in leukocyte and lymphocyte counts and was cytotoxic to the intestinal epithelium and the mucosa-associated lymphoid tissue, altering the intestinal barrier and increasing susceptibility to various associated diseases.

  1. Determination of boron concentration in biopsy-sized tissue samples

    International Nuclear Information System (INIS)

    Hou, Yougjin; Fong, Katrina; Edwards, Benjamin; Autry-Conwell, Susan; Boggan, James

    2000-01-01

    Inductively coupled plasma mass spectrometry (ICP-MS) is the most sensitive analytical method for boron determination. However, because boron is volatile and ubiquitous in nature, low-concentration boron sample measurement remains a challenge. In this study, an improved ICP-MS method was developed for quantitation of tissue samples with low (less than 10 ppb) and high (100 ppb) boron concentrations. The addition of an ammonia-mannitol solution converts volatile boric acid to the non-volatile ammonium borate in the spray chamber and with the formation of a boron-mannitol complex, the boron memory effect and background are greatly reduced. This results in measurements that are more accurate, repeatable, and efficient. This improved analysis method has facilitated rapid and reliable tissue biodistribution analyses of newly developed boronated compounds for potential use in neutron capture therapy. (author)

  2. Quantitation of ranaviruses in cell culture and tissue samples

    DEFF Research Database (Denmark)

    Holopainen, Riikka; Honkanen, Jarno; Jensen, Britt Bang

    2011-01-01

    – epithelioma papulosum cyprini (EPC) and bluegill fry (BF-2) – were infected with four of the isolates (EHNV, ECV, FV3 and DFV), and the viral quantity was determined from seven time points during the first three days after infection. The qPCR was also used to determine the viral load in tissue samples from...... pike (Esox lucius) fry challenged experimentally with EHNV....

  3. Proteomic analysis of tissue samples in translational breast cancer research

    DEFF Research Database (Denmark)

    Gromov, Pavel; Moreira, José; Gromova, Irina

    2014-01-01

    In the last decade, many proteomic technologies have been applied, with varying success, to the study of tissue samples of breast carcinoma for protein expression profiling in order to discover protein biomarkers/signatures suitable for: characterization and subtyping of tumors; early diagnosis......, and both prognosis and prediction of outcome of chemotherapy. The purpose of this review is to critically appraise what has been achieved to date using proteomic technologies and to bring forward novel strategies - based on the analysis of clinically relevant samples - that promise to accelerate...

  4. Epithelial-Mesenchymal Interactions in Urinary Bladder and Small Intestine and How to Apply Them in Tissue Engineering.

    Science.gov (United States)

    Jerman, Urška Dragin; Kreft, Mateja Erdani; Veranič, Peter

    2015-12-01

    Reciprocal interactions between the epithelium and mesenchyme are essential for the establishment of proper tissue morphology during organogenesis and tissue regeneration as well as for the maintenance of cell differentiation. With this review, we highlight the importance of epithelial-mesenchymal cross talk in healthy tissue and further discuss its significance in engineering functional tissues in vitro. We focus on the urinary bladder and small intestine, organs that are often compromised by disease and are as such in need of research that would advance effective treatment or tissue replacement. To date, the understanding of epithelial-mesenchymal reciprocal interactions has enabled the development of in vitro biomimetic tissue equivalents that have provided many possibilities in treating defective, damaged, or even cancerous tissues. Although research of the past several years has advanced the field of bladder and small intestine tissue engineering, one must be aware of its current limitations in successfully and above all safely introducing tissue-engineered constructs into clinical practice. Special attention is in particular needed when treating cancerous tissues, as initially successful tumor excision and tissue reconstruction may later on result in cancer recurrence due to oncogenic signals originating from an altered stroma. Recent rather poor outcomes in pioneering clinical trials of bladder reconstructions should serve as a reminder that recreating a functional organ to replace a dysfunctional one is an objective far more difficult to reach than initially foreseen. When considering effective tissue engineering approaches for diseased tissues in humans, it is imperative to introduce animal models with dysfunctional or, even more importantly, cancerous organs, which would greatly contribute to predicting possible complications and, hence, reducing risks when translating to the clinic.

  5. Effects of honey to mobilize endogenous stem cells in efforts intestinal and ovarian tissue regeneration in rats with protein energy malnutrition

    Directory of Open Access Journals (Sweden)

    R. Heru Prasetyo

    2016-05-01

    Conclusions: Expression of CD34+ and CD45+, which significantly different in treatment 2 (2. Furthermore, increase of immune response (decrease Hsp70 expression and increased PGE2 in intestinal tissue. Increased immune response causes expression of GDF-9 in ovarian tissue. Decreased of Hsp70 expression, increased PGE2 and increased GDF-9 followed the process of regeneration of the intestinal and ovarian tissue.

  6. Tissue engineered esophagus scaffold constructed with porcine small intestinal submucosa and synthetic polymers.

    Science.gov (United States)

    Fan, Mei-Rong; Gong, Mei; Da, Lin-Cui; Bai, Lin; Li, Xiu-Qun; Chen, Ke-Fei; Li-Ling, Jesse; Yang, Zhi-Ming; Xie, Hui-Qi

    2014-02-01

    Acellular porcine small intestinal submucosa (SIS) has been successfully used for reconstructing esophagus with half circumferential defects. However, repairing full circumferential esophageal defects with SIS has been restricted due to the latter's poor mechanical properties. In the present study, synthetic polyesters biomaterial poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) and poly(lactide-co-glycolide) (PLGA) have been used to improve the mechanical properties of SIS. Feasibility of SIS/PHBHHx-PLGA composite material scaffold for esophageal tissue engineering has been assessed through a series of testing. The appropriate mixing ratio of PHBHHx and PLGA polymers has been determined as 5:5 by mechanical testing and in vitro degradation experiment. The morphology of constructed membranous and tubular scaffolds was also characterized. As confirmed by enzyme-linked immunosorbent assay, the contents of VEGF and TGF-β have respectively reached 657 ± 18 ng mL(-1) and 130 ± 4 pg mL(-1) within the SIS/PHBHHx-PLGA specimens. Biocompatibility of the SIS/PHBHHx-PLGA specimens with rat bone marrow mesenchymal stem cells (MSCs) was also evaluated by scanning electron microscopy and a live-dead cell viability assay. Actin filaments of MSCs on the composite materials were labeled. Biological safety of the extract from SIS/PHBHHx-PLGA specimens, measured as hemolysis rate, was all lower than 5%. Compared with SIS and SIS/PHBHHx-PLGA specimens, inflammatory reaction provoked by the PHBHHx-PLGA specimens in rats was however more severe. Our results have suggested that SIS/PHBHHx-PLGA composite material can offer a new approach for esophageal tissue engineering.

  7. Sex identification of polar bears from blood and tissue samples

    Science.gov (United States)

    Amstrup, Steven C.; Garner, G.W.; Cronin, M.A.; Patton, J.C.

    1993-01-01

    Polar bears (Ursus maritimus) can be adversely affected by hunting and other human perturbations because of low population densities and low reproduction rates. The sustainable take of adult females may be as low as 1.5% of the population. Females and accompanying young are most vulnerable to hunting, and hunters have not consistently reported the sex composition of the harvest, therefore a method to confirm the sexes of polar bears harvested in Alaska is needed. Evidence of the sex of harvested animals is often not available, but blood or other tissue samples often are. We extracted DNA from tissue and blood samples, and amplified segments of zinc finger (ZFX and ZFY) genes from both X and Y chromosomes with the polymerase chain reaction. Digestion of amplified portions of the X chromosome with the restriction enzyme HaeIII resulted in subdivision of the original amplified segment into four smaller fragments. Digestion with HaeIII did not subdivide the original segment amplified from the Y chromosome. The differing fragment sizes produced patterns in gel electrophoresis that distinguished samples from male and female bears 100% of the time. This technique is applicable to the investigation of many wildlife management and research questions.

  8. Theory of sampling and its application in tissue based diagnosis

    Directory of Open Access Journals (Sweden)

    Kayser Gian

    2009-02-01

    Full Text Available Abstract Background A general theory of sampling and its application in tissue based diagnosis is presented. Sampling is defined as extraction of information from certain limited spaces and its transformation into a statement or measure that is valid for the entire (reference space. The procedure should be reproducible in time and space, i.e. give the same results when applied under similar circumstances. Sampling includes two different aspects, the procedure of sample selection and the efficiency of its performance. The practical performance of sample selection focuses on search for localization of specific compartments within the basic space, and search for presence of specific compartments. Methods When a sampling procedure is applied in diagnostic processes two different procedures can be distinguished: I the evaluation of a diagnostic significance of a certain object, which is the probability that the object can be grouped into a certain diagnosis, and II the probability to detect these basic units. Sampling can be performed without or with external knowledge, such as size of searched objects, neighbourhood conditions, spatial distribution of objects, etc. If the sample size is much larger than the object size, the application of a translation invariant transformation results in Kriege's formula, which is widely used in search for ores. Usually, sampling is performed in a series of area (space selections of identical size. The size can be defined in relation to the reference space or according to interspatial relationship. The first method is called random sampling, the second stratified sampling. Results Random sampling does not require knowledge about the reference space, and is used to estimate the number and size of objects. Estimated features include area (volume fraction, numerical, boundary and surface densities. Stratified sampling requires the knowledge of objects (and their features and evaluates spatial features in relation to

  9. Fusion of intestinal epithelial cells with bone marrow derived cells is dispensable for tissue homeostasis

    NARCIS (Netherlands)

    de Jong, Joan H.; Rodermond, Hans M.; Zimberlin, Cheryl D.; Lascano, Valeria; de Sousa E Melo, Felipe; Richel, Dick J.; Medema, Jan Paul; Vermeulen, Louis

    2012-01-01

    The epithelial lining of the intestine is characterized by an immense cellular turn-over ascertaining an extensive regenerative capacity. Multiple reports suggest that besides the local intestinal stem cell pool, circulating cells of bone marrow origin (BMDCs) contribute to this process by fusing

  10. Intestinal Tissues Induce an SNP Mutation in Pseudomonas aeruginosa That Enhances Its Virulence: Possible Role in Anastomotic Leak

    OpenAIRE

    Olivas, Andrea D.; Shogan, Benjamin D.; Valuckaite, Vesta; Zaborin, Alexander; Belogortseva, Natalya; Musch, Mark; Meyer, Folker; L.Trimble, William; An, Gary; Gilbert, Jack; Zaborina, Olga; Alverdy, John C.

    2012-01-01

    The most feared complication following intestinal resection is anastomotic leakage. In high risk areas (esophagus/rectum) where neoadjuvant chemoradiation is used, the incidence of anastomotic leaks remains unacceptably high (≈ 10%) even when performed by specialist surgeons in high volume centers. The aims of this study were to test the hypothesis that anastomotic leakage develops when pathogens colonizing anastomotic sites become in vivo transformed to express a tissue destroying phenotype....

  11. How You Can Help Medical Research: Donating Your Blood, Tissue, and Other Samples

    Science.gov (United States)

    ... HUMAN SERVICES National Institutes of Health Donating Your Blood, Tissue, and Other Samples You have the choice to donate samples, such as blood and tissue, for medical research. Medical researchers use samples to ...

  12. Radionuclides in animal tissue samples from various regions of Austria

    International Nuclear Information System (INIS)

    Tatzber, F.

    1981-06-01

    An investigation of the concentration of radioactive substances in animal species from various regions of Austria has been carried out. For bone and liver of deer, radionuclide concentrations typical for central Europe were found. The content of 90 Sr were higher in gasteropod shells than in deer bone. Similar concentrations of 90 Sr were found in isopods as in snail shells related to fresh weight, but related to Ca content the values in isopods were higher than in all other animals. Based on these results, a study of snail shells and of isopods as bioindicators for 90 Sr content in environmental control is indicated. In tissue samples of the same species, but from different regions of Austria, the fallout radionuclide concentrations were found to be related to altitude ( 90 Sr) and to the amount of precipitation ( 137 Cs). These correlation differences could point to a different deposition behaviours of 90 Sr and 137 Cs, the former being deposited mainly with solid precipitation. This seems plausible since aerosols carried over continental distances show a high sulfate content and alkaline earth metal sulfates are less soluble than alkali sulfates. Examination of absolute concentration values related to fresh tissue weight show high fallout radionuclide concentrations, as compared to natural radionuclide concentration, especially in hard tissues. These fallout levels constitute a significant radioactive load on the biosphere. Due to the long physical half-life of 90 Sr and 137 Cs, this situation will remain virtually unchanged during the next decades, even if no further nuclear weapons tests are carried out. (G.G.)

  13. Measurement of phthalates in small samples of mammalian tissue

    International Nuclear Information System (INIS)

    Acott, P.D.; Murphy, M.G.; Ogborn, M.R.; Crocker, J.F.S.

    1987-01-01

    Di-(2-ethylhexyl)-phthalate (DEHP) is a phthalic acid ester that is used as a plasticizer in polyvinyl chloride products, many of which have widespread medical application. DEHP has been shown to be leached from products used for storage and delivery of blood transfusions during procedures such as plasmaphoresis, hemodialysis and open heart surgery. Results of studies in this laboratory have suggested that there is an association between the absorption and deposition of DEHP (and/or related chemicals) in the kidney and the acquired renal cystic disease (ACD) frequently seen in patients who have undergone prolonged dialysis treatment. In order to determine the relationship between the two, it has been necessary to establish a method for extracting and accurately quantitating minute amounts of these chemicals in small tissue samples. The authors have now established such a method using kidneys from normal rats and from a rat model for ACD

  14. Dendritic Cells Produce CXCL13 and Participate in the Development of Murine Small Intestine Lymphoid Tissues

    OpenAIRE

    McDonald, Keely G.; McDonough, Jacquelyn S.; Dieckgraefe, Brian K.; Newberry, Rodney D.

    2010-01-01

    In the adult intestine, luminal microbiota induce cryptopatches to transform into isolated lymphoid follicles (ILFs), which subsequently act as sites for the generation of IgA responses. The events leading to this conversion are incompletely understood. Dendritic cells (DCs) are components of cryptopatches (CPs) and ILFs and were therefore evaluated in this process. We observed that the adult murine intestine contains clusters of DCs restricted to the CP/ILF continuum. A numerical and cell as...

  15. Fusion of intestinal epithelial cells with bone marrow derived cells is dispensable for tissue homeostasis

    OpenAIRE

    de Jong, Joan H.; Rodermond, Hans M.; Zimberlin, Cheryl D.; Lascano, Valeria; De Sousa E Melo, Felipe; Richel, Dick J.; Medema, Jan Paul; Vermeulen, Louis

    2012-01-01

    The epithelial lining of the intestine is characterized by an immense cellular turn-over ascertaining an extensive regenerative capacity. Multiple reports suggest that besides the local intestinal stem cell pool, circulating cells of bone marrow origin (BMDCs) contribute to this process by fusing with the epithelial lineage. However, the functional relevance of these observations is unknown. In the present study we employ a model system in which we cannot only detect cell fusion but also exam...

  16. Effect of a cocoa diet on the small intestine and gut-associated lymphoid tissue composition in an oral sensitization model in rats.

    Science.gov (United States)

    Camps-Bossacoma, Mariona; Pérez-Cano, Francisco J; Franch, Àngels; Untersmayr, Eva; Castell, Margarida

    2017-04-01

    Previous studies have attributed to the cocoa powder the capacity to attenuate the immune response in a rat oral sensitization model. To gain a better understanding of cocoa-induced mechanisms at small intestinal level, 3-week-old female Lewis rats were fed either a standard diet or a diet containing 10% cocoa for 4 weeks with or without concomitant oral sensitization with ovalbumin (OVA). Thereafter, we evaluated the lymphocyte composition of the Peyer's patches (PPL), small intestine epithelium (IEL) and lamina propria (LPL). Likewise, gene expression of several immune molecules was quantified in the small intestine. Moreover, histological samples were used to evaluate the proportion of goblet cells, IgA+ cells and granzyme+cells as well. In cocoa-fed animals, we identified a five-time reduction in the percentage of IgA+ cells in intestinal tissue together with a decreased proportion of TLR4+ IEL. Analyzing the lymphocyte composition, almost a double proportion of TCRγδ+cells and an increase of NK cell percentage in PPL and IEL were found. In addition, a rise in CD25+, CD103+ and CD62L- cell proportions was observed in CD4+ PPL from cocoa-fed animals, along with a decrease in gene expression of CD11b, CD11c and IL-10. These results suggest that changes in PPL and IEL composition and in the gene expression induced by the cocoa diet could be involved, among other mechanisms, on its tolerogenic effect. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Late effects of intraoperative radiation therapy on retroperitoneal tissues, intestine, and bile duct in a large animal model

    Energy Technology Data Exchange (ETDEWEB)

    Sindelar, W.F.; Tepper, J.E.; Kinslla, T.J.; Barnes, M.; DeLuca, A.M.; Terrill, R.; Matthews, D.; Johnstone, P.A.S. [National Institutes of Health, Bethesda, MD (United States); Anderson, W.J. [Terre Haute Center for Medical Education, IN (United States); Bollinger, B.K. [National Naval Medical Center, Bethesda, MD (United States)

    1994-07-01

    The late histopathological effects of intraoperative radiotherapy (IORT) on retroperitoneal tissues, intestine, and bile duct were investigated in dogs. Fourteen adult foxhounds were subjected to laparotomy and varying doses (0-45 Gy) of IORT (11 MeV electrons) delivered to retroperitoneal tissues including the great vessels and ureters, to a loop of defunctionalized small bowel, or to the extrahepatic bile duct. One control animal received an aortic transection and reanastomosis at the time of laparotomy; another control received laparotomy alone. This paper describes the late effects of single-fraction IORT occurring 3-5 years following treatment. Dogs receiving IORT to the retroperitoneum through a 4 X 15 cm portal showed few gross or histologic abnormalities at 20 Gy. At doses ranging from 30-45 Gy, radiation changes in normal tissues were consistently observed. Retroperitoneal fibrosis with encasement of the ureters and great vessels developed at doses {ge}30 Gy. Radiation changes were present in the aorta and vena cava at doses {ge}40 Gy. A 30 Gy dog developed an in-field malignant osteosarcoma at 3 years which invaded the vertebral column and compressed the spinal cord. A 40 Gy animal developed obstruction of the right ureter with fatal septic hydronephrosis at 4 years. Animals receiving IORT through a 5 cm IORT portal to an upper abdominal field which included a defunctionalized loop of small bowel, showed few gross or histologic abnormalities at a dose of 20 Gy. At 30 Gy, hyaline degeneration of the intestinal muscularis layer of the bowel occurred. At a dose of 45 Gy, internal intestinal fistulae developed. One 30 Gy animal developed right ureteral obstruction and hydronephrosis at 5 years. A dog receiving 30 Gy IORT through a 5 cm portal to the extrahepatic bile duct showed diffuse fibrosis through the gastroduodenal ligament. These canine studies contribute to the area of late tissue tolerance to IORT. 7 refs., 3 figs., 5 tabs.

  18. Late effects of intraoperative radiation therapy on retroperitoneal tissues, intestine, and bile duct in a large animal model

    International Nuclear Information System (INIS)

    Sindelar, W.F.; Tepper, J.E.; Kinslla, T.J.; Barnes, M.; DeLuca, A.M.; Terrill, R.; Matthews, D.; Johnstone, P.A.S.; Anderson, W.J.; Bollinger, B.K.

    1994-01-01

    The late histopathological effects of intraoperative radiotherapy (IORT) on retroperitoneal tissues, intestine, and bile duct were investigated in dogs. Fourteen adult foxhounds were subjected to laparotomy and varying doses (0-45 Gy) of IORT (11 MeV electrons) delivered to retroperitoneal tissues including the great vessels and ureters, to a loop of defunctionalized small bowel, or to the extrahepatic bile duct. One control animal received an aortic transection and reanastomosis at the time of laparotomy; another control received laparotomy alone. This paper describes the late effects of single-fraction IORT occurring 3-5 years following treatment. Dogs receiving IORT to the retroperitoneum through a 4 X 15 cm portal showed few gross or histologic abnormalities at 20 Gy. At doses ranging from 30-45 Gy, radiation changes in normal tissues were consistently observed. Retroperitoneal fibrosis with encasement of the ureters and great vessels developed at doses ≥30 Gy. Radiation changes were present in the aorta and vena cava at doses ≥40 Gy. A 30 Gy dog developed an in-field malignant osteosarcoma at 3 years which invaded the vertebral column and compressed the spinal cord. A 40 Gy animal developed obstruction of the right ureter with fatal septic hydronephrosis at 4 years. Animals receiving IORT through a 5 cm IORT portal to an upper abdominal field which included a defunctionalized loop of small bowel, showed few gross or histologic abnormalities at a dose of 20 Gy. At 30 Gy, hyaline degeneration of the intestinal muscularis layer of the bowel occurred. At a dose of 45 Gy, internal intestinal fistulae developed. One 30 Gy animal developed right ureteral obstruction and hydronephrosis at 5 years. A dog receiving 30 Gy IORT through a 5 cm portal to the extrahepatic bile duct showed diffuse fibrosis through the gastroduodenal ligament. These canine studies contribute to the area of late tissue tolerance to IORT. 7 refs., 3 figs., 5 tabs

  19. Cloning of the rat ecotropic retroviral receptor and studies of its expression in intestinal tissues

    Energy Technology Data Exchange (ETDEWEB)

    Puppi, M.; Henning, S.J. [Baylor College of Medicine, Houston, TX (United States)

    1995-05-01

    A long-term goal of our laboratory is to establish a rat model to study the feasibility of using the intestinal tract as a site for somatic gene therapy. As a step toward that goal, the current study reports the cloning of the rat ecotropic retroviral receptor (EcoR) cDNA and the study of various aspects of its expression in the intestinal cDNA library with mouse EcoR cDNA. A clone of approximately 7 kb, designated MP10, was obtained. Partial sequencing of MP10 from the 5{prime} end revealed a level of similarity of 92% compared with mouse EcoR. The presence of a 5{prime} untranslated region and a 3{prime} poly(A)tract, together with the overall size of the cDNA, suggest that is very close to being a full-length cDNA for this large transcript. Northern blots with MP10 showed an RNA of approximately 7.9 kb present along the entire length of the small intestine and somewhat less abundant in the colon. Developmental studies showed high levels of EcoR in fetal rat intestine, a decline in the early postnatal period, then a gradual rise to adulthood. Caco-2 cells were used to assess the expression of EcoR in proliferating compared with differentiated intestinal epithelial cells. EcoR mRNA was found to be very much more abundant in nondifferentiated cells and declined to low levels as the cells underwent spontaneous differentiation. These patterns of EcoR expression indicate that ecotropic retroviruses should be suitable vectors with which to attempt gene transfer into the intestinal epithelium. In addition, since the endogenous role of EcoR is as the y{sup +} cationic amino acid transporter, these data have significance for understanding patterns of amino acid transport in the intestinal epithelium. 37 refs., 4 figs.

  20. Gene Expression Changes in Mouse Intestinal Tissue Following Whole-Body Proton or Gamma-Irradiation

    Science.gov (United States)

    Purgason, Ashley; Zhang, Ye; Mangala, Lingegowda; Nie, Ying; Gridley, Daila; Hamilton, Stanley R.; Seidel, Derek V.; Wu, Honglu

    2014-01-01

    Crew members face potential consequences following exposure to the space radiation environment including acute radiation syndrome and cancer. The space radiation environment is ample with protons, and numerous studies have been devoted to the understanding of the health consequences of proton exposures. In this project, C57BL/6 mice underwent whole-body exposure to 250 MeV of protons at doses of 0, 0.1, 0.5, 2 and 6 Gy and the gastrointestinal (GI) tract of each animal was dissected four hours post-irradiation. Standard H&E staining methods to screen for morphologic changes in the tissue showed an increase in apoptotic lesions for even the lowest dose of 0.1 Gy, and the percentage of apoptotic cells increased with increasing dose. Results of gene expression changes showed consistent up- or down- regulation, up to 10 fold, of a number of genes across exposure doses that may play a role in proton-induced oxidative stress including Gpx2. A separate study in C57BL/6 mice using the same four hour time point but whole-body gamma-irradiation showed damage to the small intestine with lesions appearing at the smallest dose of 0.05 Gy and increasing with increasing absorbed dose. Expressions of genes associated with oxidative stress processes were analyzed at four hours and twenty-four hours after exposure to gamma rays. We saw a much greater number of genes with significant up- or down-regulation twenty-four hours post-exposure as compared to the four hour time point. At both four hours and twenty-four hours post-exposure, Duox1 and Mpo underwent up-regulation for the highest dose of 6 Gy. Both protons and gamma rays lead to significant variation in gene expressions and these changes may provide insight into the mechanism of injury seen in the GI tract following radiation exposure. We have also completed experiments using a BALB/c mouse model undergoing whole-body exposure to protons. Doses of 0, 0.1, 1 and 2 Gy were used and results will be compared to the work mentioned

  1. Exploring drug solubility in fasted human intestinal fluid aspirates: Impact of inter-individual variability, sampling site and dilution.

    Science.gov (United States)

    de la Cruz-Moreno, Mariangeles Pérez; Montejo, Consuelo; Aguilar-Ros, Antonio; Dewe, Walthère; Beck, Benoît; Stappaerts, Jef; Tack, Jan; Augustijns, Patrick

    2017-08-07

    One of the main factors defining intestinal drug absorption is the solubility of the compound in the gastrointestinal environment. This study reports the solubility of a series of 27 commonly used acidic, neutral and basic drugs in human intestinal fluid samples collected from the duodenum or jejunum of healthy volunteers under fasted state conditions. The interindividual variability as well as the impact of factors such as pH, sampling site and bile salts on the solubility in human intestinal fluids was investigated. The solubility measurements were evaluated using a statistical experimental design. Variability in solubility across volunteers and sampling sites was highly compound-specific and appeared to be substantial for weak acids and bases and for lipophilic drugs. Both pH of the samples and the abundance of amphiphilic components were responsible for the variability observed in the solubility values obtained. The results confirm strong interindividual differences in intraluminal solubility, especially for compounds with high lipophilicity and/or compounds with a pKa value within the physiological pH range. It is important to recognize this variability in intestinal drug solubility as it may considerably influence the therapeutic outcome among patients. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Ret receptor tyrosine kinase sustains proliferation and tissue maturation in intestinal epithelia.

    Science.gov (United States)

    Perea, Daniel; Guiu, Jordi; Hudry, Bruno; Konstantinidou, Chrysoula; Milona, Alexandra; Hadjieconomou, Dafni; Carroll, Thomas; Hoyer, Nina; Natarajan, Dipa; Kallijärvi, Jukka; Walker, James A; Soba, Peter; Thapar, Nikhil; Burns, Alan J; Jensen, Kim B; Miguel-Aliaga, Irene

    2017-10-16

    Expression of the Ret receptor tyrosine kinase is a defining feature of enteric neurons. Its importance is underscored by the effects of its mutation in Hirschsprung disease, leading to absence of gut innervation and severe gastrointestinal symptoms. We report a new and physiologically significant site of Ret expression in the intestine: the intestinal epithelium. Experiments in Drosophila indicate that Ret is expressed both by enteric neurons and adult intestinal epithelial progenitors, which require Ret to sustain their proliferation. Mechanistically, Ret is engaged in a positive feedback loop with Wnt/Wingless signalling, modulated by Src and Fak kinases. We find that Ret is also expressed by the developing intestinal epithelium of mice, where its expression is maintained into the adult stage in a subset of enteroendocrine/enterochromaffin cells. Mouse organoid experiments point to an intrinsic role for Ret in promoting epithelial maturation and regulating Wnt signalling. Our findings reveal evolutionary conservation of the positive Ret/Wnt signalling feedback in both developmental and homeostatic contexts. They also suggest an epithelial contribution to Ret loss-of-function disorders such as Hirschsprung disease. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  3. Dendritic cells produce CXCL13 and participate in the development of murine small intestine lymphoid tissues.

    Science.gov (United States)

    McDonald, Keely G; McDonough, Jacquelyn S; Dieckgraefe, Brian K; Newberry, Rodney D

    2010-05-01

    In the adult intestine, luminal microbiota induce cryptopatches to transform into isolated lymphoid follicles (ILFs), which subsequently act as sites for the generation of IgA responses. The events leading to this conversion are incompletely understood. Dendritic cells (DCs) are components of cryptopatches (CPs) and ILFs and were therefore evaluated in this process. We observed that the adult murine intestine contains clusters of DCs restricted to the CP/ILF continuum. A numerical and cell associative hierarchy in the adult intestine and a chronologic hierarchy in the neonatal intestine demonstrated that these clusters form after the coalescence of CD90+ cells to form CPs and before the influx of B220+ B lymphocytes to form ILFs. Cluster formation was dependent on lymphotoxin and the lymphotoxin beta receptor and independent of lymphocytes. The ILF DC population was distinguished from that of the lamina propria by the absence of CD4+CD11c+ cells and an increased proportion of CD11c+B220+ cells. The formation of clusters was not limited by DC numbers but was induced by luminal microbiota. Moreover, in the absence of the chemokine CXCL13, CP transformation into ILF was arrested. Furthermore, ILF DCs express CXCL13, and depletion of DCs resulted in regression of ILFs and disorganization of CPs. These results reveal DC participation in ILF transformation and maintenance and suggest that in part this may be due to CXCL13 production by these cells.

  4. Comparative MiRNA Expressional Profiles and Molecular Networks in Human Small Bowel Tissues of Necrotizing Enterocolitis and Spontaneous Intestinal Perforation.

    Directory of Open Access Journals (Sweden)

    Pak Cheung Ng

    Full Text Available Necrotizing enterocolitis (NEC and spontaneous intestinal perforation (SIP are acute intestinal conditions which could result in mortality and severe morbidity in preterm infants. Our objective was to identify dysregulated micro-RNAs (miRNAs in small bowel tissues of NEC and SIP, and their possible roles in disease pathophysiology.We performed differential miRNA arrays on tissues of NEC (n = 4, SIP (n = 4 and surgical-control (Surg-CTL; n = 4, and validated target miRNAs by qPCR (n = 10 each group. The association of target miRNAs with 52 dysregulated mRNAs was investigated by bioinformatics on functional and base-pair sequence algorithms, and correlation in same tissue samples.We presented the first miRNA profiles of NEC, SIP and Surg-CTL intestinal tissues in preterm infants. Of 28 validated miRNAs, 21 were significantly different between NEC or SIP and Surg-CTL. Limited overlapping in the aberrant expression of miRNAs between NEC and SIP indicated their distinct molecular mechanisms. A proposed network of dysregulated miRNA/mRNA pairs in NEC suggested interaction at bacterial receptor TLR4 (miR-31, miR-451, miR-203, miR-4793-3p, mediated via key transcription factors NFKB2 (miR-203, AP-1/FOSL1 (miR-194-3p, FOXA1 (miR-21-3p, miR-431 and miR-1290 and HIF1A (miR-31, and extended downstream to pathways of angiogenesis, arginine metabolism, cell adhesion and chemotaxis, extracellular matrix remodeling, hypoxia/oxidative stress, inflammation and muscle contraction. In contrast, upregulation of miR-451 and miR-223 in SIP suggested modulation of G-protein-mediated muscle contraction.The robust response of miRNA dysregulation in NEC and SIP, and concerted involvement of specific miRNAs in the molecular networks indicated their crucial roles in mucosa integrity and disease pathophysiology.

  5. Characterization of stable RNAs from the resected intestinal tissues of individuals with either Crohn’s disease or ulcerative colitis

    Science.gov (United States)

    Roy, Guylaine; Mercure, Stéphane; Beuvon, Frédéric; Perreault, Jean-Pierre

    2010-01-01

    Circular RNAs reminiscent of viroids and the human hepatitis delta virus have been proposed as possible nonconventional pathogens responsible for Crohn’s disease and ulcerative colitis, two inflammatory bowel diseases. Consequently, RNA was extracted from various areas of intestinal tissues from individuals with either Crohn’s disease or ulcerative colitis as well as several appropriate control diseases, and analyzed by two-dimensional gel electrophoresis. No circular viroid-like RNAs (28S ribosomal RNA cleavage product characterized previously; a 5.8S ribosomal RNA conformer; and a fragment homologous to transcripts from DNA CpG islands. The two last RNAs were detected prior to visible morphological tissue alterations, suggesting that they are produced early during the inflammation and that they have value as molecular diagnostic tools for the inflammatory bowel diseases. The potential cellular mechanisms producing these RNAs and their involvement in inflammatory bowel disease are discussed. PMID:9599669

  6. Morphologic observation of mucosa-associated lymphoid tissue in the large intestine of Bactrian camels (Camelus bactrianus).

    Science.gov (United States)

    ZhaXi, Yingpai; Wang, Wenhui; Zhang, Wangdong; Gao, Qiang; Guo, Minggang; Jia, Shuai

    2014-07-01

    The structure and distribution of the mucosa-associated lymphoid tissue (MALT) throughout the large intestine of 10 Bactrian camels were comparatively studied by anatomical and histological methods. The results showed that Peyer's patches (PPs) were mainly located on the mucosal surfaces of the entire ileocecal orifice, the beginning of the cecum and the first third of the colon. The shape of PPs gradually changed from "scrotiform" to "faviform" along the large intestine with the scrotiform PP as the major type in the ileocecal orifice. The distribution density also gradually decreased from the ileocecal orifice to the colon. The histological observations further revealed that the MALT in the form of PPs or isolated lymphoid follicles (ILF) and lamina propria lymphocytes was mainly present in the lamina propria and submucosa from the entire ileocecal orifice, where the muscularis mucosa is usually incomplete, to the colonic forepart. In addition, lymphoid tissue was much more abundant in the lamina propria and submucosa of the ileocecal orifice as compared to the cecum and colon. Statistically, the MALT of the ileocecal orifice contained a higher number of lymphoid follicles (37.7/10 mm(2) ) than that of the cecum, colon, or rectum (P lymphoid follicles were clearly visible. Together, our data suggest that the ileocecal orifice constitutes the main inductive site for the mucosal immunity in the large intestine of the Bactrian camel; and that scrotiform PPs are likely to the result of long-term adaptation of the Bactrian camel to the harsh living environment. © 2014 Wiley Periodicals, Inc.

  7. Signal transduction pathways participating in homeostasis and malignant transformation of the intestinal tissue

    Czech Academy of Sciences Publication Activity Database

    Krausová, Michaela; Kořínek, Vladimír

    2012-01-01

    Roč. 59, č. 6 (2012), s. 708-718 ISSN 0028-2685 R&D Projects: GA ČR GAP305/11/1780; GA ČR GAP305/12/2347; GA ČR GAP304/11/1252; GA ČR GD204/09/H058 Keywords : colorectal cancer * epithelium * gut * intestine * mouse models * stem cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.574, year: 2012

  8. The evaluation of risk-benefit ratio for gut tissue sampling in HIV cure research.

    Science.gov (United States)

    Mehraj, Vikram; Ghali, Peter; Ramendra, Rayoun; Costiniuk, Cecilia; Lebouché, Bertrand; Ponte, Rosalie; Reinhard, Robert; Sousa, Jose; Chomont, Nicolas; Cohen, Eric A; Ancuta, Petronela; Routy, Jean-Pierre

    2017-10-01

    Antiretroviral therapy (ART) does not cure HIV infection due to the persistence of HIV reservoirs in long-lived memory CD4 T cells present in the blood, lymph nodes, intestinal tract, and other tissues. Interest grows in obtaining gut-tissue samples for HIV persistence studies, which poses an ethical challenge to provide study volunteers with adequate information on risks and benefits. Herein we assess the risks and benefits of undergoing gut biopsy procedures for HIV pathogenesis and reservoir studies. A group discussion was organised with physicians and community representatives on performing either a flexible sigmoidoscopy or a colonoscopy. Consensus was reached on conducting colonoscopy in persons ≥50 years. Thirty HIV-infected, ART-treated and nine uninfected participants were recruited. Colonoscopy was performed to collect 30 gut mucosal biopsies. When present, polyps were removed and abnormal mucosal findings were biopsied for pathological analysis. Participants were interviewed on potential discomfort following colonoscopic examination. The HIV-infected and uninfected groups were comparable in terms of age and gender with more men who have sex with men (MSM) in the former group. Abnormal colonoscopic findings were observed in 43.6% of all the participants and did not differ by HIV status. In total, 24 polyps were removed with a higher mean number of polyps removed in HIV-infected versus uninfected participants (1.7 vs 1.0, P =0.013). The number of polyps marginally correlated with inverted CD4:CD8 ratio. Based on our findings, colonoscopic examination was safe to use for gut biopsy procedures where almost half of the participants had polyps removed. Participation in the study provided colon cancer screening as an ancillary benefit that participants could have received in standard medical care, thus mitigating burdens of invasive procedures. Dialogue between community representatives and clinical researchers can increase participation and advance HIV cure

  9. A Comparison of Molecular and Histopathological Changes in Mouse Intestinal Tissue Following Whole-Body Proton- or Gamma-Irradiation

    Science.gov (United States)

    Purgason, Ashley; Mangala, Lingegowda; Zhang, Ye; Hamilton, Stanley; Wu, Honglu

    2010-01-01

    There are many consequences following exposure to the space radiation environment which can adversely affect the health of a crew member. Acute radiation syndrome (ARS) involving nausea and vomiting, damage to radio-sensitive tissue such as the blood forming organs and gastrointestinal tract, and cancer are some of these negative effects. The space radiation environment is ample with protons and contains gamma rays as well. Little knowledge exists to this point, however, regarding the effects of protons on mammalian systems; conversely several studies have been performed observing the effects of gamma rays on different animal models. For the research presented here, we wish to compare our previous work looking at whole-body exposure to protons using a mouse model to our studies of mice experiencing whole-body exposure to gamma rays as part of the radio-adaptive response. Radio-adaptation is a well-documented phenomenon in which cells exposed to a priming low dose of radiation prior to a higher dose display a reduction in endpoints like chromosomal aberrations, cell death, micronucleus formation, and more when compared to their counterparts receiving high dose-irradiation only. Our group has recently completed a radio-adaptive experiment with C57BL/6 mice. For both this study and the preceding proton research, the gastrointestinal tract of each animal was dissected four hours post-irradiation and the isolated small intestinal tissue was fixed in formalin for histopathological examination or snap-frozen in liquid nitrogen for RNA isolation. Histopathologic observation of the tissue using standard H&E staining methods to screen for morphologic changes showed an increase in apoptotic lesions for even the lowest doses of 0.1 Gy of protons and 0.05 Gy of gamma rays, and the percentage of apoptotic cells increased with increasing dose. A smaller percentage of crypts showed 3 or more apoptotic lesions in animals that received 6 Gy of gamma-irradiation compared to mice

  10. Regulation of serum amyloid A3 (SAA3) in mouse colonic epithelium and adipose tissue by the intestinal microbiota.

    Science.gov (United States)

    Reigstad, Christopher S; Lundén, Gunnel Ostergren; Felin, Jenny; Bäckhed, Fredrik

    2009-06-09

    The gut microbiota has been proposed as an environmental factor that affects the development of metabolic and inflammatory diseases in mammals. Recent reports indicate that gut bacteria-derived lipopolysaccharide (LPS) can initiate obesity and insulin resistance in mice; however, the molecular interactions responsible for microbial regulation of host metabolism and mediators of inflammation have not been studied in detail. Hepatic serum amyloid A (SAA) proteins are markers and proposed mediators of inflammation that exhibit increased levels in serum of insulin-resistant mice. Adipose tissue-derived SAA3 displays monocyte chemotactic activity and may play a role in metabolic inflammation associated with obesity and insulin resistance. To investigate a potential mechanistic link between the intestinal microbiota and induction of proinflammatory host factors, we performed molecular analyses of germ-free, conventionally raised and genetically modified Myd88-/- mouse models. SAA3 expression was determined to be significantly augmented in adipose (9.9+/-1.9-fold; P<0.001) and colonic tissue (7.0+/-2.3-fold; P<0.05) by the presence of intestinal microbes. In the colon, we provided evidence that SAA3 is partially regulated through the Toll-like receptor (TLR)/MyD88/NF-kappaB signaling axis. We identified epithelial cells and macrophages as cellular sources of SAA3 in the colon and found that colonic epithelial expression of SAA3 may be part of an NF-kappaB-dependent response to LPS from gut bacteria. In vitro experiments showed that LPS treatments of both epithelial cells and macrophages induced SAA3 expression (27.1+/-2.5-fold vs. 1.6+/-0.1-fold, respectively). Our data suggest that LPS, and potentially other products of the indigenous gut microbiota, might elevate cytokine expression in tissues and thus exacerbate chronic low-grade inflammation observed in obesity.

  11. Regulation of serum amyloid A3 (SAA3 in mouse colonic epithelium and adipose tissue by the intestinal microbiota.

    Directory of Open Access Journals (Sweden)

    Christopher S Reigstad

    Full Text Available The gut microbiota has been proposed as an environmental factor that affects the development of metabolic and inflammatory diseases in mammals. Recent reports indicate that gut bacteria-derived lipopolysaccharide (LPS can initiate obesity and insulin resistance in mice; however, the molecular interactions responsible for microbial regulation of host metabolism and mediators of inflammation have not been studied in detail. Hepatic serum amyloid A (SAA proteins are markers and proposed mediators of inflammation that exhibit increased levels in serum of insulin-resistant mice. Adipose tissue-derived SAA3 displays monocyte chemotactic activity and may play a role in metabolic inflammation associated with obesity and insulin resistance. To investigate a potential mechanistic link between the intestinal microbiota and induction of proinflammatory host factors, we performed molecular analyses of germ-free, conventionally raised and genetically modified Myd88-/- mouse models. SAA3 expression was determined to be significantly augmented in adipose (9.9+/-1.9-fold; P<0.001 and colonic tissue (7.0+/-2.3-fold; P<0.05 by the presence of intestinal microbes. In the colon, we provided evidence that SAA3 is partially regulated through the Toll-like receptor (TLR/MyD88/NF-kappaB signaling axis. We identified epithelial cells and macrophages as cellular sources of SAA3 in the colon and found that colonic epithelial expression of SAA3 may be part of an NF-kappaB-dependent response to LPS from gut bacteria. In vitro experiments showed that LPS treatments of both epithelial cells and macrophages induced SAA3 expression (27.1+/-2.5-fold vs. 1.6+/-0.1-fold, respectively. Our data suggest that LPS, and potentially other products of the indigenous gut microbiota, might elevate cytokine expression in tissues and thus exacerbate chronic low-grade inflammation observed in obesity.

  12. Ret receptor tyrosine kinase sustains proliferation and tissue maturation in intestinal epithelia

    DEFF Research Database (Denmark)

    Perea, Daniel; Guiu, Jordi; Hudry, Bruno

    2017-01-01

    with Wnt/Wingless signalling, modulated by Src and Fak kinases. We find that Ret is also expressed by the developing intestinal epithelium of mice, where its expression is maintained into the adult stage in a subset of enteroendocrine/enterochromaffin cells. Mouse organoid experiments point to an intrinsic...... role for Ret in promoting epithelial maturation and regulating Wnt signalling. Our findings reveal evolutionary conservation of the positive Ret/Wnt signalling feedback in both developmental and homeostatic contexts. They also suggest an epithelial contribution to Ret loss-of-function disorders...

  13. Converging biofabrication and organoid technologies : the next frontier in hepatic and intestinal tissue engineering?

    NARCIS (Netherlands)

    Schneeberger, Kerstin; Spee, Bart; Costa, Pedro; Sachs, Norman; Clevers, Hans; Malda, Jos

    2017-01-01

    Adult tissue stem cells can form self-organizing 3D organoids in vitro. Organoids resemble small units of their organ of origin and have great potential for tissue engineering, as well as models of disease. However, current culture technology limits the size, architecture and complexity of

  14. Heterotopic pancreatic tissue of the stomach leading to gastric diverticulum and upper gastro-intestinal bleeding

    Directory of Open Access Journals (Sweden)

    Silviu Stoian

    2016-11-01

    Full Text Available Heterotopic pancreatic tissue of the stomach is a rare condition. Gastric diverticulum is also a rare condition, mostly located at the fornix. Therefore, the existence of a pyloric gastric diverticulum containing a submucosal tumor proved to be heterotopic pancreatic tissue of the stomach is an extremely rare condition. The patient was a young thin male with epigastralgia chronically treated for gastritis/ulcer. Following an episode of melena, he underwent gastroscopy that diagnosed antral gastric diverticulum containing a polyp. The lesion was surgically removed. The pathology report stated: heterotopic pancreatic tissue of the stomach with secondary development of a traction diverticulum. Heterotopic pancreas tissue of the stomach is a rare condition but the association with gastric diverticulum is completely unusual. The possibility of the ectopic tissue leading to secondary diverticulum development should be considered.

  15. T-lymphocyte responses to intestinally absorbed antigens can contribute to adipose tissue inflammation and glucose intolerance during high fat feeding.

    Directory of Open Access Journals (Sweden)

    Yuehui Wang

    Full Text Available BACKGROUND: Obesity is associated with inflammation of visceral adipose tissues, which increases the risk for insulin resistance. Animal models suggest that T-lymphocyte infiltration is an important early step, although it is unclear why these cells are attracted. We have recently demonstrated that dietary triglycerides, major components of high fat diets, promote intestinal absorption of a protein antigen (ovalbumin, "OVA". The antigen was partly transported on chylomicrons, which are prominently cleared in adipose tissues. We hypothesized that intestinally absorbed gut antigens may cause T-lymphocyte associated inflammation in adipose tissue. METHODOLOGY/PRINCIPAL FINDINGS: Triglyceride absorption promoted intestinal absorption of OVA into adipose tissue, in a chylomicron-dependent manner. Absorption tended to be higher in mesenteric than subcutaneous adipose tissue, and was lowest in gonadal tissue. OVA immunoreactivity was detected in stromal vascular cells, including endothelial cells. In OVA-sensitized mice, OVA feeding caused marked accumulation of CD3+ and osteopontin+ cells in mesenteric adipose tissue. The accumulating T-lymphocytes were mainly CD4+. As expected, high-fat (60% kCal diets promoted mesenteric adipose tissue inflammation compared to low-fat diets (10% Kcal, as reflected by increased expression of osteopontin and interferon-gamma. Immune responses to dietary OVA further increased diet-induced osteopontin and interferon-gamma expression in mesenteric adipose. Inflammatory gene expression in subcutaneous tissue did not respond significantly to OVA or dietary fat content. Lastly, whereas OVA responses did not significantly affect bodyweight or adiposity, they significantly impaired glucose tolerance. CONCLUSIONS/SIGNIFICANCE: Our results suggest that loss or lack of immunological tolerance to intestinally absorbed T-lymphocyte antigens can contribute to mesenteric adipose tissue inflammation and defective glucose metabolism

  16. Effect of bovine colostrum feeding in comparison with milk replacer and natural feeding on the immune responses and colonisation of enterotoxigenic Escherichia coli in the intestinal tissue of piglets

    DEFF Research Database (Denmark)

    Sugiharto, Sugiharto; Poulsen, Ann-Sofie Riis; Canibe, Nuria

    2015-01-01

    -fed and Sow-Milk groups. The expression level of IL-2 was higher (P≤ 0·051) in piglets from the MR-fed group than in those from the other treatment groups. In conclusion, feeding BC rather than MR to the piglets reduced the colonisation of intestine by ETEC and modulated the intestinal immune system, whereas......The present study investigated the effect of feeding bovine colostrum (BC) to piglets in comparison with feeding a milk replacer (MR) and conventional rearing by the sow on the intestinal immune system and number of enterotoxigenic Escherichia coli (ETEC) colonising the intestinal tissue. Piglets...... (23-d-old) were allocated to one of the following four groups: (1) killed at the beginning of the experiment (Base); (2) separated from the sow and fed BC (BC-fed); (3) separated from the sow and fed a MR (MR-fed); (4) kept with the sow (Sow-Milk). Blood was sampled on days 1 and 8, and faecal samples...

  17. Evaluation of intestinal sampling sites in pigs at slaughter for assessing antibiotic resistance level in swine herds

    DEFF Research Database (Denmark)

    Jensen, Annette Nygaard; Thanou, Olga; Axelsdottir, Aslaug

    2012-01-01

    In the EU project SafeOrganic, the objective is to compare the level of antibiotic resistance in conventional pig herds with the level in organic pig herds, where a restricted use of antimicrobials is expected to result in less resistant bacteria. For such survey, sampling at the abattoir opposed......-colon in some pigs indicating a relatively short intestinal passage time after ingestion and then the risk of finding bacteria not originating from the host pig but from the environment. However, the proportion of the TET resistant E. coli in the large intestine appeared relatively stable over time, though...... generally a little lower than in rectal swabs. Accordingly, it seems that testing for the level of TET resistant E. coli at slaughter can allow for comparison of the presence of TET resistance also at herd level. Ongoing investigations of pigs from different herds should allow for further conclusions...

  18. Alcaligenes is Commensal Bacteria Habituating in the Gut-Associated Lymphoid Tissue for the Regulation of Intestinal IgA Responses.

    Science.gov (United States)

    Kunisawa, Jun; Kiyono, Hiroshi

    2012-01-01

    Secretory-immunoglobulin A (S-IgA) plays an important role in immunological defense in the intestine. It has been known for a long time that microbial stimulation is required for the development and maintenance of intestinal IgA production. Recent advances in genomic technology have made it possible to detect uncultivable commensal bacteria in the intestine and identify key bacteria in the regulation of innate and acquired mucosal immune responses. In this review, we focus on the immunological function of Peyer's patches (PPs), a major gut-associated lymphoid tissue, in the induction of intestinal IgA responses and the unique immunological interaction of PPs with commensal bacteria, especially Alcaligenes, a unique indigenous bacteria habituating inside PPs.

  19. Effects of fabrication on the mechanics, microstructure and micromechanical environment of small intestinal submucosa scaffolds for vascular tissue engineering.

    Science.gov (United States)

    Sánchez-Palencia, Diana M; D'Amore, Antonio; González-Mancera, Andrés; Wagner, William R; Briceño, Juan C

    2014-08-22

    In small intestinal submucosa scaffolds for functional tissue engineering, the impact of scaffold fabrication parameters on success rate may be related to the mechanotransductory properties of the final microstructural organization of collagen fibers. We hypothesized that two fabrication parameters, 1) preservation (P) or removal (R) of a dense collagen layer present in SIS and 2) SIS in a final dehydrated (D) or hydrated (H) state, have an effect on scaffold void area, microstructural anisotropy (fiber alignment) and mechanical anisotropy (global mechanical compliance). We further integrated our experimental measurements in a constitutive model to explore final effects on the micromechanical environment inside the scaffold volume. Our results indicated that PH scaffolds might exhibit recurrent and large force fluctuations between layers (up to 195 pN), while fluctuations in RH scaffolds might be larger (up to 256 pN) but not as recurrent. In contrast, both PD and RD groups were estimated to produce scarcer and smaller fluctuations (not larger than 50 pN). We concluded that the hydration parameter strongly affects the micromechanics of SIS and that an adequate choice of fabrication parameters, assisted by the herein developed method, might leverage the use of SIS for functional tissue engineering applications, where forces at the cellular level are of concern in the guidance of new tissue formation. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Ecotoxicoparasitology: Understanding mercury concentrations in gut contents, intestinal helminths and host tissues of Alaskan gray wolves (Canis lupus)

    Science.gov (United States)

    McGrew, Ashley K.; O'Hara, Todd M.; Stricker, Craig A.; Castellini, Margaret; Beckmen, Kimberlee B.; Salman, Mo D.; Ballweber, Lora R.

    2015-01-01

    Some gastrointestinal helminths acquire nutrients from the lumen contents in which they live; thus, they may be exposed to non-essential elements, such as mercury (Hg), during feeding. The objectives of this study were: 1) determine the total mercury concentrations ([THg]) in Gray wolves (Canis lupus) and their parasites, and 2) use stable isotopes to evaluate the trophic relationships within the host. [THg] and stable isotopes (C and N) were determined for helminths, host tissues, and lumen contents from 88 wolves. Sixty-three wolves contained grossly visible helminths (71.5%). The prevalence of taeniids and ascarids was 63.6% (56/88) and 20.5% (18/88), respectively. Nine of these 63 wolves contained both taeniids and ascarids (14.3%). All ascarids were determined to beToxascaris leonina. Taenia species present included T. krabbei and T. hydatigena. Within the GI tract, [THg] in the lumen contents of the proximal small intestine were significantly lower than in the distal small intestine. There was a significant positive association between hepatic and taeniid [THg]. Bioaccumulation factors (BAF) ranged from < 1 to 22.9 in taeniids, and 1.1 to 12.3 in T. leonina. Taeniid and ascarid BAF were significantly higher than 1, suggesting that both groups are capable of THg accumulation in their wolf host. δ13C in taeniids was significantly lower than in host liver and skeletal muscle. [THg] in helminths and host tissues, in conjunction with stable isotope (C and N) values, provides insight into food-web dynamics of the host GI tract, and aids in elucidating ecotoxicoparasitologic relationships. Variation of [THg] throughout the GI tract, and between parasitic groups, underscores the need to further evaluate the effect(s) of feeding niche, and the nutritional needs of parasites, as they relate to toxicant exposure and distribution within the host.

  1. RAPID PROCESSING OF ARCHIVAL TISSUE SAMPLES FOR PROTEOMIC ANALYSIS USING PRESSURE-CYCLING TECHNOLOGY

    Directory of Open Access Journals (Sweden)

    Vinuth N. Puttamallesh1,2

    2017-06-01

    Full Text Available Advent of mass spectrometry based proteomics has revolutionized our ability to study proteins from biological specimen in a high-throughput manner. Unlike cell line based studies, biomedical research involving tissue specimen is often challenging due to limited sample availability. In addition, investigation of clinically relevant research questions often requires enormous amount of time for sample collection prospectively. Formalin fixed paraffin embedded (FFPE archived tissue samples are a rich source of tissue specimen for biomedical research. However, there are several challenges associated with analysing FFPE samples. Protein cross-linking and degradation of proteins particularly affects proteomic analysis. We demonstrate that barocycler that uses pressure-cycling technology enables efficient protein extraction and processing of small amounts of FFPE tissue samples for proteomic analysis. We identified 3,525 proteins from six 10µm esophageal squamous cell carcinoma (ESCC tissue sections. Barocycler allows efficient protein extraction and proteolytic digestion of proteins from FFPE tissue sections at par with conventional methods.

  2. Expression of matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-2 in radiation exposed small intestinal mucosa of the rat

    International Nuclear Information System (INIS)

    Kwag, Hyon Joo; Lee, Kyoung Ja; Rhee, Chung Sik

    2003-01-01

    The matrix metalloproteinases (MMPs) are a family of enzymes whose main function is the degradation of the extracellular matrix. Several studies have revealed that MMPs and TIMPs are related to the wound healing process and in photoaging caused by ultraviolet irradiation. However, the expressions of MMP and TIMP after irradiation have not, to the best of our knowledge, been studied. This study investigates the expressions of MMP-2 and TIMP-2 in rat intestinal mucosa following irradiation. The entire abdomen of Sprague-Dawley rats was irradiated using a single dose method. The rats were sacrificed on day 1, 2, 3, 5, 7 and 14 following irradiation. Histopathological observations were made using hematoxilin and eosin staining. The expressions of MMP-2 and TIMP-2 were examined using immunohistochemistry, immunoblotting and ELISA. Radiation induced damage, associated with atrophic villi, and infiltration of inflammatory cells was observed from the first postirradiation day, and severe tissue damage was observed on the second and the third postirradiation days. An increase in mitosis and the number of regenerating crypts, as evidence of regeneration, were most noticeable on the fifth postirradiation day. From the immunohistochemistry, the MMP-2 expression was observed from the first postirradiation day, but was most conspicuous on the third and the fifth postirradiation days. The TIMP-2 expression was most conspicuous on the fifth postirradiation day. From the immunoblotting, the MMP-2 expression was strongly positive on the third postirradiation day, and that of TIMP-2 showed a strong positive response on the fifth postirradiation day. In ELISA, tests, the expressions of MMP-2 and TIMP-2. were increased in the postirradiation groups compared to those of the normal controls, and showed a maximum increase on the fifth postirradiation day. These results were statistically significant. The expressions of MMP-2 and TIMP-2 were increased in the intestinal mucosa of the rats

  3. GeLC-MS: A Sample Preparation Method for Proteomics Analysis of Minimal Amount of Tissue.

    Science.gov (United States)

    Makridakis, Manousos; Vlahou, Antonia

    2017-10-10

    Application of various proteomics methodologies have been implemented for the global and targeted proteome analysis of many different types of biological samples such as tissue, urine, plasma, serum, blood, and cell lines. Among the aforementioned biological samples, tissue has an exceptional role into clinical research and practice. Disease initiation and progression is usually located at the tissue level of different organs, making the analysis of this material very important for the understanding of the disease pathophysiology. Despite the significant advances in the mass spectrometry instrumentation, tissue proteomics still faces several challenges mainly due to increased sample complexity and heterogeneity. However, the most prominent challenge is attributed to the invasive procedure of tissue sampling which restricts the availability of fresh frozen tissue to minimal amounts and limited number of samples. Application of GeLC-MS sample preparation protocol for tissue proteomics analysis can greatly facilitate making up for these difficulties. In this chapter, a step by step guide for the proteomics analysis of minute amounts of tissue samples using the GeLC-MS sample preparation protocol, as applied by our group in the analysis of multiple different types of tissues (vessels, kidney, bladder, prostate, heart) is provided.

  4. Growth and parameters of microflora in intestinal and faecal samples of piglets due to application of a phytogenic feed additive.

    Science.gov (United States)

    Muhl, A; Liebert, F

    2007-10-01

    A commercial phytogenic feed additive (PFA), containing the fructopolysaccharide inulin, an essential oil mix (carvacrol, thymol), chestnut meal (tannins) and cellulose powder as carrier substance, was examined for effects on growth and faecal and intestinal microflora of piglets. Two experiments (35 days) were conducted, each with 40 male castrated weaned piglets. In experiment 1, graded levels of the PFA were supplied (A1: control; B1: 0.05% PFA; C1: 0.1% PFA; D1: 0.15% PFA) in diets based on wheat, barley, soybean meal and fish meal with lysine as the limiting amino acid. In experiment 2, a similar diet with 0.1% of the PFA (A2: control; B2: 0.1% PFA; C2: +0.35% lysine; D2: 0.1% PFA + 0.35% lysine) and lysine supplementation was utilized. During experiment 1, no significant effect of the PFA on growth, feed intake and feed conversion rate was observed (p > 0.05). Lysine supplementation in experiment 2 improved growth performance significantly, but no significant effect of the PFA was detected. Microbial counts in faeces (aerobes, Gram negatives, anaerobes and lactobacilli) during the first and fifth week did not indicate any significant PFA effect (p > 0.05). In addition, microflora in intestinal samples was not significantly modified by supplementing the PFA (p > 0.05). Lysine supplementation indicated lysine as limiting amino acid in the basal diet, but did not influence the microbial counts in faeces and small intestine respectively.

  5. Versatile electrochemial sensor for tissue culturing and sample handling

    DEFF Research Database (Denmark)

    Bakmand, Tanya; Kwasny, Dorota; Al Atraktchi, Fatima Al-Zahraa

    2014-01-01

    in microfluidic devices for sample preparation. In this work we present the development of the sensor system along with results on characterization by impedance spectroscopy and cyclic voltammetry. Furthermore we present recent results on integration of the sensor as well as amperometric detection of dopamine...

  6. Final LDRD report : development of sample preparation methods for ChIPMA-based imaging mass spectrometry of tissue samples.

    Energy Technology Data Exchange (ETDEWEB)

    Maharrey, Sean P.; Highley, Aaron M.; Behrens, Richard, Jr.; Wiese-Smith, Deneille

    2007-12-01

    The objective of this short-term LDRD project was to acquire the tools needed to use our chemical imaging precision mass analyzer (ChIPMA) instrument to analyze tissue samples. This effort was an outgrowth of discussions with oncologists on the need to find the cellular origin of signals in mass spectra of serum samples, which provide biomarkers for ovarian cancer. The ultimate goal would be to collect chemical images of biopsy samples allowing the chemical images of diseased and nondiseased sections of a sample to be compared. The equipment needed to prepare tissue samples have been acquired and built. This equipment includes an cyro-ultramicrotome for preparing thin sections of samples and a coating unit. The coating unit uses an electrospray system to deposit small droplets of a UV-photo absorbing compound on the surface of the tissue samples. Both units are operational. The tissue sample must be coated with the organic compound to enable matrix assisted laser desorption/ionization (MALDI) and matrix enhanced secondary ion mass spectrometry (ME-SIMS) measurements with the ChIPMA instrument Initial plans to test the sample preparation using human tissue samples required development of administrative procedures beyond the scope of this LDRD. Hence, it was decided to make two types of measurements: (1) Testing the spatial resolution of ME-SIMS by preparing a substrate coated with a mixture of an organic matrix and a bio standard and etching a defined pattern in the coating using a liquid metal ion beam, and (2) preparing and imaging C. elegans worms. Difficulties arose in sectioning the C. elegans for analysis and funds and time to overcome these difficulties were not available in this project. The facilities are now available for preparing biological samples for analysis with the ChIPMA instrument. Some further investment of time and resources in sample preparation should make this a useful tool for chemical imaging applications.

  7. Cytokine gene expression profiles in chicken spleen and intestinal tissues during Ascaridia galli infection

    DEFF Research Database (Denmark)

    Pleidrup, Janne A.; Norup, Liselotte R.; Dalgaard, Tina S.

    2014-01-01

    , the anti-inflammatory cytokines IL-10 and TGF-beta 4 and the proinflammatory cytokine IL-17F were determined over a period of 3 weeks in A. galli and non-A. galli-infected chickens. A characteristic Th2 response was observed in the jejunum of A. galli-infected chickens with increased expression of IL-13...... and decreased expression of IFN-gamma from day 14 post infection. At the putative time of larvae invasion into the intestinal mucosa (day 7), an increased expression of IFN-gamma, IL-10, and TGF-beta 4 was observed in the spleen. At the putative onset of the innate immune response (day 10), a decreased...... expression of jejunal IFN-gamma and IL-13 was observed. Finally, at the expected period of an adaptive immune response (days 14-21) a general decreased expression of IFN-gamma and TGF-beta 4 in spleen and IFN-gamma in jejunum was followed by a decreased expression of IFN-gamma and IL-10 at day 21 in caecal...

  8. Handbook of Human Tissue Sources. A National Resource of Human Tissue Samples

    Science.gov (United States)

    1999-01-01

    assess exposure to infectious and toxic agents during deployment, to estimate immunity prevalence for a variety of diseases, and to determine the...the Missouri Lions Eye Tissue Banks in three Missouri cities, Glaucoma Screening, Eyeglass Recycling, Amblyopia Screening, Indigent Patient Care, and

  9. Elemental distribution and sample integrity comparison of freeze-dried and frozen-hydrated biological tissue samples with nuclear microprobe

    Energy Technology Data Exchange (ETDEWEB)

    Vavpetič, P., E-mail: primoz.vavpetic@ijs.si [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Vogel-Mikuš, K. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Jeromel, L. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Ogrinc Potočnik, N. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); FOM-Institute AMOLF, Science Park 104, 1098 XG Amsterdam (Netherlands); Pongrac, P. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Department of Plant Physiology, University of Bayreuth, Universitätstr. 30, 95447 Bayreuth (Germany); Drobne, D.; Pipan Tkalec, Ž.; Novak, S.; Kos, M.; Koren, Š.; Regvar, M. [Biotechnical Faculty, Department of Biology, University of Ljubljana, Jamnikarjeva 101, SI-1000 Ljubljana (Slovenia); Pelicon, P. [Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia)

    2015-04-01

    The analysis of biological samples in frozen-hydrated state with micro-PIXE technique at Jožef Stefan Institute (JSI) nuclear microprobe has matured to a point that enables us to measure and examine frozen tissue samples routinely as a standard research method. Cryotome-cut slice of frozen-hydrated biological sample is mounted between two thin foils and positioned on the sample holder. The temperature of the cold stage in the measuring chamber is kept below 130 K throughout the insertion of the samples and the proton beam exposure. Matrix composition of frozen-hydrated tissue is consisted mostly of ice. Sample deterioration during proton beam exposure is monitored during the experiment, as both Elastic Backscattering Spectrometry (EBS) and Scanning Transmission Ion Microscopy (STIM) in on–off axis geometry are recorded together with the events in two PIXE detectors and backscattered ions from the chopper in a single list-mode file. The aim of this experiment was to determine differences and similarities between two kinds of biological sample preparation techniques for micro-PIXE analysis, namely freeze-drying and frozen-hydrated sample preparation in order to evaluate the improvements in the elemental localisation of the latter technique if any. In the presented work, a standard micro-PIXE configuration for tissue mapping at JSI was used with five detection systems operating in parallel, with proton beam cross section of 1.0 × 1.0 μm{sup 2} and a beam current of 100 pA. The comparison of the resulting elemental distributions measured at the biological tissue prepared in the frozen-hydrated and in the freeze-dried state revealed differences in elemental distribution of particular elements at the cellular level due to the morphology alteration in particular tissue compartments induced either by water removal in the lyophilisation process or by unsatisfactory preparation of samples for cutting and mounting during the shock-freezing phase of sample preparation.

  10. Elemental distribution and sample integrity comparison of freeze-dried and frozen-hydrated biological tissue samples with nuclear microprobe

    International Nuclear Information System (INIS)

    Vavpetič, P.; Vogel-Mikuš, K.; Jeromel, L.; Ogrinc Potočnik, N.; Pongrac, P.; Drobne, D.; Pipan Tkalec, Ž.; Novak, S.; Kos, M.; Koren, Š.; Regvar, M.; Pelicon, P.

    2015-01-01

    The analysis of biological samples in frozen-hydrated state with micro-PIXE technique at Jožef Stefan Institute (JSI) nuclear microprobe has matured to a point that enables us to measure and examine frozen tissue samples routinely as a standard research method. Cryotome-cut slice of frozen-hydrated biological sample is mounted between two thin foils and positioned on the sample holder. The temperature of the cold stage in the measuring chamber is kept below 130 K throughout the insertion of the samples and the proton beam exposure. Matrix composition of frozen-hydrated tissue is consisted mostly of ice. Sample deterioration during proton beam exposure is monitored during the experiment, as both Elastic Backscattering Spectrometry (EBS) and Scanning Transmission Ion Microscopy (STIM) in on–off axis geometry are recorded together with the events in two PIXE detectors and backscattered ions from the chopper in a single list-mode file. The aim of this experiment was to determine differences and similarities between two kinds of biological sample preparation techniques for micro-PIXE analysis, namely freeze-drying and frozen-hydrated sample preparation in order to evaluate the improvements in the elemental localisation of the latter technique if any. In the presented work, a standard micro-PIXE configuration for tissue mapping at JSI was used with five detection systems operating in parallel, with proton beam cross section of 1.0 × 1.0 μm 2 and a beam current of 100 pA. The comparison of the resulting elemental distributions measured at the biological tissue prepared in the frozen-hydrated and in the freeze-dried state revealed differences in elemental distribution of particular elements at the cellular level due to the morphology alteration in particular tissue compartments induced either by water removal in the lyophilisation process or by unsatisfactory preparation of samples for cutting and mounting during the shock-freezing phase of sample preparation

  11. In vivo imaging of enteric neurogenesis in the deep tissue of mouse small intestine.

    Science.gov (United States)

    Goto, Kei; Kato, Go; Kawahara, Isao; Luo, Yi; Obata, Koji; Misawa, Hiromi; Ishikawa, Tatsuya; Kuniyasu, Hiroki; Nabekura, Junich; Takaki, Miyako

    2013-01-01

    One of the challenges of using imaging techniques as a tool to study cellular physiology has been the inability to resolve structures that are not located near the surface of the preparation. Nonlinear optical microscopy, in particular two photon-excited fluorescence microscopy (2PM), has overcome this limitation, providing deeper optical penetration (several hundred µm) in ex vivo and in vivo preparations. We have used this approach in the gut to achieve the first in vivo imaging of enteric neurons and nerve fibers in the mucosa, submucosa, submucosal and myenteric plexuses, and circular and longitudinal muscles of the small intestine in H-line: Thy1 promoter GFP mice. Moreover, we obtained clear three-dimensional imaging of enteric neurons that were newly generated after gut transection and reanastomosis. Neurogenesis was promoted by oral application of the 5-HT(4)-receptor agonist, mosapride citrate (MOS). The number of newly generated neurons observed in mice treated with MOS for one week was 421±89 per 864,900 µm(2) (n = 5), which was significantly greater than that observed in preparations treated with MOS plus an antagonist (113±76 per 864,900 µm(2)) or in 4 week vehicle controls (100±34 per 864,900 µm(2)) (n = 4 both). Most neurons were located within 100 µm of the surface. These results confirm that activation of enteric neural 5-HT(4)-receptor by MOS promotes formation of new enteric neurons. We conclude that in vivo 2PM imaging made it possible to perform high-resolution deep imaging of the living mouse whole gut and reveal formation of new enteric neurons promoted by 5-HT(4)-receptor activation.

  12. In vivo imaging of enteric neurogenesis in the deep tissue of mouse small intestine.

    Directory of Open Access Journals (Sweden)

    Kei Goto

    Full Text Available One of the challenges of using imaging techniques as a tool to study cellular physiology has been the inability to resolve structures that are not located near the surface of the preparation. Nonlinear optical microscopy, in particular two photon-excited fluorescence microscopy (2PM, has overcome this limitation, providing deeper optical penetration (several hundred µm in ex vivo and in vivo preparations. We have used this approach in the gut to achieve the first in vivo imaging of enteric neurons and nerve fibers in the mucosa, submucosa, submucosal and myenteric plexuses, and circular and longitudinal muscles of the small intestine in H-line: Thy1 promoter GFP mice. Moreover, we obtained clear three-dimensional imaging of enteric neurons that were newly generated after gut transection and reanastomosis. Neurogenesis was promoted by oral application of the 5-HT(4-receptor agonist, mosapride citrate (MOS. The number of newly generated neurons observed in mice treated with MOS for one week was 421±89 per 864,900 µm(2 (n = 5, which was significantly greater than that observed in preparations treated with MOS plus an antagonist (113±76 per 864,900 µm(2 or in 4 week vehicle controls (100±34 per 864,900 µm(2 (n = 4 both. Most neurons were located within 100 µm of the surface. These results confirm that activation of enteric neural 5-HT(4-receptor by MOS promotes formation of new enteric neurons. We conclude that in vivo 2PM imaging made it possible to perform high-resolution deep imaging of the living mouse whole gut and reveal formation of new enteric neurons promoted by 5-HT(4-receptor activation.

  13. Tissue levels and post-prandial secretion of the intestinal growth factor, glucagon-like peptide-2, in controls and inflammatory bowel disease

    DEFF Research Database (Denmark)

    Schmidt, Peter T; Ljung, Tryggve; Hartmann, Bolette

    2005-01-01

    BACKGROUND AND AIM: Glucagon-like peptide-2 (GLP-2) and peptide YY (PYY) are produced in endocrine L-cells of the intestine and secreted in response to food intake. GLP-2 has a trophic effect on the intestinal epithelium, whereas PYY has pro-absorptive effects. It can be speculated that...... the fasting plasma levels nor the meal responses of GLP-2 and PYY differed between controls and IBD patients. CONCLUSION: The similar responses of GLP-2 and PYY in patients and controls do not support the suggestion that L-cell secretion is altered in IBD. The decreased tissue PYY concentrations may...

  14. Intestinal transport and tissue biodistributions of two radium species using variations in dietary calcium

    International Nuclear Information System (INIS)

    Domel, R.U.; Beal, A.M.

    1996-01-01

    Full text: Natural leaching of uranium ore bodies can result in 226 Ra pollution of adjacent waterways and consequent incorporation of radium into the food chain. In the Magela flood plain, NT, the freshwater mussel (Velesunio angasi) concentrates radium in its tissues as a phosphate compound. Calcium has been demonstrated to be a competitive cation and thus influence the uptake and biodistribution of radium. In this set of experiments the influence of three levels of dietary calcium (0%, 1 .15% and 6. 15%) were compared in adult and juvenile rats at the end of one week feed trials. Increasing dietary calcium from 1. 15% to 6. 15% in combination with radium chloride dosage reduced radium uptake by bone in juveniles and adults. In contrast, juveniles fed mussel radium had increased radium uptake by bone. The increased combination with mussel radium was without effect in adults. For radium chloride but not mussel radium, the radium uptake by soft tissues was higher in juvenile rats fed with increased dietary calcium. Reducing calcium diet from 1.15% to 0% increased radium uptake by bone from radium chloride in both age groups and also uptake by bone from mussel radium in juveniles, whereas the effect on bone uptake in adults was not significant. The juveniles also obtained a higher radium dose in the soft tissues from the radium chloride but not the mussel radium treatment with the calcium deficient diet. Extrapolation of the data from the animal model to humans suggests that eating these mussels in the absence or presence of dietary calcium, carries with it only a low risk of exceeding the Annual Limit of Intake (ALI) set for members of the public, even in juveniles

  15. The application of fluorescence in situ hybridization (FISH technique for studying the microbial communities in intestinal tissues of white shrimp (Penaeus vannamei

    Directory of Open Access Journals (Sweden)

    Supamattaya, K.

    2005-02-01

    Full Text Available Fluorescence in situ hybridization technique is very useful for the evaluation of microbial communities in various environments. It is possible to apply this technique to study the intestinal microflora in white shrimp (Penaeus vannamei. Different fixatives and storage temperature were tested in this technique. It was found that fixation with 10% buffered formalin for 12 hours and changed to 70% ethanol shown positive results when compared to the fixation with Davidson's fixative or RF fixative. The best signaling was obtainedfrom the samples which were stored in -20ºC. By using the DNA probe targeted to the Eubacteria domain (EUB338 probe, 5′-GCT GCC TCC CGT AGG AGT-3′ labeled with fluorescein as a hybridizing probe, it was found that most intestinal microflora were aggregated with the intestinal contents, or dispersed in the lumen. There was not evidence of the attachment of the microflora with the intestinal epithelium in this study.

  16. Tissue Microarray Technology for Molecular Applications: Investigation of Cross-Contamination between Tissue Samples Obtained from the Same Punching Device

    Directory of Open Access Journals (Sweden)

    Erik Vassella

    2015-04-01

    Full Text Available Background: Tissue microarray (TMA technology allows rapid visualization of molecular markers by immunohistochemistry and in situ hybridization. In addition, TMA instrumentation has the potential to assist in other applications: punches taken from donor blocks can be placed directly into tubes and used for nucleic acid analysis by PCR approaches. However, the question of possible cross-contamination between samples punched with the same device has frequently been raised but never addressed. Methods: Two experiments were performed. (1 A block from mycobacterium tuberculosis (TB positive tissue and a second from an uninfected patient were aligned side-by-side in an automated tissue microarrayer. Four 0.6 mm punches were cored from each sample and placed inside their corresponding tube. Between coring of each donor block, a mechanical cleaning step was performed by insertion of the puncher into a paraffin block. This sequence of coring and cleaning was repeated three times, alternating between positive and negative blocks. A fragment from the 6110 insertion sequence specific for mycobacterium tuberculosis was analyzed; (2 Four 0.6 mm punches were cored from three KRAS mutated colorectal cancer blocks, alternating with three different wild-type tissues using the same TMA instrument (sequence of coring: G12D, WT, G12V, WT, G13D and WT. Mechanical cleaning of the device between each donor block was made. Mutation analysis by pyrosequencing was carried out. This sequence of coring was repeated manually without any cleaning step between blocks. Results/Discussion: In both analyses, all alternating samples showed the expected result (samples 1, 3 and 5: positive or mutated, samples 2, 4 and 6: negative or wild-type. Similar results were obtained without cleaning step. These findings suggest that no cross-contamination of tissue samples occurs when donor blocks are punched using the same device, however a cleaning step is nonetheless recommended. Our

  17. Ecotoxicoparasitology: Understanding mercury concentrations in gut contents, intestinal helminths and host tissues of Alaskan gray wolves (Canis lupus)

    Energy Technology Data Exchange (ETDEWEB)

    McGrew, Ashley K. [Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80523-1619 (United States); O' Hara, Todd M. [Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80523-1619 (United States); Wildlife Toxicology Laboratory, Department of Veterinary Medicine, University of Alaska Fairbanks, Fairbanks, AK 99775 (United States); Stricker, Craig A. [U. S. Geological Survey, Fort Collins Science Center, Denver, CO 80225 (United States); Margaret Castellini, J. [Wildlife Toxicology Laboratory, Department of Veterinary Medicine, University of Alaska Fairbanks, Fairbanks, AK 99775 (United States); Beckmen, Kimberlee B. [Alaska Department of Fish & Game, Fairbanks, AK (United States); Salman, Mo D. [Animal Population Health Institute, Department of Clinical Sciences, Colorado State University, Fort Collins, CO 80523-1644 (United States); Ballweber, Lora R. [Department of Microbiology, Immunology, and Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, CO 80523-1619 (United States)

    2015-12-01

    Some gastrointestinal helminths acquire nutrients from the lumen contents in which they live; thus, they may be exposed to non-essential elements, such as mercury (Hg), during feeding. The objectives of this study were: 1) determine the total mercury concentrations ([THg]) in Gray wolves (Canis lupus) and their parasites, and 2) use stable isotopes to evaluate the trophic relationships within the host. [THg] and stable isotopes (C and N) were determined for helminths, host tissues, and lumen contents from 88 wolves. Sixty-three wolves contained grossly visible helminths (71.5%). The prevalence of taeniids and ascarids was 63.6% (56/88) and 20.5% (18/88), respectively. Nine of these 63 wolves contained both taeniids and ascarids (14.3%). All ascarids were determined to be Toxascaris leonina. Taenia species present included T. krabbei and T. hydatigena. Within the GI tract, [THg] in the lumen contents of the proximal small intestine were significantly lower than in the distal small intestine. There was a significant positive association between hepatic and taeniid [THg]. Bioaccumulation factors (BAF) ranged from < 1 to 22.9 in taeniids, and 1.1 to 12.3 in T. leonina. Taeniid and ascarid BAF were significantly higher than 1, suggesting that both groups are capable of THg accumulation in their wolf host. δ{sup 13}C in taeniids was significantly lower than in host liver and skeletal muscle. [THg] in helminths and host tissues, in conjunction with stable isotope (C and N) values, provides insight into food-web dynamics of the host GI tract, and aids in elucidating ecotoxicoparasitologic relationships. Variation of [THg] throughout the GI tract, and between parasitic groups, underscores the need to further evaluate the effect(s) of feeding niche, and the nutritional needs of parasites, as they relate to toxicant exposure and distribution within the host. - Highlights: • [THg] and stable isotopes together provide insight on host-parasite-Hg interactions. • A

  18. Comparison of diagnostic efficacy between CLE, tissue sampling, and CLE combined with tissue sampling for undetermined pancreaticobiliary strictures: a meta-analysis.

    Science.gov (United States)

    Gao, Ya-Dong; Qu, Ya-Wei; Liu, Hai-Feng

    2018-04-01

    The accurate diagnosis of undetermined pancreaticobiliary strictures remains challenging. Current ERCP-guided tissue sampling methods are of low sensitivity. Confocal laser endomicroscopy (CLE) is a new procedure and allows real optical biopsies that may improve the diagnosis of undetermined pancreaticobiliary strictures. The aim of this meta-analysis was to determine the diagnostic yield of CLE, tissue sampling, and CLE combined with tissue sampling for undetermined pancreaticobiliary strictures. Pubmed, Embase, and the Cochrane Library database were reviewed for relevant studies. Pooled estimates of sensitivity and specificity with 95% confidence intervals (CIs) were calculated using the random-effects meta-analysis model. The summary receiver-operating characteristic (SROC) curve was constructed, and the area under the receiver operating characteristic curve (AUC) was calculated. Twelve studies involving 591 patients were enrolled in our analysis. The overall sensitivity and the specificity estimate of CLE for discriminating benign and malignant pancreaticobiliary strictures were 87% (95%CI, 83-91%) and 76% (95%CI, 70-81%), respectively. The AUC to assess the diagnostic efficacy was 0.8705. For tissue sampling, the overall sensitivity and the specificity estimate were 64% (95%CI, 57-70%) and 94% (95%CI, 90-97%), respectively. The AUC to assess the diagnostic efficacy was 0.8040. A combination of both methods increased the sensitivity (93%; 95%CI, 88-96%) with a specificity of 82% (95%CI, 74-89%). The AUC to assess the diagnostic efficacy was 0.9377. There was no publication bias by Deeks' Funnel Plot with p = .936. Compared with tissue sampling, CLE may increase the sensitivity for the diagnosis of malignant pancreaticobiliary strictures. A combination of both can effectively diagnose malignant pancreaticobiliary strictures.

  19. Effects of Re-heating Tissue Samples to Core Body Temperature on High-Velocity Ballistic Projectile-tissue Interactions.

    Science.gov (United States)

    Humphrey, Caitlin; Henneberg, Maciej; Wachsberger, Christian; Maiden, Nicholas; Kumaratilake, Jaliya

    2017-11-01

    Damage produced by high-speed projectiles on organic tissue will depend on the physical properties of the tissues. Conditioning organic tissue samples to human core body temperature (37°C) prior to conducting ballistic experiments enables their behavior to closely mimic that of living tissues. To minimize autolytic changes after death, the tissues are refrigerated soon after their removal from the body and re-heated to 37°C prior to testing. This research investigates whether heating 50-mm-cube samples of porcine liver, kidney, and heart to 37°C for varying durations (maximum 7 h) can affect the penetration response of a high-speed, steel sphere projectile. Longer conditioning times for heart and liver resulted in a slight loss of velocity/energy of the projectile, but the reverse effect occurred for the kidney. Possible reasons for these trends include autolytic changes causing softening (heart and liver) and dehydration causing an increase in density (kidney). © 2017 American Academy of Forensic Sciences.

  20. Differences between intestinal segments and soybean meal-induced changes in intestinal mucus composition of common carp Cyprinus carpio L.

    NARCIS (Netherlands)

    Marel, van der M.C.; Propsting, M.J.; Battermann, F.; Jung-Schroers, V.; Hubner, A.; Rombout, J.H.W.M.; Steinhagen, D.

    2014-01-01

    The alimentary tract is a possible site where pathogens and toxins can enter. The alimentary tract is protected, amongst others, by mucus. In this study, tissue samples and crude mucus preparations from different parts of the intestinal tract of Cyprinus carpio (from intestinal bulb onto the

  1. A stromal cell population in the large intestine identified by tissue factor expression that is lost during colorectal cancer progression.

    Science.gov (United States)

    Eriksson, Oskar; Asplund, Anna; Hegde, Geeta; Edqvist, Per-Henrik; Navani, Sanjay; Pontén, Fredrik; Siegbahn, Agneta

    2016-11-30

    Colorectal cancer (CRC) is a major cause of morbidity and mortality, and the composition of the tumour stroma is a strong predictor of survival in this cancer type. Tissue factor (TF) functions as the trigger of haemostasis together with its ligand coagulation factor VII/VIIa, and TF expression has been found in tumour cells of colorectal tumours. However, TF expression in the CRC tumour stroma or its relationship to patient outcome has not yet been studied. To address this question we developed and validated a specific anti-TF antibody using standardised methods within the Human Protein Atlas project. We used this antibody to investigate TF expression in normal colorectal tissue and CRC using immunofluorescence and immunohistochemistry in two patient cohorts. TF was strongly expressed in a cell population immediately adjacent to the colorectal epithelium. These TF-positive cells were ACTA2-negative but weakly vimentin-positive, defining a specific population of pericryptal sheath cells. In colorectal tumours, TF-positive sheath cells were progressively lost after the adenoma-to-carcinoma transition, demonstrating downregulation of this source of TF in CRC. Furthermore, loss of sheath cell TF was significantly associated with poor overall and disease-specific survival in rectal but not colon cancers. In conclusion, we demonstrate that TF is a marker of a specific cell population in the large intestine, which is lost during CRC progression. Our results highlight the role of the tumour stroma in this cancer type and suggest TF to be a potential prognostic biomarker in rectal cancers through the identification of pericryptal sheath cells.

  2. Increased Expression of Peripheral Blood Leukocyte Genes Implicate CD14+ Tissue Macrophages in Cellular Intestine Allograft Rejection

    Science.gov (United States)

    Ashokkumar, Chethan; Ningappa, Mylarappa; Ranganathan, Sarangarajan; Higgs, Brandon W.; Sun, Qing; Schmitt, Lori; Snyder, Sara; Dobberstein, Jennifer; Branca, Maria; Jaffe, Ronald; Zeevi, Adriana; Squires, Robert; Alissa, Feras; Shneider, Benjamin; Soltys, Kyle; Bond, Geoffrey; Abu-Elmagd, Kareem; Humar, Abhinav; Mazariegos, George; Hakonarson, Hakon; Sindhi, Rakesh

    2011-01-01

    Recurrent rejection shortens graft survival after intestinal transplantation (ITx) in children, most of whom also experience early acute cellular rejection (rejectors). To elucidate mechanisms common to early and recurrent rejection, we used a test cohort of 20 recipients to test the hypothesis that candidate peripheral blood leukocyte genes that trigger rejection episodes would be evident late after ITx during quiescent periods in genome-wide gene expression analysis and would achieve quantitative real-time PCR replication pre-ITx (another quiescent period) and in the early post-ITx period during first rejection episodes. Eight genes were significantly up-regulated among rejectors in the late post-ITx and pre-ITx periods, compared with nonrejectors: TBX21, CCL5, GNLY, SLAMF7, TGFBR3, NKG7, SYNE1, and GK5. Only CCL5 was also up-regulated in the early post-ITx period. Among resting peripheral blood leukocyte subsets in randomly sampled nonrejectors, CD14+ monocytes expressed the CCL5 protein maximally. Compared with nonrejectors, rejectors demonstrated higher counts of both circulating CCL5+CD14+ monocytes and intragraft CD14+ monocyte-derived macrophages in immunohistochemistry of postperfusion and early post-ITx biopsies from the test and an independent replication cohort. Donor-specific alloreactivity measured with CD154+ T-cytotoxic memory cells correlated with the CCL5 gene and intragraft CD14+ monocyte-derived macrophages at graft reperfusion and early post-ITx. CCL5 gene up-regulation and CD14+ macrophages likely prime cellular ITx rejection. Infiltration of reperfused intestine allografts with CD14+ macrophages may predict rejection events. PMID:21854741

  3. Fermentation of mucin by bifidobacteria from rectal samples of humans and rectal and intestinal samples of animals

    Czech Academy of Sciences Publication Activity Database

    Killer, Jiří; Marounek, Milan

    2011-01-01

    Roč. 56, č. 2 (2011), s. 85-89 ISSN 0015-5632 Institutional research plan: CEZ:AV0Z50450515 Keywords : mucin * bifidobacteria * rectal samples Subject RIV: EE - Microbiology, Virology Impact factor: 0.677, year: 2011

  4. A method for high purity intestinal epithelial cell culture from adult human and murine tissues for the investigation of innate immune function.

    Science.gov (United States)

    Graves, Christina L; Harden, Scott W; LaPato, Melissa; Nelson, Michael; Amador, Byron; Sorenson, Heather; Frazier, Charles J; Wallet, Shannon M

    2014-12-01

    Intestinal epithelial cells (IECs) serve as an important physiologic barrier between environmental antigens and the host intestinal immune system. Thus, IECs serve as a first line of defense and may act as sentinel cells during inflammatory insults. Despite recent renewed interest in IEC contributions to host immune function, the study of primary IEC has been hindered by lack of a robust culture technique, particularly for small intestinal and adult tissues. Here, a novel adaptation for culture of primary IEC is described for human duodenal organ donor tissue as well as duodenum and colon of adult mice. These epithelial cell cultures display characteristic phenotypes and are of high purity. In addition, the innate immune function of human primary IEC, specifically with regard to Toll-like receptor (TLR) expression and microbial ligand responsiveness, is contrasted with a commonly used intestinal epithelial cell line (HT-29). Specifically, TLR expression at the mRNA level and production of cytokine (IFNγ and TNFα) in response to TLR agonist stimulation is assessed. Differential expression of TLRs as well as innate immune responses to ligand stimulation is observed in human-derived cultures compared to that of HT-29. Thus, use of this adapted method to culture primary epithelial cells from adult human donors and from adult mice will allow for more appropriate studies of IECs as innate immune effectors. Published by Elsevier B.V.

  5. Culture methods of allograft musculoskeletal tissue samples in Australian bacteriology laboratories.

    Science.gov (United States)

    Varettas, Kerry

    2013-12-01

    Samples of allograft musculoskeletal tissue are cultured by bacteriology laboratories to determine the presence of bacteria and fungi. In Australia, this testing is performed by 6 TGA-licensed clinical bacteriology laboratories with samples received from 10 tissue banks. Culture methods of swab and tissue samples employ a combination of solid agar and/or broth media to enhance micro-organism growth and maximise recovery. All six Australian laboratories receive Amies transport swabs and, except for one laboratory, a corresponding biopsy sample for testing. Three of the 6 laboratories culture at least one allograft sample directly onto solid agar. Only one laboratory did not use a broth culture for any sample received. An international literature review found that a similar combination of musculoskeletal tissue samples were cultured onto solid agar and/or broth media. Although variations of allograft musculoskeletal tissue samples, culture media and methods are used in Australian and international bacteriology laboratories, validation studies and method evaluations have challenged and supported their use in recovering fungi and aerobic and anaerobic bacteria.

  6. Giardia duodenalis infection reduces granulocyte infiltration in an in vivo model of bacterial toxin-induced colitis and attenuates inflammation in human intestinal tissue.

    Science.gov (United States)

    Cotton, James A; Motta, Jean-Paul; Schenck, L Patrick; Hirota, Simon A; Beck, Paul L; Buret, Andre G

    2014-01-01

    Giardia duodenalis (syn. G. intestinalis, G. lamblia) is a predominant cause of waterborne diarrheal disease that may lead to post-infectious functional gastrointestinal disorders. Although Giardia-infected individuals could carry as much as 106 trophozoites per centimetre of gut, their intestinal mucosa is devoid of overt signs of inflammation. Recent studies have shown that in endemic countries where bacterial infectious diseases are common, Giardia infections can protect against the development of diarrheal disease and fever. Conversely, separate observations have indicated Giardia infections may enhance the severity of diarrheal disease from a co-infecting pathogen. Polymorphonuclear leukocytes or neutrophils (PMNs) are granulocytic, innate immune cells characteristic of acute intestinal inflammatory responses against bacterial pathogens that contribute to the development of diarrheal disease following recruitment into intestinal tissues. Giardia cathepsin B cysteine proteases have been shown to attenuate PMN chemotaxis towards IL-8/CXCL8, suggesting Giardia targets PMN accumulation. However, the ability of Giardia infections to attenuate PMN accumulation in vivo and how in turn this effect may alter the host inflammatory response in the intestine has yet to be demonstrated. Herein, we report that Giardia infection attenuates granulocyte tissue infiltration induced by intra-rectal instillation of Clostridium difficile toxin A and B in an isolate-dependent manner. This attenuation of granulocyte infiltration into colonic tissues paralled decreased expression of several cytokines associated with the recruitment of PMNs. Giardia trophozoite isolates that attenuated granulocyte infiltration in vivo also decreased protein expression of cytokines released from inflamed mucosal biopsy tissues collected from patients with active Crohn's disease, including several cytokines associated with PMN recruitment. These results demonstrate for the first time that certain

  7. Giardia duodenalis infection reduces granulocyte infiltration in an in vivo model of bacterial toxin-induced colitis and attenuates inflammation in human intestinal tissue.

    Directory of Open Access Journals (Sweden)

    James A Cotton

    Full Text Available Giardia duodenalis (syn. G. intestinalis, G. lamblia is a predominant cause of waterborne diarrheal disease that may lead to post-infectious functional gastrointestinal disorders. Although Giardia-infected individuals could carry as much as 106 trophozoites per centimetre of gut, their intestinal mucosa is devoid of overt signs of inflammation. Recent studies have shown that in endemic countries where bacterial infectious diseases are common, Giardia infections can protect against the development of diarrheal disease and fever. Conversely, separate observations have indicated Giardia infections may enhance the severity of diarrheal disease from a co-infecting pathogen. Polymorphonuclear leukocytes or neutrophils (PMNs are granulocytic, innate immune cells characteristic of acute intestinal inflammatory responses against bacterial pathogens that contribute to the development of diarrheal disease following recruitment into intestinal tissues. Giardia cathepsin B cysteine proteases have been shown to attenuate PMN chemotaxis towards IL-8/CXCL8, suggesting Giardia targets PMN accumulation. However, the ability of Giardia infections to attenuate PMN accumulation in vivo and how in turn this effect may alter the host inflammatory response in the intestine has yet to be demonstrated. Herein, we report that Giardia infection attenuates granulocyte tissue infiltration induced by intra-rectal instillation of Clostridium difficile toxin A and B in an isolate-dependent manner. This attenuation of granulocyte infiltration into colonic tissues paralled decreased expression of several cytokines associated with the recruitment of PMNs. Giardia trophozoite isolates that attenuated granulocyte infiltration in vivo also decreased protein expression of cytokines released from inflamed mucosal biopsy tissues collected from patients with active Crohn's disease, including several cytokines associated with PMN recruitment. These results demonstrate for the first time

  8. High-throughput simultaneous analysis of RNA, protein, and lipid biomarkers in heterogeneous tissue samples.

    Science.gov (United States)

    Reiser, Vladimír; Smith, Ryan C; Xue, Jiyan; Kurtz, Marc M; Liu, Rong; Legrand, Cheryl; He, Xuanmin; Yu, Xiang; Wong, Peggy; Hinchcliffe, John S; Tanen, Michael R; Lazar, Gloria; Zieba, Renata; Ichetovkin, Marina; Chen, Zhu; O'Neill, Edward A; Tanaka, Wesley K; Marton, Matthew J; Liao, Jason; Morris, Mark; Hailman, Eric; Tokiwa, George Y; Plump, Andrew S

    2011-11-01

    With expanding biomarker discovery efforts and increasing costs of drug development, it is critical to maximize the value of mass-limited clinical samples. The main limitation of available methods is the inability to isolate and analyze, from a single sample, molecules requiring incompatible extraction methods. Thus, we developed a novel semiautomated method for tissue processing and tissue milling and division (TMAD). We used a SilverHawk atherectomy catheter to collect atherosclerotic plaques from patients requiring peripheral atherectomy. Tissue preservation by flash freezing was compared with immersion in RNAlater®, and tissue grinding by traditional mortar and pestle was compared with TMAD. Comparators were protein, RNA, and lipid yield and quality. Reproducibility of analyte yield from aliquots of the same tissue sample processed by TMAD was also measured. The quantity and quality of biomarkers extracted from tissue prepared by TMAD was at least as good as that extracted from tissue stored and prepared by traditional means. TMAD enabled parallel analysis of gene expression (quantitative reverse-transcription PCR, microarray), protein composition (ELISA), and lipid content (biochemical assay) from as little as 20 mg of tissue. The mean correlation was r = 0.97 in molecular composition (RNA, protein, or lipid) between aliquots of individual samples generated by TMAD. We also demonstrated that it is feasible to use TMAD in a large-scale clinical study setting. The TMAD methodology described here enables semiautomated, high-throughput sampling of small amounts of heterogeneous tissue specimens by multiple analytical techniques with generally improved quality of recovered biomolecules.

  9. Correlation of NOX1 and NOX2 expression in ulcerative colitis tissue with intestinal mucosal oxidative stress response and barrier function injury

    Directory of Open Access Journals (Sweden)

    Tao Ma

    2017-10-01

    Full Text Available Objective: To study the correlation of NOX1 and NOX2 expression in ulcerative colitis tissue with intestinal mucosal oxidative stress response and barrier function injury. Methods: A total of 69 patients who were diagnosed with ulcerative colitis in Yan’an People’s Hospital between May 2015 and March 2017 were selected as the UC group of the research, and 78 patients who were diagnosed with colon polyps were selected as the polyps group of the research. The ulcerative colitis lesion and polyp lesion were collected to detect the expression of NOX1 and NOX2, the generation of oxygen free radicals as well as the contents of apoptosis molecules and mucosal barrier molecules. Results: The mRNA expression and protein expression of NOX1 and NOX2 in the intestinal mucosa of UC group were significantly higher than those of polyps group; LPO, MDA, AOPP, NO, PDCD5 and Bax levels in intestinal mucosa of UC group were significantly higher than those of polyps group and positively correlated with the mRNA expression and protein expression of NOX1 and NOX2 while Bcl-2, Cdx1, Cdx2, galectin-1, galectin-3, OCLN, cingulin and ZO-1 levels were significantly lower than those of polyps group and negatively correlated with the mRNA expression and protein expression of NOX1 and NOX2. Conclusion: The high expression of NOX1 and NOX2 in ulcerative colitis tissue can activate the intestinal mucosal oxidative stress response and result in the intestinal mucosal barrier function injury.

  10. Efficient and scalable serial extraction of DNA and RNA from frozen tissue samples.

    Science.gov (United States)

    Mathot, Lucy; Lindman, Monica; Sjöblom, Tobias

    2011-01-07

    Advances in cancer genomics have created a demand for scalable sample processing. We here present a process for serial extraction of nucleic acids from the same frozen tissue sample based on magnetic silica particles. The process is automation friendly with high recoveries of pure DNA and RNA suitable for analysis.

  11. Mitochondrial DNA from archived tissue samples kept in formalin for forensic odontology studies.

    Science.gov (United States)

    Pandey, Rahul; Mehrotra, Divya; Kowtal, Pradnya; Mahdi, Abbas Ali; Sarin, Rajiv

    2014-01-01

    Samples used for DNA isolation to be used for forensic odontology studies are often limited. The possibility to use tissue samples stored in formalin for a prolonged period, which contains nucleic acids of questionable quality, opens exciting possibilities for genetic and molecular biology studies useful in speciality of forensic odontology. The present study defines substantial modification of existing protocols for total genomic isolation including mitochondrial DNA and proves the utility of such obtained mitochondrial DNA in microsatellite analyses. 50 dental tissue samples which were kept in neutral buffered formalin liquid bottles were taken for DNA isolation and subsequent analysis. For the isolation of total genomic DNA from tissue samples, a new protocol with substantial modifications from routine ones was adopted by us. Total genomic DNA from matched blood samples were extracted using standard phenol-chloroform extraction method. Polymerase Chain Reaction and Sequencing of such extracted DNA samples for mitochondrial D loop region were successful and the results were comparable with DNA extracted from normal sources of samples. The present study reports for the first time that nucleic acids extracted from human dental tissue samples under prolonged formalin fixation times can be used for forensic odontology studies using the described methodology.

  12. Online recovery of radiocesium from soil, tissue paper and plant samples by supercritical fluid extraction

    International Nuclear Information System (INIS)

    Kanekar, A.S.; Pathak, P.N.; Mohapatra, P.K.

    2014-01-01

    The feasibility of recovery of radio-cesium from soil, tissue papers, and plant samples has been evaluated by supercritical fluid extraction (SFE) route employing calix(4)arene-mono(crown-6) (CC) dissolved in acetonitrile. These studies showed that quantitative recovery of 137 Cs from soil samples was difficult under the conditions of these studies. However, experiments performed on tissue papers (cellulose matrix) showed quantitative recovery of 137 Cs. On the other hand, 137 Cs recovery from plant samples varied between ∼50 % (for stems) and ∼67.2 % (for leaves) employing 1x10 -3 M CC + 4 M HNO 3 dissolved in acetonitrile. (author)

  13. Impacts of maternal selenium supply and nutritional plane on visceral tissues and intestinal biology in 180-day-old offspring in sheep.

    Science.gov (United States)

    Yunusova, R D; Neville, T L; Vonnahme, K A; Hammer, C J; Reed, J J; Taylor, J B; Redmer, D A; Reynolds, L P; Caton, J S

    2013-05-01

    Objectives were to investigate the effects of maternal Se supply and nutritional plane during gestation on offspring visceral tissues and indices of intestinal growth, vascularity, and function at 180 d of age. Rambouillet ewe lambs (n = 82, approximately 240 d of age; 52 ± 0.8 kg BW at breeding) were allocated to a 2 × 3 factorial arrangement of treatments. Treatments included dietary Se [adequate Se (ASe, 9.5 µg/kg BW) or high Se (HSe, 81.8 µg/kg BW)] initiated at breeding and nutritional plane [60% (restricted, RES), 100% (control, CON), and 140% (high, HI) of requirements] initiated at d 50 of gestation. Ewes were fed pelleted diets and housed individually indoors. At parturition, lambs were immediately removed and fed artificial colostrum for the first 20 h followed by ad libitum access to milk replacer. At 180 ± 2 d of age, lambs were euthanized and tissues were harvested. Birth weight was affected by nutritional treatments (P lambs from CON being greater than RES. Expression of most mRNA for measured angiogenic factors and receptors was not altered (P ≤ 0.13) by maternal treatments; however, expression of glucagon-like peptide-2 (GLP-2) was decreased (P = 0.07) in offspring from RES compared with CON ewes. Offspring from ewes fed HI diets had increased (P = 0.08) jejunal mucosal maltase activity. In conclusion, maternal Se supply and nutritional plane during gestation resulted in measurable changes in offspring visceral tissues and intestinal biology, including perirenal fat, blood volume, intestinal mass, total jejunal crypt cell proliferation, area per capillary in jejunal villi, GLP-2 mRNA expression, and maltase activity at 180 d. Additional work is needed to determine impacts on intestinal function and nutrient uptake.

  14. Sample preparation for mass spectrometry imaging of plant tissues: a review

    Directory of Open Access Journals (Sweden)

    Yonghui eDong

    2016-02-01

    Full Text Available Mass spectrometry imaging (MSI is a mass spectrometry based molecular ion imaging technique. It provides the means for ascertaining the spatial distribution of a large variety of analytes directly on tissue sample surfaces without any labeling or staining steps. These advantages make it an attractive molecular histology tool in medical, pharmaceutical and biological research. Likewise, MSI has started gaining popularity in plant sciences; yet, information regarding sample preparation methods specific for plant tissues is still limited. Sample preparation is a crucial step that is directly associated with the quality and authenticity of the imaging results, it therefore demands in-depth studies based on the characteristics of plant samples. In this review, a sample preparation pipeline is discussed in detail and illustrated through selected practical examples. In particular, special concerns regarding sample preparation for plant imaging are critically evaluated. Finally, the applications of various MSI techniques in plants are reviewed according to different classes of plant metabolites.

  15. Sample Preparation for Mass Spectrometry Imaging of Plant Tissues: A Review.

    Science.gov (United States)

    Dong, Yonghui; Li, Bin; Malitsky, Sergey; Rogachev, Ilana; Aharoni, Asaph; Kaftan, Filip; Svatoš, Aleš; Franceschi, Pietro

    2016-01-01

    Mass spectrometry imaging (MSI) is a mass spectrometry based molecular ion imaging technique. It provides the means for ascertaining the spatial distribution of a large variety of analytes directly on tissue sample surfaces without any labeling or staining agents. These advantages make it an attractive molecular histology tool in medical, pharmaceutical, and biological research. Likewise, MSI has started gaining popularity in plant sciences; yet, information regarding sample preparation methods for plant tissues is still limited. Sample preparation is a crucial step that is directly associated with the quality and authenticity of the imaging results, it therefore demands in-depth studies based on the characteristics of plant samples. In this review, a sample preparation pipeline is discussed in detail and illustrated through selected practical examples. In particular, special concerns regarding sample preparation for plant imaging are critically evaluated. Finally, the applications of MSI techniques in plants are reviewed according to different classes of plant metabolites.

  16. Rapid column extraction method for actinides and strontium in fish and other animal tissue samples

    International Nuclear Information System (INIS)

    Maxwell III, S.L.; Faison, D.M.

    2008-01-01

    The analysis of actinides and radiostrontium in animal tissue samples is very important for environmental monitoring. There is a need to measure actinide isotopes and strontium with very low detection limits in animal tissue samples, including fish, deer, hogs, beef and shellfish. A new, rapid separation method has been developed that allows the measurement of plutonium, neptunium, uranium, americium, curium and strontium isotopes in large animal tissue samples (100-200 g) with high chemical recoveries and effective removal of matrix interferences. This method uses stacked TEVA Resin R , TRU Resin R and DGA Resin R cartridges from Eichrom Technologies (Darien, IL, USA) that allows the rapid separation of plutonium (Pu), neptunium (Np), uranium (U), americium (Am), and curium (Cm) using a single multi-stage column combined with alphaspectrometry. Strontium is collected on Sr Resin R from Eichrom Technologies (Darien, IL, USA). After acid digestion and furnace heating of the animal tissue samples, the actinides and 89/90 Sr are separated using column extraction chromatography. This method has been shown to be effective over a wide range of animal tissue matrices. Vacuum box cartridge technology with rapid flow rates is used to minimize sample preparation time. (author)

  17. Measurement of the hyperelastic properties of 44 pathological ex vivo breast tissue samples

    Science.gov (United States)

    O'Hagan, Joseph J.; Samani, Abbas

    2009-04-01

    The elastic and hyperelastic properties of biological soft tissues have been of interest to the medical community. There are several biomedical applications where parameters characterizing such properties are critical for a reliable clinical outcome. These applications include surgery planning, needle biopsy and brachtherapy where tissue biomechanical modeling is involved. Another important application is interpreting nonlinear elastography images. While there has been considerable research on the measurement of the linear elastic modulus of small tissue samples, little research has been conducted for measuring parameters that characterize the nonlinear elasticity of tissues included in tissue slice specimens. This work presents hyperelastic measurement results of 44 pathological ex vivo breast tissue samples. For each sample, five hyperelastic models have been used, including the Yeoh, N = 2 polynomial, N = 1 Ogden, Arruda-Boyce, and Veronda-Westmann models. Results show that the Yeoh, polynomial and Ogden models are the most accurate in terms of fitting experimental data. The results indicate that almost all of the parameters corresponding to the pathological tissues are between two times to over two orders of magnitude larger than those of normal tissues, with C11 showing the most significant difference. Furthermore, statistical analysis indicates that C02 of the Yeoh model, and C11 and C20 of the polynomial model have very good potential for cancer classification as they show statistically significant differences for various cancer types, especially for invasive lobular carcinoma. In addition to the potential for use in cancer classification, the presented data are very important for applications such as surgery planning and virtual reality based clinician training systems where accurate nonlinear tissue response modeling is required.

  18. Measurement of the hyperelastic properties of 44 pathological ex vivo breast tissue samples

    Energy Technology Data Exchange (ETDEWEB)

    O' Hagan, Joseph J; Samani, Abbas [Department of Electrical and Computer Engineering, University of Western Ontario, London, ON (Canada)], E-mail: asamani@uwo.ca

    2009-04-21

    The elastic and hyperelastic properties of biological soft tissues have been of interest to the medical community. There are several biomedical applications where parameters characterizing such properties are critical for a reliable clinical outcome. These applications include surgery planning, needle biopsy and brachtherapy where tissue biomechanical modeling is involved. Another important application is interpreting nonlinear elastography images. While there has been considerable research on the measurement of the linear elastic modulus of small tissue samples, little research has been conducted for measuring parameters that characterize the nonlinear elasticity of tissues included in tissue slice specimens. This work presents hyperelastic measurement results of 44 pathological ex vivo breast tissue samples. For each sample, five hyperelastic models have been used, including the Yeoh, N = 2 polynomial, N = 1 Ogden, Arruda-Boyce, and Veronda-Westmann models. Results show that the Yeoh, polynomial and Ogden models are the most accurate in terms of fitting experimental data. The results indicate that almost all of the parameters corresponding to the pathological tissues are between two times to over two orders of magnitude larger than those of normal tissues, with C{sub 11} showing the most significant difference. Furthermore, statistical analysis indicates that C{sub 02} of the Yeoh model, and C{sub 11} and C{sub 20} of the polynomial model have very good potential for cancer classification as they show statistically significant differences for various cancer types, especially for invasive lobular carcinoma. In addition to the potential for use in cancer classification, the presented data are very important for applications such as surgery planning and virtual reality based clinician training systems where accurate nonlinear tissue response modeling is required.

  19. X-ray scattering for the characterization of lyophilized breast tissue samples

    Science.gov (United States)

    Elshemey, Wael M.; Mohamed, Fayrouz S.; Khater, Ibrahim M.

    2013-09-01

    This work investigates the possibility of characterizing breast cancer by measuring the X-ray scattering profiles of lyophilized excised breast tissue samples. Since X-ray scattering from water-rich tissue is dominated by scattering from water, the removal of water by lyophilization would enhance the characterization process. In the present study, X-ray scattering profiles of 22 normal, 22 malignant and 10 benign breast tissue samples are measured. The cut-offs of scatter diagrams, sensitivity, specificity and diagnostic accuracy of three characterization parameters (full width at half maximum (FWHM) for the peak at 1.1 nm-1, area under curve (AUC), and ratio of 1st to 2nd scattering peak intensities (I1/I2%)) are calculated and compared to the data from non-lyophilized samples. Results show increased sensitivity (up to 100%) of the present data on lyophilized breast tissue samples compared to previously reported data for non-lyophilized samples while the specificity (up to 95.4%), diagnostic accuracy (up to 95.4%) and receiver operating characteristic (ROC) curve values (up to 0.9979) for both sets of data are comparable. The present study shows significant differences between normal samples and each of malignant and benign samples. Only subtle differences exist between malignant and benign lyophilized breast tissue samples where FWHM=0.7±0.1 and 0.8±0.3, AUC=1.3±0.2 and 1.4±0.2 and I1/I2%=44.9±11.0 and 52.4±7.6 for malignant and benign samples respectively.

  20. Validation of proton stopping power ratio estimation based on dual energy CT using fresh tissue samples

    Science.gov (United States)

    Taasti, Vicki T.; Michalak, Gregory J.; Hansen, David C.; Deisher, Amanda J.; Kruse, Jon J.; Krauss, Bernhard; Muren, Ludvig P.; Petersen, Jørgen B. B.; McCollough, Cynthia H.

    2018-01-01

    Dual energy CT (DECT) has been shown, in theoretical and phantom studies, to improve the stopping power ratio (SPR) determination used for proton treatment planning compared to the use of single energy CT (SECT). However, it has not been shown that this also extends to organic tissues. The purpose of this study was therefore to investigate the accuracy of SPR estimation for fresh pork and beef tissue samples used as surrogates of human tissues. The reference SPRs for fourteen tissue samples, which included fat, muscle and femur bone, were measured using proton pencil beams. The tissue samples were subsequently CT scanned using four different scanners with different dual energy acquisition modes, giving in total six DECT-based SPR estimations for each sample. The SPR was estimated using a proprietary algorithm (syngo.via DE Rho/Z Maps, Siemens Healthcare, Forchheim, Germany) for extracting the electron density and the effective atomic number. SECT images were also acquired and SECT-based SPR estimations were performed using a clinical Hounsfield look-up table. The mean and standard deviation of the SPR over large volume-of-interests were calculated. For the six different DECT acquisition methods, the root-mean-square errors (RMSEs) for the SPR estimates over all tissue samples were between 0.9% and 1.5%. For the SECT-based SPR estimation the RMSE was 2.8%. For one DECT acquisition method, a positive bias was seen in the SPR estimates, having a mean error of 1.3%. The largest errors were found in the very dense cortical bone from a beef femur. This study confirms the advantages of DECT-based SPR estimation although good results were also obtained using SECT for most tissues.

  1. Collecting and Storing Tissue, Blood, and Bone Marrow Samples From Patients With Rhabdomyosarcoma or Other Soft Tissue Sarcoma

    Science.gov (United States)

    2017-12-11

    Adult Rhabdomyosarcoma; Childhood Desmoplastic Small Round Cell Tumor; Chordoma; Desmoid Tumor; Metastatic Childhood Soft Tissue Sarcoma; Nonmetastatic Childhood Soft Tissue Sarcoma; Previously Treated Childhood Rhabdomyosarcoma; Previously Untreated Childhood Rhabdomyosarcoma; Recurrent Adult Soft Tissue Sarcoma; Recurrent Childhood Rhabdomyosarcoma; Recurrent Childhood Soft Tissue Sarcoma; Stage I Adult Soft Tissue Sarcoma; Stage II Adult Soft Tissue Sarcoma; Stage III Adult Soft Tissue Sarcoma; Stage IV Adult Soft Tissue Sarcoma

  2. Characteristics of electrically injured skin from human hand tissue samples using Fourier transform infrared microspectroscopy.

    Science.gov (United States)

    Li, Shi-Ying; Zou, Dong-Hua; Luo, Yi-Wen; Sun, Qi-Ran; Deng, Kai-Fei; Chen, Yi-Jiu; Huang, Ping

    2014-01-01

    This technical note describes a method for distinguishing normal skin tissue samples from those electrically injured by Fourier transform infrared microspectroscopy (FTIR MSP). Furthermore, the infrared spectral features of electrically injured cells and tissues were evaluated to identify molecular changes in epidermal cells. In the present study, 20 human hand tissue samples were evaluated macroscopically and histopathologically. The electrically injured skin samples were subdivided into 2 regions [normal cell regions (NCRs) and polarized cell regions (PCRs)] and 14 major spectral absorption bands were selected. The spectral results showed that the band absorbance at 1080, 1126, 1172, 1242, 1307, 1403, 1456, 1541, 2852, 2925, 2957, 3075, and 3300cm(-1) increased significantly both in the stratum and non-stratum corneum of the PCRs in electrically injured skin tissues samples. No significant difference was found between normal skin and the NCR of the electrically injured skin samples. The band absorbance ratios of A1172/A1126, A1456/A1403, and A2925/A2957 were significantly increased, whereas the A1652/A1541 ratio was decreased in the PCR of the stratum corneum and non-stratum corneum. Baseline changes from 4000 to near 1737cm(-1) were observed in the spectra of the electrically injured skin samples, which were interpreted in terms of the pathological process involved in electrical injury. FTIR-MSP presents a useful method to provide objective spectral markers for the assisted diagnosis of electrical marks. © 2013.

  3. Sample Preparation of Corn Seed Tissue to Prevent Analyte Relocations for Mass Spectrometry Imaging

    Science.gov (United States)

    Kim, Shin Hye; Kim, Jeongkwon; Lee, Young Jin; Lee, Tae Geol; Yoon, Sohee

    2017-08-01

    Corn seed tissue sections were prepared by the tape support method using an adhesive tape, and mass spectrometry imaging (MSI) was performed. The effect of heat generated during sample preparation was investigated by time-of-flight secondary mass spectrometry (TOF-SIMS) imaging of corn seed tissue prepared by the tape support and the thaw-mounted methods. Unlike thaw-mounted sample preparation, the tape support method does not cause imaging distortion because of the absence of heat, which can cause migration of the analytes on the sample. By applying the tape-support method, the corn seed tissue was prepared without structural damage and MSI with accurate spatial information of analytes was successfully performed.

  4. Sample Preparation of Corn Seed Tissue to Prevent Analyte Relocations for Mass Spectrometry Imaging.

    Science.gov (United States)

    Kim, Shin Hye; Kim, Jeongkwon; Lee, Young Jin; Lee, Tae Geol; Yoon, Sohee

    2017-08-01

    Corn seed tissue sections were prepared by the tape support method using an adhesive tape, and mass spectrometry imaging (MSI) was performed. The effect of heat generated during sample preparation was investigated by time-of-flight secondary mass spectrometry (TOF-SIMS) imaging of corn seed tissue prepared by the tape support and the thaw-mounted methods. Unlike thaw-mounted sample preparation, the tape support method does not cause imaging distortion because of the absence of heat, which can cause migration of the analytes on the sample. By applying the tape-support method, the corn seed tissue was prepared without structural damage and MSI with accurate spatial information of analytes was successfully performed. Graphical Abstract ᅟ.

  5. Pathologic characteristics of gut-associated lymphoid tissues and lymphocyte apoptosis in mouse intestine after neutron-and γ-irradiation

    International Nuclear Information System (INIS)

    Fu Kaifei; Peng Ruiyun; Gao Yabing; Wang Dewen; Chen Haoyu; Wu Xiaohong; Yang Yi; Hu Wenhua; Ma Junjie

    2004-01-01

    Objective: To compare the pathologic characteristics of gut-associated lymphoid tissues and lymphocyte apoptosis in neutron-irradiated mouse small intestines with those in γ-irradiated ones. Methods: Altogether 350 BALB/c mice were irradiated with different doses of neutrons or γ-rays, and were sacrificed on 6 h,12 h,125 d, 7 d, 14 d, 21 d and 28 d after irradiation and their total intestines were removed. Then the pathologic changes and death mode of lymphocytes in gut-associated lymphoid tissues were studied comparatively with light microscopy, electron microscopy and in situ terminal labeling method. Results: The basic pathologic changes of gut-associated lymphoid tissues after neutron irradiation included degeneration, apoptosis and necrosis of lymphocytes. The number of lymphocytes also decreased. There was no obvious regeneration after 4.0 and 5.5 Gy neutron irradiation, while after 2.5 Gy regeneration and recovery appeared, which were, there fore, dose-dependent. In the 2.5 Gy neutron group, the numbers of lymphocytes of intramucosal and submucous lymphoid tissues decreased, and karyopyknosis and a great quantity of nuclear fragments could also be observed at 6 h-3 d after irradiation. However, on the 3rd day regeneration of crypt epithelial cells appeared. On the 5th day hyperplasia of submucous lymphocytic tissues appeared, but recovery to normal level was not achieved till 14 d after irradiation. The basic pathologic changes after γ-irradiation were similar to that of neutron irradiation. Regeneration and recovery appeared in the 5.5 Gy group while no obvious regeneration in the 12.0 Gy group. The results of in situ terminal labeling indicated that at 6 h after irradiation the number of apoptotic cells in gut-associated lymphoid tissues of each group increased obviously, while in 4.0 Gy neutron group and 12.0 Gy γ-ray group it was more abundant. Conclusion: Both 2.5-5.5 Gy neutron and 5.5-12.0 Gy γ-ray irradiation can induce obvious injuries in gut

  6. Trace element contamination in feather and tissue samples from Anna’s hummingbirds

    Science.gov (United States)

    Mikoni, Nicole A.; Poppenga, Robert H.; Ackerman, Joshua T.; Foley, Janet E.; Hazlehurst, Jenny; Purdin, Güthrum; Aston, Linda; Hargrave, Sabine; Jelks, Karen; Tell, Lisa A.

    2017-01-01

    Trace element contamination (17 elements; Be, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, As, Se, Mo, Cd, Ba, Hg, Tl, and Pb) of live (feather samples only) and deceased (feather and tissue samples) Anna's hummingbirds (Calypte anna) was evaluated. Samples were analyzed using inductively coupled plasma-mass spectrometry (ICP-MS; 17 elements) and atomic absorption spectrophotometry (Hg only). Mean plus one standard deviation (SD) was considered the benchmark, and concentrations above the mean + 1 SD were considered elevated above normal. Contour feathers were sampled from live birds of varying age, sex, and California locations. In order to reduce thermal impacts, minimal feathers were taken from live birds, therefore a novel method was developed for preparation of low mass feather samples for ICP-MS analysis. The study found that the novel feather preparation method enabled small mass feather samples to be analyzed for trace elements using ICP-MS. For feather samples from live birds, all trace elements, with the exception of beryllium, had concentrations above the mean + 1 SD. Important risk factors for elevated trace element concentrations in feathers of live birds were age for iron, zinc, and arsenic, and location for iron, manganese, zinc, and selenium. For samples from deceased birds, ICP-MS results from body and tail feathers were correlated for Fe, Zn, and Pb, and feather concentrations were correlated with renal (Fe, Zn, Pb) or hepatic (Hg) tissue concentrations. Results for AA spectrophotometry analyzed samples from deceased birds further supported the ICP-MS findings where a strong correlation between mercury concentrations in feather and tissue (pectoral muscle) samples was found. These study results support that sampling feathers from live free-ranging hummingbirds might be a useful, non-lethal sampling method for evaluating trace element exposure and provides a sampling alternative since their small body size limits traditional sampling of blood and tissues. The

  7. Impact of tissue sampling on accuracy of Ki67 immunohistochemistry evaluation in breast cancer.

    Science.gov (United States)

    Besusparis, Justinas; Plancoulaine, Benoit; Rasmusson, Allan; Augulis, Renaldas; Green, Andrew R; Ellis, Ian O; Laurinaviciene, Aida; Herlin, Paulette; Laurinavicius, Arvydas

    2016-08-30

    Gene expression studies have identified molecular subtypes of breast cancer with implications to chemotherapy recommendations. For distinction of these types, a combination of immunohistochemistry (IHC) markers, including proliferative activity of tumor cells, estimated by Ki67 labeling index is used. Clinical studies are frequently based on IHC performed on tissue microarrays (TMA) with variable tissue sampling. This raises the need for evidence-based sampling criteria for individual IHC biomarker studies. We present a novel tissue sampling simulation model and demonstrate its application on Ki67 assessment in breast cancer tissue taking intratumoral heterogeneity into account. Whole slide images (WSI) of 297 breast cancer sections, immunohistochemically stained for Ki67, were subjected to digital image analysis (DIA). Percentage of tumor cells stained for Ki67 was computed for hexagonal tiles super-imposed on the WSI. From this, intratumoral Ki67 heterogeneity indicators (Haralick's entropy values) were extracted and used to dichotomize the tumors into homogeneous and heterogeneous subsets. Simulations with random selection of hexagons, equivalent to 0.75 mm circular diameter TMA cores, were performed. The tissue sampling requirements were investigated in relation to tumor heterogeneity using linear regression and extended error analysis. The sampling requirements were dependent on the heterogeneity of the biomarker expression. To achieve a coefficient error of 10 %, 5-6 cores were needed for homogeneous cases, 11-12 cores for heterogeneous cases; in mixed tumor population 8 TMA cores were required. Similarly, to achieve the same accuracy, approximately 4,000 nuclei must be counted when the intratumor heterogeneity is mixed/unknown. Tumors of low proliferative activity would require larger sampling (10-12 TMA cores, or 6,250 nuclei) to achieve the same error measurement results as for highly proliferative tumors. Our data show that optimal tissue sampling for

  8. X-ray scattering for the characterization of lyophilized breast tissue samples

    International Nuclear Information System (INIS)

    Elshemey, Wael M.; Mohamed, Fayrouz S.; Khater, Ibrahim M.

    2013-01-01

    This work investigates the possibility of characterizing breast cancer by measuring the X-ray scattering profiles of lyophilized excised breast tissue samples. Since X-ray scattering from water-rich tissue is dominated by scattering from water, the removal of water by lyophilization would enhance the characterization process. In the present study, X-ray scattering profiles of 22 normal, 22 malignant and 10 benign breast tissue samples are measured. The cut-offs of scatter diagrams, sensitivity, specificity and diagnostic accuracy of three characterization parameters (full width at half maximum (FWHM) for the peak at 1.1 nm −1 , area under curve (AUC), and ratio of 1st to 2nd scattering peak intensities (I 1 /I 2 %)) are calculated and compared to the data from non-lyophilized samples. Results show increased sensitivity (up to 100%) of the present data on lyophilized breast tissue samples compared to previously reported data for non-lyophilized samples while the specificity (up to 95.4%), diagnostic accuracy (up to 95.4%) and receiver operating characteristic (ROC) curve values (up to 0.9979) for both sets of data are comparable. The present study shows significant differences between normal samples and each of malignant and benign samples. Only subtle differences exist between malignant and benign lyophilized breast tissue samples where FWHM=0.7±0.1 and 0.8±0.3, AUC=1.3±0.2 and 1.4±0.2 and I 1 /I 2 %=44.9±11.0 and 52.4±7.6 for malignant and benign samples respectively. - Highlights: • X-ray scattering profiles of breast tissue samples are acquired. • Three X-ray profile characterization parameters are calculated. • The cut-offs, sensitivity, specificity and diagnostic accuracy are calculated. • They are compared to the data from non-lyophilized samples. • Results show increased sensitivity in case of lyophilized samples

  9. Automated MALDI Matrix Coating System for Multiple Tissue Samples for Imaging Mass Spectrometry

    Science.gov (United States)

    Mounfield, William P.; Garrett, Timothy J.

    2012-03-01

    Uniform matrix deposition on tissue samples for matrix-assisted laser desorption/ionization (MALDI) is key for reproducible analyte ion signals. Current methods often result in nonhomogenous matrix deposition, and take time and effort to produce acceptable ion signals. Here we describe a fully-automated method for matrix deposition using an enclosed spray chamber and spray nozzle for matrix solution delivery. A commercial air-atomizing spray nozzle was modified and combined with solenoid controlled valves and a Programmable Logic Controller (PLC) to control and deliver the matrix solution. A spray chamber was employed to contain the nozzle, sample, and atomized matrix solution stream, and to prevent any interference from outside conditions as well as allow complete control of the sample environment. A gravity cup was filled with MALDI matrix solutions, including DHB in chloroform/methanol (50:50) at concentrations up to 60 mg/mL. Various samples (including rat brain tissue sections) were prepared using two deposition methods (spray chamber, inkjet). A linear ion trap equipped with an intermediate-pressure MALDI source was used for analyses. Optical microscopic examination showed a uniform coating of matrix crystals across the sample. Overall, the mass spectral images gathered from tissues coated using the spray chamber system were of better quality and more reproducible than from tissue specimens prepared by the inkjet deposition method.

  10. Intestinal ischemia-reperfusion injury augments intestinal mucosal injury and bacterial translocation in jaundiced rats.

    Science.gov (United States)

    Yüksek, Yunus Nadi; Kologlu, Murat; Daglar, Gül; Doganay, Mutlu; Dolapci, Istar; Bilgihan, Ayse; Dolapçi, Mete; Kama, Nuri Aydin

    2004-01-01

    The aim of this study was to evaluate local effects and degree of bacterial translocation related with intestinal ischemia-reperfusion injury in a rat obstructive jaundice model. Thirty adult Sprague-Dawley rats (200-250 g) were divided into three groups; including Group 1 (jaundice group), Group 2 (jaundice-ischemia group) and Group 3 (ischemia group). All rats had 2 laparotomies. After experimental interventions, tissue samples for translocation; liver and ileum samples for histopathological examination, 25 cm of small intestine for mucosal myeloperoxidase and malondialdehyde levels and blood samples for biochemical analysis were obtained. Jaundiced rats had increased liver enzyme levels and total and direct bilirubin levels (p<0.05). Intestinal mucosal myeloperoxidase and malondialdehyde levels were found to be high in intestinal ischemia-reperfusion groups (p<0.05). Intestinal mucosal damage was more severe in rats with intestinal ischemia-reperfusion after bile duct ligation (p<0.05). Degree of bacterial translocation was also found to be significantly high in these rats (p<0.05). Intestinal mucosa is disturbed more severely in obstructive jaundice with the development of ischemia and reperfusion. Development of intestinal ischemia-reperfusion in obstructive jaundice increases bacterial translocation.

  11. Phase-Contrast Hounsfield Units of Fixated and Non-Fixated Soft-Tissue Samples.

    Directory of Open Access Journals (Sweden)

    Marian Willner

    Full Text Available X-ray phase-contrast imaging is a novel technology that achieves high soft-tissue contrast. Although its clinical impact is still under investigation, the technique may potentially improve clinical diagnostics. In conventional attenuation-based X-ray computed tomography, radiological diagnostics are quantified by Hounsfield units. Corresponding Hounsfield units for phase-contrast imaging have been recently introduced, enabling a setup-independent comparison and standardized interpretation of imaging results. Thus far, the experimental values of few tissue types have been reported; these values have been determined from fixated tissue samples. This study presents phase-contrast Hounsfield units for various types of non-fixated human soft tissues. A large variety of tissue specimens ranging from adipose, muscle and connective tissues to liver, kidney and pancreas tissues were imaged by a grating interferometer with a rotating-anode X-ray tube and a photon-counting detector. Furthermore, we investigated the effects of formalin fixation on the quantitative phase-contrast imaging results.

  12. Three dimensional imaging of paraffin embedded human lung tissue samples by micro-computed tomography.

    Directory of Open Access Journals (Sweden)

    Anna E Scott

    Full Text Available Understanding the three-dimensional (3-D micro-architecture of lung tissue can provide insights into the pathology of lung disease. Micro computed tomography (µCT has previously been used to elucidate lung 3D histology and morphometry in fixed samples that have been stained with contrast agents or air inflated and dried. However, non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data.FFPE human lung tissue samples (n = 4 were scanned using a Nikon metrology µCT scanner. Semi-automatic techniques were used to segment the 3D structure of airways and blood vessels. Airspace size (mean linear intercept, Lm was measured on µCT images and on matched histological sections from the same FFPE samples imaged by light microscopy to validate µCT imaging.The µCT imaging protocol provided contrast between tissue and paraffin in FFPE samples (15 mm x 7 mm. Resolution (voxel size 6.7 µm in the reconstructed images was sufficient for semi-automatic image segmentation of airways and blood vessels as well as quantitative airspace analysis. The scans were also used to scout for regions of interest, enabling time-efficient preparation of conventional histological sections. The Lm measurements from µCT images were not significantly different to those from matched histological sections.We demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory µCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis.

  13. Improved application of the electrophoretic tissue clearing technology, CLARITY, to intact solid organs including brain, pancreas, liver, kidney, lung, and intestine.

    Science.gov (United States)

    Lee, Hyunsu; Park, Jae-Hyung; Seo, Incheol; Park, Sun-Hyun; Kim, Shin

    2014-12-21

    Mapping of tissue structure at the cellular, circuit, and organ-wide scale is important for understanding physiological and biological functions. A bio-electrochemical technique known as CLARITY used for three-dimensional anatomical and phenotypical mapping within transparent intact tissues has been recently developed. This method provided a major advance in understanding the structure-function relationships in circuits of the nervous system and organs by using whole-body clearing. Thus, in the present study, we aimed to improve the original CLARITY procedure and developed specific CLARITY protocols for various intact organs. We determined the optimal conditions for reducing bubble formation, discoloration, and depositing of black particles on the surface of tissue, which allowed production of clearer organ images. We also determined the appropriate replacement cycles of clearing solution for each type of organ, and convincingly demonstrated that 250-280 mA is the ideal range of electrical current for tissue clearing. We then acquired each type of cleared organs including brain, pancreas, liver, lung, kidney, and intestine. Additionally, we determined the images of axon fibers of hippocampal region, the Purkinje layer of cerebellum, and vessels and cellular nuclei of pancreas. CLARITY is an innovative biochemical technology for the structural and molecular analysis of various types of tissue. We developed improved CLARITY methods for clearing of the brain, pancreas, lung, intestine, liver, and kidney, and identified the appropriate experimental conditions for clearing of each specific tissue type. These optimized methods will be useful for the application of CLARITY to various types of organs.

  14. Sample dilution : A methodological pitfall in the measurement of tissue but not serum ACE-activity

    NARCIS (Netherlands)

    Koiter, J; Navis, G; de Jong, PE; van Gilst, WH; de Zeeuw, D

    Many tissue ACE-assays suffer from underestimation of the ACE-activity at low sample dilutions. However, measurement of ACE-activity as the amount of hippuric acid produced by cleavage of the commonly used substrate hippuryl-histidyl-leucine might circumvent this problem. In this study, we

  15. DNA damage in preserved specimens and tissue samples: a molecular assessment

    Directory of Open Access Journals (Sweden)

    Cantin Elizabeth

    2008-10-01

    Full Text Available Abstract The extraction of genetic information from preserved tissue samples or museum specimens is a fundamental component of many fields of research, including the Barcode of Life initiative, forensic investigations, biological studies using scat sample analysis, and cancer research utilizing formaldehyde-fixed, paraffin-embedded tissue. Efforts to obtain genetic information from these sources are often hampered by an inability to amplify the desired DNA as a consequence of DNA damage. Previous studies have described techniques for improved DNA extraction from such samples or focused on the effect of damaging agents – such as light, oxygen or formaldehyde – on free nucleotides. We present ongoing work to characterize lesions in DNA samples extracted from preserved specimens. The extracted DNA is digested to single nucleosides with a combination of DNase I, Snake Venom Phosphodiesterase, and Antarctic Phosphatase and then analyzed by HPLC-ESI-TOF-MS. We present data for moth specimens that were preserved dried and pinned with no additional preservative and for frog tissue samples that were preserved in either ethanol, or formaldehyde, or fixed in formaldehyde and then preserved in ethanol. These preservation methods represent the most common methods of preserving animal specimens in museum collections. We observe changes in the nucleoside content of these samples over time, especially a loss of deoxyguanosine. We characterize the fragmentation state of the DNA and aim to identify abundant nucleoside lesions. Finally, simple models are introduced to describe the DNA fragmentation based on nicks and double-strand breaks.

  16. Augmenter of liver regeneration gene expression in human colon cancer cell lines and clinical tissue samples.

    Science.gov (United States)

    Gatzidou, Elisavet; Mantzourani, Marina; Giaginis, Constantinos; Giagini, Athina; Patsouris, Efstratios; Kouraklis, Gregory; Theocharis, Stamatios

    2015-01-01

    Augmenter of liver regeneration (ALR) is an hepatotrophic factor responsible for the increased regenerative capacity of mammalian liver and ALR gene expression has been well-documented in liver cirrhosis and hepatocellular carcinoma tissue samples. The present study aimed to quantify and evaluate ALR gene expression in human colon cancer cell lines and tissue samples. Total RNA was isolated from 6 colorectal cancer cell lines and 23 primary colorectal tumors, cDNA was prepared and ALR mRNA expression analysis was performed using quantitative real-time PCR. ALR mRNA expression was confirmed in all 6 colorectal cancer cell lines (SW480, SW620, DLD-1, RKO, COLO-205 and HTC-116) and an epithelial one (WISH). DLD-1 cell line showed the highest ALR mRNA levels, followed by RKO, COLO-205, HCT-116, SW480, SW620 and WISH cell lines. ALR gene expression levels were detected in all cancer tissue samples (N=23), being significantly increased in well/moderately compared to poorly differentiated tumors (p=0.0208). ALR gene expression levels were increased in Dukes' stage A/B compared to stage C tumors, at a non significant level (p=0.2842). ALR mRNA levels were slightly higher in colon cancer tissues compared to adjacent non-neoplastic ones (N=19), at a non significant level (p=0.2122). The present study verified for the first time the ALR gene expression in both human colon cancer cell lines and clinical samples. Enhanced ALR gene expression was negatively correlated with advanced histopathological grade and stage in both colon cancer cell lines and human tissue samples, implicating ALR participation at the early stage of colon malignant progression.

  17. Preservation and rapid purification of DNA from decomposing human tissue samples.

    Science.gov (United States)

    Sorensen, Amy; Rahman, Elizabeth; Canela, Cassandra; Gangitano, David; Hughes-Stamm, Sheree

    2016-11-01

    One of the key features to be considered in a mass disaster is victim identification. However, the recovery and identification of human remains are sometimes complicated by harsh environmental conditions, limited facilities, loss of electricity and lack of refrigeration. If human remains cannot be collected, stored, or identified immediately, bodies decompose and DNA degrades making genotyping more difficult and ultimately decreasing DNA profiling success. In order to prevent further DNA damage and degradation after collection, tissue preservatives may be used. The goal of this study was to evaluate three customized (modified TENT, DESS, LST) and two commercial DNA preservatives (RNAlater and DNAgard ® ) on fresh and decomposed human skin and muscle samples stored in hot (35°C) and humid (60-70% relative humidity) conditions for up to three months. Skin and muscle samples were harvested from the thigh of three human cadavers placed outdoors for up to two weeks. In addition, the possibility of purifying DNA directly from the preservative solutions ("free DNA") was investigated in order to eliminate lengthy tissue digestion processes and increase throughput. The efficiency of each preservative was evaluated based on the quantity of DNA recovered from both the "free DNA" in solution and the tissue sample itself in conjunction with the quality and completeness of downstream STR profiles. As expected, DNA quantity and STR success decreased with time of decomposition. However, a marked decrease in DNA quantity and STR quality was observed in all samples after the bodies entered the bloat stage (approximately six days of decomposition in this study). Similar amounts of DNA were retrieved from skin and muscle samples over time, but slightly more complete STR profiles were obtained from muscle tissue. Although higher amounts of DNA were recovered from tissue samples than from the surrounding preservative, the average number of reportable alleles from the "free DNA" was

  18. A method for the determination of potassium concentration in organic tissue samples

    International Nuclear Information System (INIS)

    Maciel, A.C.A.

    1976-12-01

    An original method has been developed to detect small variations of potassium in several samples of organic tissue. These variations are relative to elements that are biologically representative, such as carbon, oxygen, and nitrogen. The samples are irradiated with a beam of protons from a Van de Graaff accelerator (4MV). Vacancies are created in the K-shell of potassium, and x-rays are emitted when these vacancies are filled with outer electrons. These X-rays and the protons elastically scattered by the nuclei of carbon, nitrogen and oxygen are detected and their energy spectra are analysed by computer programs especially elaborated for this purpose. A technique for routine preparation of samples in the laboratory was developed including the production of aluminum support layers, and the preparation of organic tissue samples with a low temperature microtome. The unique features of this method are that it does not destroy the tissue, permitting further analysis with the microscope, and the normalization of the amount of potassium using other elements (C,O,N) instead of the total mass of the sample. (Author) [pt

  19. Three-Dimensional Coculture of Meniscal Cells and Mesenchymal Stem Cells in Collagen Type I Hydrogel on a Small Intestinal Matrix-A Pilot Study Toward Equine Meniscus Tissue Engineering.

    Science.gov (United States)

    Kremer, Antje; Ribitsch, Iris; Reboredo, Jenny; Dürr, Julia; Egerbacher, Monika; Jenner, Florien; Walles, Heike

    2017-05-01

    Meniscal injuries are the most frequently encountered soft tissue injuries in the equine stifle joint. Due to the inherent limited repair potential of meniscal tissue, meniscal injuries do not only affect the meniscus itself but also lead to impaired joint homeostasis and secondary osteoarthritis. The presented study compares 3D coculture constructs of primary equine mesenchymal stem cells (MSC) and meniscus cells (MC) seeded on three different scaffolds-a cell-laden collagen type I hydrogel (Col I gel), a tissue-derived small intestinal matrix scaffold (SIS-muc) and a combination thereof-for their qualification to be applied for meniscus tissue engineering. To investigate cell attachment of primary MC and MSC on SIS-muc matrix SEM pictures were performed. For molecular analysis, lyophilized samples of coculture constructs with different cell ratios (100% MC, 100% MSC, and 50% MC and 50% MSC, 20% MC, and 80% MSC) were digested and analyzed for DNA and GAG content. Active matrix remodeling of 3D coculture models was indicated by matrix metalloproteinases detection. For comparison of tissue-engineered constructs with the histologic architecture of natural equine menisci, paired lateral and medial menisci of 15 horses representing different age groups were examined. A meniscus phenotype with promising similarity to native meniscus tissue in its GAG/DNA expression in addition to Col I, Col II, and Aggrecan production was achieved using a scaffold composed of Col I gel on SIS-muc combined with a coculture of MC and MSC. The results encourage further development of this scaffold-cell combination for meniscus tissue engineering.

  20. Comparison of different biopsy forceps models for tissue sampling in eosinophilic esophagitis.

    Science.gov (United States)

    Bussmann, Christian; Schoepfer, Alain M; Safroneeva, Ekaterina; Haas, Nadine; Godat, Sébastien; Sempoux, Christine; Simon, Hans-Uwe; Straumann, Alex

    2016-12-01

    Background and aims: Eosinophilic esophagitis (EoE) is a mixed inflammatory and fibrostenotic disease. Unlike superficial inflammatory changes, subepithelial fibrosis is not routinely sampled in esophageal biopsies. This study aimed to evaluate the efficacy and safety of deep esophageal sampling with four different types of biopsy forceps. Patients and methods: In this cross-sectional study, esophageal biopsies were taken in 30 adult patients by one expert endoscopist. Biopsies sampled from distal esophagus using a static jaw forceps (Olympus, FB-11K-1) were compared with proximal biopsies sampled with static jaw (Olympus, FB-45Q-1), alligator jaw (Olympus, FB-210K), and large-capacity forceps (Boston Scientific, Radial Jaw 4). One pathologist calculated the surface area of epithelial and subepithelial layers in hematoxylin and eosin (H&E)-stained biopsies. Results: Subepithelial tissue was acquired in 97 % (static jaw FB-11K-1), 93 % (static jaw FB-45Q-1), 80 % (alligator jaw), and 55 % (large-capacity) of samples. Median (interquartile [IQR]) ratios of surface area of epithelial to subepithelial tissue were: static jaw FB-45Q-1, 1.07 (0.65 - 4.465); static jaw FB-11K-1, 1.184 (0.608 - 2.545); alligator jaw, 2.353 (1.312 - 4.465); and large-capacity, 2.71 (1.611 - 4.858). The static jaw models obtained a larger surface area of subepithelial tissue compared with the alligator jaw ( P   90 % of biopsies and appear to be superior to alligator or large-capacity forceps in sampling larger amounts of subepithelial tissue. © Georg Thieme Verlag KG Stuttgart · New York.

  1. Do anesthetics and sampling strategies affect transcription analysis of fish tissues?

    Directory of Open Access Journals (Sweden)

    Hevrøy Ernst M

    2007-06-01

    Full Text Available Abstract Background The aim of the current examination was to evaluate if sedation and anesthetic treatment techniques affect the quality of RNA extracted from liver, gill, head kidney and brain tissues in Atlantic salmon Salmo salar L. Blood parameters were measured and tissue specimens sampled in six groups of fish; one control group (0 minutes, two groups kept in pure seawater in 90 liter tanks for 30 and 120 minutes, two groups treated with the anesthetic isoeugenol for 30 and 120 minutes, and one group kept in pure seawater for 105 minutes and then anaesthetized with metacaine for 15 minutes. RNA quality was assessed with the NanoDrop ND-1000 spectrophotometer (260/280 and 260/230 nm ratios and with the Agilent Bioanalyzer (28S/18S ratio and RIN data in samples either preserved in liquefied nitrogen (N2 or in RNAlater. In addition, the transcriptional levels of two fast-responding genes were quantified in gill and brain tissues. Results The results show that physiological stress during sampling does not affect the quality of RNA extracted from fish specimens. However, prolonged sedation (2 hours resulted in a metabolic alkalosis that again affected the transcriptional levels of genes involved in ionoregulation and respiration. In gills, Na+-K+-ATPase α1b was significantly downregulated and hypoxia inducible factor 1 (HIF1 significantly upregulated after two hours of treatment with isoeugenol, suggesting that this commonly used sedative affects osmo-regulation and respiration in the fish. The results also suggest that for tissue preservation in general it is better to flash-freeze fish specimens in liquefied N2 than to use RNAlater. Conclusion Prolonged sedation may affect the transcription of fast-responding genes in tissues of fish. Two hours of sedation with isoeugenol resulted in downregulation of the Na+-K+-ATPase α1b gene and upregulation of the HIF1 gene in gills of Atlantic salmon. The quality of RNA extracted from tissue specimens

  2. Spatially-Resolved Proteomics: Rapid Quantitative Analysis of Laser Capture Microdissected Alveolar Tissue Samples

    Energy Technology Data Exchange (ETDEWEB)

    Clair, Geremy; Piehowski, Paul D.; Nicola, Teodora; Kitzmiller, Joseph A.; Huang, Eric L.; Zink, Erika M.; Sontag, Ryan L.; Orton, Daniel J.; Moore, Ronald J.; Carson, James P.; Smith, Richard D.; Whitsett, Jeffrey A.; Corley, Richard A.; Ambalavanan, Namasivayam; Ansong, Charles

    2016-12-22

    Global proteomics approaches allow characterization of whole tissue lysates to an impressive depth. However, it is now increasingly recognized that to better understand the complexity of multicellular organisms, global protein profiling of specific spatially defined regions/substructures of tissues (i.e. spatially-resolved proteomics) is essential. Laser capture microdissection (LCM) enables microscopic isolation of defined regions of tissues preserving crucial spatial information. However, current proteomics workflows entail several manual sample preparation steps and are challenged by the microscopic mass-limited samples generated by LCM, and that impact measurement robustness, quantification, and throughput. Here, we coupled LCM with a fully automated sample preparation workflow that with a single manual step allows: protein extraction, tryptic digestion, peptide cleanup and LC-MS/MS analysis of proteomes from microdissected tissues. Benchmarking against the current state of the art in ultrasensitive global proteomic analysis, our approach demonstrated significant improvements in quantification and throughput. Using our LCM-SNaPP proteomics approach, we characterized to a depth of more than 3,400 proteins, the ontogeny of protein changes during normal lung development in laser capture microdissected alveolar tissue containing ~4,000 cells per sample. Importantly, the data revealed quantitative changes for 350 low abundance transcription factors and signaling molecules, confirming earlier transcript-level observations and defining seven modules of coordinated transcription factor/signaling molecule expression patterns, suggesting that a complex network of temporal regulatory control directs normal lung development with epigenetic regulation fine-tuning pre-natal developmental processes. Our LCM-proteomics approach facilitates efficient, spatially-resolved, ultrasensitive global proteomics analyses in high-throughput that will be enabling for several clinical and

  3. [Optical properties of human normal small intestine tissue with theoretical model of optics about biological tissues at Ar+ laser and 532 nm laser and their linearly polarized laser irradiation in vitro].

    Science.gov (United States)

    Wei, Hua-jiang; Xing, Da; Wu, Guo-yong; Jin, Ying; Gu, Huai-min

    2004-05-01

    A double-integrating-spheres system, basic principle of measuring technology of ray radiation, and optical model of biological tissues were used for the study. Optical properties of human normal small intestine tissue at 476.5, 488, 496.5, 514.5 and 532 nm laser and their linearly polarized laser irradiation were studied. The results of measurement showed that the total attenuation coefficient and scattering coefficient of the tissue at these wavelengths of laser and their linearly polarized laser irradiation increased with decreasing wavelengths. And obviously there was a distinction at 514.5 to 532 nm wavelength between lasers and their linearly polarized laser irradiation. Absorption coefficient of tissue at these wavelengths of laser and their linearly polarized laser irradiation increased with decreasing wavelengths. Absorption coefficient of tissue at 514.5 to 532 nm wavelength of laser was obviously decreasing, which was independent of these wavelengths of laser or their linearly polarized laser irradiation. Mean cosine of scattering of tissue at these wavelengths of laser and their linearly polarized laser irradiation also increased with decreasing wavelengths. But penetration depth of tissue at these wavelengths of laser and their linearly polarized laser irradiation also increased with increasing of wavelengths. Refractive index of tissue between these wavelengths of laser was within 1.38 to 1.48. Absorption coefficient, scattering coefficient, total attenuation coefficient, effective attenuation coefficients of tissue in Kubelka-Munk two-flux model at the same wavelength of laser and their linearly polarized laser irradiation showed no prominent distinction (P>0.01). Absorption coefficient, scattering coefficient, total attenuation coefficient, effective attenuation coefficients of tissue in Kubelka-Munk two-flux model at different wavelength of laser and their linearly polarized laser irradiation showed obvious distinction. Optical properties of tissue

  4. Screening of Viral Pathogens from Pediatric Ileal Tissue Samples after Vaccination

    Directory of Open Access Journals (Sweden)

    Laura Hewitson

    2014-01-01

    Full Text Available In 2010, researchers reported that the two US-licensed rotavirus vaccines contained DNA or DNA fragments from porcine circovirus (PCV. Although PCV, a common virus among pigs, is not thought to cause illness in humans, these findings raised several safety concerns. In this study, we sought to determine whether viruses, including PCV, could be detected in ileal tissue samples of children vaccinated with one of the two rotavirus vaccines. A broad spectrum, novel DNA detection technology, the Lawrence Livermore Microbial Detection Array (LLMDA, was utilized, and confirmation of viral pathogens using the polymerase chain reaction (PCR was conducted. The LLMDA technology was recently used to identify PCV from one rotavirus vaccine. Ileal tissue samples were analyzed from 21 subjects, aged 15–62 months. PCV was not detected in any ileal tissue samples by the LLMDA or PCR. LLMDA identified a human rotavirus A from one of the vaccinated subjects, which is likely due to a recent infection from a wild type rotavirus. LLMDA also identified human parechovirus, a common gastroenteritis viral infection, from two subjects. Additionally, LLMDA detected common gastrointestinal bacterial organisms from the Enterobacteriaceae, Bacteroidaceae, and Streptococcaceae families from several subjects. This study provides a survey of viral and bacterial pathogens from pediatric ileal samples, and may shed light on future studies to identify pathogen associations with pediatric vaccinations.

  5. Combined micro-XRF and TXRF methodology for quantitative elemental imaging of tissue samples.

    Science.gov (United States)

    Wróbel, Paweł M; Bała, Sławomir; Czyzycki, Mateusz; Golasik, Magdalena; Librowski, Tadeusz; Ostachowicz, Beata; Piekoszewski, Wojciech; Surówka, Artur; Lankosz, Marek

    2017-01-01

    Local differences in structural properties of biological specimens pose a major limitation to quantitative X-ray fluorescence imaging. This is because both the various tissue compartments of different density and variation in the sample thickness upon frequently used freeze-drying come up with the different values of the sample mass per unit area to be taken into account. Even though several solutions to tackle this problem based on the home-made standards for quantification in terms of thickness- and density-independent elemental mass fractions have been proposed, this issue is not addressed enough due to the samples' heterogeneity. In our recent study, we propose a calculation scheme based on combined external-standard micro X-ray fluorescence (micro-XRF) imaging and internal-standard total reflection X-ray fluorescence (TXRF) analysis to determine the corrected elemental mass fraction distributions in commonly analysed rat tissues: kidney, liver and spleen. The results of TXRF analysis of digested large tissue sections together with the mean values of elemental masses per unit area obtained with micro-XRF were employed to determine the average masses per unit area of the samples. The correction for variation of the tissue thickness and density was done through with the use of Compton intensities. Importantly, by its versatility, our novel approach can be used to produce elemental contrast in a variety of biological specimens where local variations in either the sample density or thickness are no longer the issue. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Application of neutron activation analysis for determination of trace element Pt in the rat's tissue sample

    International Nuclear Information System (INIS)

    Wardani, S.; Mulyaningsih, R.

    1996-01-01

    The neutron activation analysis (NAA) have been developed as an analytical method with fast, high sensitivity, high accuracy and high precision for medical research purposes to determine the trace elements in the rat's tissue sample. Determination of the trace elements Pt and Se have done using bovine liver standard of NBS as a reference. This standard has used also as a tool to verify an equipment and method will be used further. The measurement results shows that using the available equipment and method, the lower limit of detection for Pt was 1 ppm. Distribution of the trace elements Pt and Se in the different rat's tissue previously treated with cisplatin can be determined. The results shows that Pt content in the several rat's tissue previously treated cisplatin plus sodium selenite is much bigger than that previously treated with cisplatin only. (author)

  7. Simultaneous sampling of tissue oxygenation and oxygen consumption in skeletal muscle.

    Science.gov (United States)

    Nugent, William H; Song, Bjorn K; Pittman, Roland N; Golub, Aleksander S

    2016-05-01

    Under physiologic conditions, microvascular oxygen delivery appears to be well matched to oxygen consumption in respiring tissues. We present a technique to measure interstitial oxygen tension (PISFO2) and oxygen consumption (VO2) under steady-state conditions, as well as during the transitions from rest to activity and back. Phosphorescence Quenching Microscopy (PQM) was employed with pneumatic compression cycling to achieve 1 to 10 Hz sampling rates of interstitial PO2 and simultaneous recurrent sampling of VO2 (3/min) in the exteriorized rat spinotrapezius muscle. The compression pressure was optimized to 120-130 mmHg without adverse effect on the tissue preparation. A cycle of 5s compression followed by 15s recovery yielded a resting VO2 of 0.98 ± 0.03 ml O2/100 cm(3)min while preserving microvascular oxygen delivery. The measurement system was then used to assess VO2 dependence on PISFO2 at rest and further tested under conditions of isometric muscle contraction to demonstrate a robust ability to monitor the on-kinetics of tissue respiration and the compensatory changes in PISFO2 during contraction and recovery. The temporal and spatial resolution of this approach is well suited to studies seeking to characterize microvascular oxygen supply and demand in thin tissues. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Analysis of dissected tissues with digital holographic microscopy: quantification of inflammation mediated tissue alteration, influence of sample preparation, and reliability of numerical autofocusing

    Science.gov (United States)

    Kemper, Björn; Lenz, Philipp; Bettenworth, Dominik; Krausewitz, Philipp; Domagk, Dirk; Ketelhut, Steffi

    2015-03-01

    Quantitative phase imaging with digital holographic microscopy (DHM) allows label-free imaging of tissue sections and quantification of the spatial refractive index distribution, which is of interest for applications in digital pathology. We show that DHM allows quantitative imaging of different layers in unstained tissue samples by detection of refractive index changes. In addition, we evaluate the automated refocussing feature of DHM for application on dissected tissues and could achieve highly reproducible holographic autofocusing for unstained and moderately stained samples. Finally, it is demonstrated that in human ulcerative colitis patients the average tissue refractive index is reduced significantly in all parts of the inflamed colonic wall in comparison to patients in remission.

  9. Detection of Flavobacterium psychrophilum from fish tissue and water samples by PCR amplification

    DEFF Research Database (Denmark)

    Wiklund, T.; Madsen, Lone; Bruun, Morten Sichlau

    2000-01-01

    Rainbow trout fry syndrome and cold-water disease, caused by Flavobacterium psychrophilum, are important diseases in farmed salmonids. Some of the presently available techniques for the detection of Fl. psychrophilum are either time consuming or lack sufficient sensitivity. In the present...... investigation, the possible detection of Fl. psychrophilum from fish tissue and water samples was examined using nested PCR with DNA probes against a sequence of the 16S rRNA genes. The DNA was extracted using Chelex(R) 100 chelating resin. The primers, which were tested against strains isolated from diseased...... to be more sensitive than agar cultivation of tissue samples from the brain of rainbow trout injected with Fl. psychrophilum. In non-sterile fresh water seeded with Fl. psychrophilum the detection limit of the PCR- assay was 1.7 cfu in the PCR tube, corresponding to 110 cfu ml(-1) water. The PCR...

  10. Transgenic zebrafish reveal tissue-specific differences in estrogen signaling in response to environmental water samples

    Science.gov (United States)

    Gorelick, Daniel A.; Iwanowicz, Luke R.; Hung, Alice L.; Blazer, Vicki; Halpern, Marnie E.

    2014-01-01

    Background: Environmental endocrine disruptors (EED) are exogenous chemicals that mimic endogenous hormones, such as estrogens. Previous studies using a zebrafish transgenic reporter demonstrated that the EEDs bisphenol A and genistein preferentially activate estrogen receptors (ER) in the larval heart compared to the liver. However, it was not known whether the transgenic zebrafish reporter was sensitive enough to detect estrogens from environmental samples, whether environmental estrogens would exhibit similar tissue-specific effects as BPA and genistein or why some compounds preferentially target receptors in the heart. Methods: We tested surface water samples using a transgenic zebrafish reporter with tandem estrogen response elements driving green fluorescent protein expression (5xERE:GFP). Reporter activation was colocalized with tissue-specific expression of estrogen receptor genes by RNA in situ hybridization. Results: Selective patterns of ER activation were observed in transgenic fish exposed to river water samples from the Mid-Atlantic United States, with several samples preferentially activating receptors in embryonic and larval heart valves. We discovered that tissue-specificity in ER activation is due to differences in the expression of estrogen receptor subtypes. ERα is expressed in developing heart valves but not in the liver, whereas ERβ2 has the opposite profile. Accordingly, subtype-specific ER agonists activate the reporter in either the heart valves or the liver. Conclusion: The use of 5xERE:GFP transgenic zebrafish has revealed an unexpected tissue-specific difference in the response to environmentally relevant estrogenic compounds. Exposure to estrogenic EEDs in utero is associated with adverse health effects, with the potentially unanticipated consequence of targeting developing heart valves.

  11. Concentration of organochlorines in human brain, liver, and adipose tissue autopsy samples from Greenland.

    OpenAIRE

    Dewailly, E; Mulvad, G; Pedersen, H S; Ayotte, P; Demers, A; Weber, J P; Hansen, J C

    1999-01-01

    Organochlorines are persistent lipophilic compounds that accumulate in Inuit people living in circumpolar countries. Organochlorines accumulate as a result of the Inuits' large consumption of sea mammal fat; however, available data are limited to blood lipids, milk fat, and adipose tissue. We report results of organochlorine determination in liver, brain, omental fat, and subcutaneous abdominal fat samples collected from deceased Greenlanders between 1992 and 1994. Eleven chlorinated pesticid...

  12. Loss of Apc heterozygosity and abnormal tissue building in nascent intestinal polyps in mice carrying a truncated Apc gene.

    Science.gov (United States)

    Oshima, M; Oshima, H; Kitagawa, K; Kobayashi, M; Itakura, C; Taketo, M

    1995-05-09

    Mutations in the APC (adenomatous polyposis coli) gene appear to be responsible for not only familial adenomatous polyposis but also many sporadic cases of gastrointestinal cancers. Using homologous recombination in mouse embryonic stem cells, we constructed mice that contained a mutant gene encoding a product truncated at a 716 (Apc delta 716). Mendelian transmission of the gene caused most homozygous mice to die in utero before day 8 of gestation. The heterozygotes developed multiple polyps throughout the intestinal tract, mostly in the small intestine. The earliest polyps arose multifocally during the third week after birth, and new polyps continued to appear thereafter. Surprisingly, every nascent polyp consisted of a microadenoma covered with a layer of the normal villous epithelium. These microadenomas originated from single crypts by forming abnormal outpockets into the inner (lacteal) side of the neighboring villi. We carefully dissected such microadenomas from nascent polyps by peeling off the normal epithelium and determined their genotype by PCR: all microadenomas had already lost the wild-type Apc allele, whereas the mutant allele remained unchanged. These results indicate that loss of heterozygosity followed by formation of intravillous microadenomas is responsible for polyposis in Apc delta 716 intestinal mucosa. It is therefore unlikely that the truncated product interacts directly with the wild-type protein and causes the microadenomas by a dominant negative mechanism.

  13. Distribution of polybrominated diphenyl ethers in Japanese autopsy tissue and body fluid samples.

    Science.gov (United States)

    Hirai, Tetsuya; Fujimine, Yoshinori; Watanabe, Shaw; Nakano, Takeshi

    2012-09-01

    Brominated flame retardants are components of many plastics and are used in products such as cars, textiles, televisions, and personal computers. Human exposure to polybrominated diphenyl ether (PBDE) flame retardants has increased exponentially during the last three decades. Our objective was to measure the body burden and distribution of PBDEs and to determine the concentrations of the predominant PBDE congeners in samples of liver, bile, adipose tissue, and blood obtained from Japanese autopsy cases. Tissues and body fluids obtained from 20 autopsy cases were analyzed. The levels of 25 PBDE congeners, ranging from tri- to hexa-BDEs, were assessed. The geometric means of the sum of the concentrations of PBDE congeners having detection frequencies >50 % (ΣPBDE) in the blood, liver, bile, and adipose tissue were 2.4, 2.6, 1.4, and 4.3 ng/g lipid, respectively. The most abundant congeners were BDE-47 and BDE-153, followed by BDE-100, BDE-99, and BDE-28+33. These concentrations of PBDE congeners were similar to other reports of human exposure in Japan but were notably lower than concentrations than those reported in the USA. Significant positive correlations were observed between the concentrations of predominant congeners and ΣPBDE among the samples analyzed. The ΣPBDE concentration was highest in the adipose tissue, but PBDEs were distributed widely among the tissues and body fluids analyzed. The PBDE levels observed in the present study are similar to those reported in previous studies in Japan and significantly lower than those reported in the USA.

  14. Evaluation of sample preparation methods and optimization of nickel determination in vegetable tissues

    Directory of Open Access Journals (Sweden)

    Rodrigo Fernando dos Santos Salazar

    2011-02-01

    Full Text Available Nickel, although essential to plants, may be toxic to plants and animals. It is mainly assimilated by food ingestion. However, information about the average levels of elements (including Ni in edible vegetables from different regions is still scarce in Brazil. The objectives of this study were to: (a evaluate and optimize a method for preparation of vegetable tissue samples for Ni determination; (b optimize the analytical procedures for determination by Flame Atomic Absorption Spectrometry (FAAS and by Electrothermal Atomic Absorption (ETAAS in vegetable samples and (c determine the Ni concentration in vegetables consumed in the cities of Lorena and Taubaté in the Vale do Paraíba, State of São Paulo, Brazil. By means of the analytical technique for determination by ETAAS or FAAS, the results were validated by the test of analyte addition and recovery. The most viable method tested for quantification of this element was HClO4-HNO3 wet digestion. All samples but carrot tissue collected in Lorena contained Ni levels above the permitted by the Brazilian Ministry of Health. The most disturbing results, requiring more detailed studies, were the Ni concentrations measured in carrot samples from Taubaté, where levels were five times higher than permitted by Brazilian regulations.

  15. Threshold-dependent sample sizes for selenium assessment with stream fish tissue

    Science.gov (United States)

    Hitt, Nathaniel P.; Smith, David R.

    2015-01-01

    Natural resource managers are developing assessments of selenium (Se) contamination in freshwater ecosystems based on fish tissue concentrations. We evaluated the effects of sample size (i.e., number of fish per site) on the probability of correctly detecting mean whole-body Se values above a range of potential management thresholds. We modeled Se concentrations as gamma distributions with shape and scale parameters fitting an empirical mean-to-variance relationship in data from southwestern West Virginia, USA (63 collections, 382 individuals). We used parametric bootstrapping techniques to calculate statistical power as the probability of detecting true mean concentrations up to 3 mg Se/kg above management thresholds ranging from 4 to 8 mg Se/kg. Sample sizes required to achieve 80% power varied as a function of management thresholds and Type I error tolerance (α). Higher thresholds required more samples than lower thresholds because populations were more heterogeneous at higher mean Se levels. For instance, to assess a management threshold of 4 mg Se/kg, a sample of eight fish could detect an increase of approximately 1 mg Se/kg with 80% power (given α = 0.05), but this sample size would be unable to detect such an increase from a management threshold of 8 mg Se/kg with more than a coin-flip probability. Increasing α decreased sample size requirements to detect above-threshold mean Se concentrations with 80% power. For instance, at an α-level of 0.05, an 8-fish sample could detect an increase of approximately 2 units above a threshold of 8 mg Se/kg with 80% power, but when α was relaxed to 0.2, this sample size was more sensitive to increasing mean Se concentrations, allowing detection of an increase of approximately 1.2 units with equivalent power. Combining individuals into 2- and 4-fish composite samples for laboratory analysis did not decrease power because the reduced number of laboratory samples was compensated for by increased

  16. Impact of freezing delay time on tissue samples for metabolomic studies

    Directory of Open Access Journals (Sweden)

    Tonje Husby Haukaas

    2016-01-01

    Full Text Available Introduction: Metabolic profiling of intact tumor tissue by high resolution magic angle spinning (HR MAS MR spectroscopy (MRS provides important biological information possibly useful for clinical diagnosis and development of novel treatment strategies. However, generation of high-quality data requires that sample handling from surgical resection until analysis is performed using systematically validated procedures. In this study, we investigated the effect of post-surgical freezing delay time on global metabolic profiles and stability of individual metabolites in intact tumor tissue.Materials and methods: Tumor tissue samples collected from two patient derived breast cancer xenograft models (n=3 for each model were divided into pieces that were snap-frozen in liquid nitrogen at 0, 15, 30, 60, 90, and 120 minutes after surgical removal. In addition, one sample was analysed immediately, representing the metabolic profile of fresh tissue exposed neither to liquid nitrogen nor to room temperature. We also evaluated the metabolic effect of prolonged spinning during the HR MAS experiments in biopsies from breast cancer patiens (n=14. All samples were analyzed by proton HR MAS MRS on a Bruker Avance DRX600 spectrometer, and changes in metabolic profiles were evaluated using multivariate analysis and linear mixed modeling (LMM. Results: Multivariate analysis showed that the metabolic differences between the two breast cancer models were more prominent than variation caused by freezing delay time. No significant changes in levels of individual metabolites were observed in samples frozen within 30 minutes of resection. After this time point, levels of choline increased whereas ascorbate, creatine and glutathione (GS levels decreased. Freezing had a significant effect on several metabolites, but is an essential procedure for research and biobank purposes. Furthermore, four metabolites (glucose, glycine, glycerophosphocholine and choline were affected by

  17. Implementation of immunohistochemistry on frozen ear notch tissue samples in diagnosis of bovine viral diarrhea virus in persistently infected cattle

    Directory of Open Access Journals (Sweden)

    Bedeković Tomislav

    2011-12-01

    Full Text Available Abstract Background Bovine viral diarrhea is a contagious disease of domestic and wild ruminants and one of the most economically important diseases in cattle. Bovine viral diarrhea virus belongs to the genus Pestivirus, within the family Flaviviridae. The identification and elimination of the persistently infected animals from herds is the initial step in the control and eradication programs. It is therefore necessary to have reliable methods for diagnosis of bovine viral diarrhea virus. One of those methods is immunohistochemistry. Immunohistochemistry on formalin fixed, paraffin embedded tissue is a routine technique in diagnosis of persistently infected cattle from ear notch tissue samples. However, such technique is inappropriate due to complicated tissue fixation process and it requires more days for preparation. On the contrary, immunohistochemistry on frozen tissue was usually applied on organs from dead animals. In this paper, for the first time, the imunohistochemistry on frozen ear notch tissue samples was described. Findings Seventeen ear notch tissue samples were obtained during the period 2008-2009 from persistently infected cattle. Samples were fixed in liquid nitrogen and stored on -20°C until testing. Ear notch tissue samples from all persistently infected cattle showed positive results with good section quality and possibility to determinate type of infected cells. Conclusions Although the number of samples was limited, this study indicated that immunohistochemistry on formalin fixed paraffin embedded tissue can be successfully replaced with immunohistochemistry on frozen ear notch tissue samples in diagnosis of persistently infected cattle.

  18. A single lysis solution for the analysis of tissue samples by different proteomic technologies

    DEFF Research Database (Denmark)

    Gromov, P.; Celis, J.E.; Gromova, I.

    2008-01-01

    -based proteomics (reverse-phase lysate arrays or direct antibody arrays), allowing the direct comparison of qualitative and quantitative data yielded by these technologies when applied to the same samples. The usefulness of the CLB1 solution for gel-based proteomics was further established by 2D PAGE analysis...... taken place in molecular biology, cell biology and genomics there is a pressing need to accelerate the translation of basic discoveries into clinical applications. This need, compounded by mounting evidence that cellular model systems are unable to fully recapitulate all biological aspects of human...... dissease, is driving scientists to increasingly use clinically relevant samples for biomarker and target discovery. Tissues are heterogeneous and as a result optimization of sample preparation is critical for generating accurate, representative, and highly reproducible quantitative data. Although a large...

  19. Intestinal transport and tissue biodistributions of radium: a comparison of RaCl2 and a radium apatite from freshwater mussels

    International Nuclear Information System (INIS)

    Domel, R.U.; Beal, A.M.

    1996-01-01

    Full text: Natural leaching of uranium ore bodies can result in 226 Ra pollution of adjacent waterways and consequent incorporation of radium into the food chain. Mining has the potential to augment this effect. In the Magela flood plain, NT, the freshwater mussel (Velesunio angasi) concentrates radium in its tissues as a phosphate compound. The availability of mussel radium for uptake and tissue incorporation was assessed relative to radium chloride using rats in feed trial experiments. In addition, the influence of age, duration of dosage and various stages after parturition were investigated. A significant amount of mussel radium was available for intestinal uptake. Incorporation of mussel radium into bone was less than that of radium chloride (p = 0.0001) for both adults and juveniles. Mussel radium dose to bone was higher in juveniles (less than 0.5%) than adults (p< 0.01) after 8 weeks dosage. At the completion of a one week treatment, mussel radium and radium chloride content of bone was lowest in old adults. Incorporation of mussel radium into bone rose progressively in juveniles but not in adults over 8 weeks administration. In contrast, radium accumulation in soft tissues in all age groups was greater after mussel radium treatment than after radium chloride treatment. The skin was the tissue which accumulated the greater proportion of the soft tissue dose. Negligible amounts of mussel radium were transferred from the dams to the young by the placenta of late gestation and little was transferred in the milk during 3 weeks of lactation. However, in 3 weeks following weaning, bone and soft tissue uptake of mussel radium exceeded that of all other ages and durations of dosage. Extrapolation of the data from the animal model to humans suggests that eating these mussels, carries with it only a low risk of exceeding the Annual Limit of Intake (ALI) set for members of the public, even in juveniles

  20. Concentrations of 17 elements in the zebra mussel (Dreissena polymorpha), in different tissues of perch (Perca fluviatilis), and in perch intestinal parasites (Acanthocephalus lucii) from the subalpine lake Mondsee, Austria

    Energy Technology Data Exchange (ETDEWEB)

    Sures, B.; Steiner, W.; Rydlo, M.; Taraschewski, H.

    1999-11-01

    Concentrations of the elements Al, Ag, Ba, ca, Cd, Co, Cr, cu, Fe, Ga, Mg, Mn, Ni, Pb, Sr, Tl, and Zn were analyzed by inductively coupled plasma mass spectrometry in the acanthocephalan Acanthocephalus lucii (Mueller); in its host, Perca fluviatilis (L.), and in the soft tissue of the zebra mussel, Dreissena polymorpha (Pallas). All animals were collected from the same sampling site in a subalpine lake, Mondsee, in Austria. Most of the elements were found at significantly higher concentrations in the acanthocephalan than in different tissues (muscle, liver, and intestinal wall) of its perch host. Only Co was concentrated in the liver of perch to a level that was significantly higher than that found in the parasite. Most of the analyzed elements were also present at significantly higher concentrations in A. lucii than in D. polymorpha. Barium and Cr were the only elements recorded at higher concentrations in the mussel compared with the acanthocephalan. Thus, when comparing the accumulation of elements, the acanthocephalans appear to be even more suitable than the zebra mussels in terms of their use in the detection of metal contamination within aquatic biotopes. Spearman correlation analysis revealed that the concentrations of several elements within the parasites decreased with increasing infrapopulation. Furthermore, the levels of some elements in the perch liver were negatively correlated with the weight of A. lucii in the intestine. Thus, it emerged that not only is there competition for elements between acanthocephalans inside the gut but there is also competition for these elements between the host and the parasites. The elevated element concentrations demonstrated here in the parasitic worm A. lucii provide support for further investigations of these common helminthes and of their accumulation properties.

  1. Prions efficiently cross the intestinal barrier after oral administration: Study of the bioavailability, and cellular and tissue distribution in vivo

    Science.gov (United States)

    Urayama, Akihiko; Concha-Marambio, Luis; Khan, Uffaf; Bravo-Alegria, Javiera; Kharat, Vineetkumar; Soto, Claudio

    2016-01-01

    Natural forms of prion diseases frequently originate by oral (p.o.) infection. However, quantitative information on the gastro-intestinal (GI) absorption of prions (i.e. the bioavailability and subsequent biodistribution) is mostly unknown. The main goal of this study was to evaluate the fate of prions after oral administration, using highly purified radiolabeled PrPSc. The results showed a bi-phasic reduction of PrPSc with time in the GI, except for the ileum and colon which showed sustained increases peaking at 3–6 hr, respectively. Plasma and whole blood 125I-PrPSc reached maximal levels by 30 min and 3 hr, respectively, and blood levels were constantly higher than plasma. Upon crossing the GI-tract 125I-PrPSc became associated to blood cells, suggesting that binding to cells decreased the biological clearance of the agent. Size-exclusion chromatography revealed that oligomeric 125I-PrPSc were transported from the intestinal tract, and protein misfolding cyclic amplification showed that PrPSc in organs and blood retained the typical prion self-replicating ability. Pharmacokinetic analysis found the oral bioavailability of 125I-PrPSc to be 33.6%. Interestingly, 125I-PrPSc reached the brain in a quantity equivalent to the minimum amount needed to initiate prion disease. Our findings provide a comprehensive and quantitative study of the fate of prions upon oral infection. PMID:27573341

  2. A workflow to preserve genome-quality tissue samples from plants in botanical gardens and arboreta1

    Science.gov (United States)

    Gostel, Morgan R.; Kelloff, Carol; Wallick, Kyle; Funk, Vicki A.

    2016-01-01

    Premise of the study: Internationally, gardens hold diverse living collections that can be preserved for genomic research. Workflows have been developed for genomic tissue sampling in other taxa (e.g., vertebrates), but are inadequate for plants. We outline a workflow for tissue sampling intended for two audiences: botanists interested in genomics research and garden staff who plan to voucher living collections. Methods and Results: Standard herbarium methods are used to collect vouchers, label information and images are entered into a publicly accessible database, and leaf tissue is preserved in silica and liquid nitrogen. A five-step approach for genomic tissue sampling is presented for sampling from living collections according to current best practices. Conclusions: Collecting genome-quality samples from gardens is an economical and rapid way to make available for scientific research tissue from the diversity of plants on Earth. The Global Genome Initiative will facilitate and lead this endeavor through international partnerships. PMID:27672517

  3. Effort of fast-neutron and X-ray on expression of p53 and CyclinB1 of small intestinal tissue in mice

    International Nuclear Information System (INIS)

    Xu Dongsheng; Liu Fenju; Sun Zhiqiang; Yang Xueqin; Chen Qiu; Zhou Weifang; Chen Ling

    2008-01-01

    In this paper, effect of fast-neutron and X-rays on the expression of P53 and CyclinB1 in small intestinal tissues of mice were investigated. The mice were randomly grouped and sacrificed 6 h after irradiation. Western blot assay was applied to detect the p53 and CyclinB 1 expressions, and flow cytometry was used to monitor changes in the small intestinal cell cycle. The results showed that the expression levels of p53 increased with irradiation dose. The expression levels of CyclinB 1 increased with the dose at first, but decreased when the dose exceeded 0.28 Gy. The results from the flow cytometry showed that the amount of cells in G2/M phase increased with the dose in both occasions. It was concluded that the low doses of fast-neutron and X-rays can make the expression levels of p53 and CyclinB 1 increase, but the expression levels of CyclinB 1 decreased and the G2 arrest occurred with increased doses. (authors)

  4. Microscopy and elemental analysis in tissue samples using computed microtomography with synchrotron x-rays

    International Nuclear Information System (INIS)

    Spanne, P.; Rivers, M.L.

    1988-01-01

    The initial development shows that CMT using synchrotron x-rays can be developed to μm spatial resolution and perhaps even better. This creates a new microscopy technique which is of special interest in morphological studies of tissues, since no chemical preparation or slicing of the sample is necessary. The combination of CMT with spatial resolution in the μm range and elemental mapping with sensitivity in the ppM range results in a new tool for elemental mapping at the cellular level. 7 refs., 1 fig

  5. Internal radiation burden of coal and uranium miners. Radioactivity measurement of pathological tissue samples

    International Nuclear Information System (INIS)

    Kobor, J.

    1994-01-01

    The components of natural radioactivity exposure of miners are the external gamma radiation, inhalation of 222 Rn and its short-halflife daughter products, and exposure to long isotope radiation ( 238 U to 226 Ra, 232 Th, 210 Pb to 210 Po). Pathological tissue samples were examined by alpha and beta radioactivity measurements to determine the contribution of radiation burden from incorporated long-halflife natural isotopes, and to determine the possible fluctuations at various populations of miners. (N.T.) 19 refs.; 7 figs.; 3 tabs

  6. The intestinal T cell response to alpha-gliadin in adult celiac disease is focused on a single deamidated glutamine targeted by tissue transglutaminase

    DEFF Research Database (Denmark)

    Arentz-Hansen, H; Körner, R; Molberg, O

    2000-01-01

    The great majority of patients that are intolerant of wheat gluten protein due to celiac disease (CD) are human histocompatibility leukocyte antigen (HLA)-DQ2(+), and the remaining few normally express HLA-DQ8. These two class II molecules are chiefly responsible for the presentation of gluten...... peptides to the gluten-specific T cells that are found only in the gut of CD patients but not of controls. Interestingly, tissue transglutaminase (tTG)-mediated deamidation of gliadin plays an important role in recognition of this food antigen by intestinal T cells. Here we have used recombinant antigens...... for T cell recognition. Gluten-specific T cell lines from 16 different adult patients all responded to one or both of these deamidated peptides, indicating that these epitopes are highly relevant to disease pathology. Binding studies showed that the deamidated peptides displayed an increased affinity...

  7. Persistent synthetic chlorinated hydrocarbons in albatross tissue samples from Midway Atoll

    Energy Technology Data Exchange (ETDEWEB)

    Jones, P.D.; Hannah, D.J.; Buckland, S.J. [ESR:Environmental, Lower Hutt (New Zealand)] [and others

    1996-10-01

    Anthropogenic organic contaminants have been found in even the most remote locations. To assess the global distribution and possible effects of such contaminants, the authors examined the tissues of two species of albatross collected from Midway Atoll in the central North Pacific Ocean. These birds have an extensive feeding range covering much of the subtropical and northern Pacific Ocean. Anthropogenic contaminants were found at relatively great concentrations in these birds. The sum of 19 polychlorinated biphenyl (PCB) congeners ranged from 177 ng/g wet weight in eggs to 2,750 ng/g wet weight in adult fat. Total toxic equivalents (TEQs) derived from polychlorinated dibenzo-p-dioxins (PCDDs) and dibenzofurans (PCDFs) ranged from 17.2 to 297 pg/g wet weight in the same tissues, while the inclusion of TEQs from PCBs increased these values to 48.4 and 769 pg/g wet weight, respectively. While contaminant concentrations varied between species and tissues, the contaminant profile was relatively uniform. The profile of contaminants detected was unusual in that much of the TEQs was contributed by two pentachlorinated congeners (2,3,4,7,8-pentachlorinated dibenzo-p-dioxin), and the profiles of PCB congeners did not match known sources. When compared to other studies the concentrations detected in the Midway Atoll samples were near or above the thresholds known to cause adverse effects in other fish-eating bird species.

  8. Use of Fluorescein Isothiocyanate-Inulin as a Marker for Intestinal Ischemic Injury.

    Science.gov (United States)

    AlKukhun, Abedalrazaq; Caturegli, Giorgio; Munoz-Abraham, Armando Salim; Judeeba, Sami; Patron-Lozano, Roger; Morotti, Raffaella; Rodriguez-Davalos, Manuel I; Geibel, John P

    2017-06-01

    Intestinal ischemia is observed in conditions such as mesenteric ischemia, or during traumatic events such as intestinal transplantation. Intestinal ischemia leads to pathophysiologic disruptions that present as increased fluid secretion into the intestinal lumen. We propose a novel method to detect real-time ischemic injury that is used in an in vitro model applicable to intestinal transplantation. Small intestine segments from rats were procured. The segments were attached to customized perfusion chambers. Both intestines were perfused on the vascular side with a Ringer buffer solution. The experimental buffer solution was bubbled with 100% nitrogen to mimic ischemia. Both lumens were perfused with 3 mL HEPES-Ringer solution containing 50 μM fluorescein isothiocyanate (FITC)-inulin. Intraluminal samples were collected at 15-minute intervals to measure FITC-inulin concentration using a nanofluorospectrophotometer. Intestinal tissue samples were processed and evaluated by a blinded pathologist using the Park/Chiu scoring system for grading intestinal ischemia. Samples collected from the ischemic intestine showed a significant decrease in FITC-inulin fluorescence compared with the control intestine, indicating enhanced fluid secretion. Histopathologic samples from the experimental arm exhibited higher scores of ischemic injury in comparison with the control arm, confirming the FITC-inulin as a correlation to ischemia. Fluorescein isothiocyanate-inulin can be used as a real-time volume marker to monitor the ischemic state of intestinal tissue. A positive correlation between the degree of fluid shift and presence of ischemic injury. The changes in fluorescence signal provide a potential selective method to measure real-time fluid changes inside an intestinal graft to evaluate viability. Copyright © 2016 American College of Surgeons. Published by Elsevier Inc. All rights reserved.

  9. Intestinal Cancer

    Science.gov (United States)

    ... connects your stomach to your large intestine. Intestinal cancer is rare, but eating a high-fat diet ... increase your risk. Possible signs of small intestine cancer include Abdominal pain Weight loss for no reason ...

  10. Biomarker discovery in heterogeneous tissue samples -taking the in-silico deconfounding approach

    Directory of Open Access Journals (Sweden)

    Parida Shreemanta K

    2010-01-01

    Full Text Available Abstract Background For heterogeneous tissues, such as blood, measurements of gene expression are confounded by relative proportions of cell types involved. Conclusions have to rely on estimation of gene expression signals for homogeneous cell populations, e.g. by applying micro-dissection, fluorescence activated cell sorting, or in-silico deconfounding. We studied feasibility and validity of a non-negative matrix decomposition algorithm using experimental gene expression data for blood and sorted cells from the same donor samples. Our objective was to optimize the algorithm regarding detection of differentially expressed genes and to enable its use for classification in the difficult scenario of reversely regulated genes. This would be of importance for the identification of candidate biomarkers in heterogeneous tissues. Results Experimental data and simulation studies involving noise parameters estimated from these data revealed that for valid detection of differential gene expression, quantile normalization and use of non-log data are optimal. We demonstrate the feasibility of predicting proportions of constituting cell types from gene expression data of single samples, as a prerequisite for a deconfounding-based classification approach. Classification cross-validation errors with and without using deconfounding results are reported as well as sample-size dependencies. Implementation of the algorithm, simulation and analysis scripts are available. Conclusions The deconfounding algorithm without decorrelation using quantile normalization on non-log data is proposed for biomarkers that are difficult to detect, and for cases where confounding by varying proportions of cell types is the suspected reason. In this case, a deconfounding ranking approach can be used as a powerful alternative to, or complement of, other statistical learning approaches to define candidate biomarkers for molecular diagnosis and prediction in biomedicine, in

  11. Correlates of professional burnout in a sample of employees of cell and tissue banks in Poland.

    Science.gov (United States)

    Kamiński, Artur; Rozenek, Hanna; Banasiewicz, Jolanta; Wójtowicz, Stanisław; Błoński, Artur; Owczarek, Krzysztof

    2018-02-03

    Job Demands-Resources model proposes that the development of burnout follows excessive job demands and lack of job resources. Job demands are predictive of feeling of exhaustion, and lack of job resources-disengagement from work. This pilot study investigated professional burnout and its correlates in employees of Polish cell and tissue banks, many of whom were involved in procurement and processing of tissues from deceased donors, as it was hypothesized that job burnout in this population might influence the effectiveness of cell and tissue transplantation network in our country. This study utilized the Polish version of the Oldenburg Burnout Inventory (OLBI), which measures the two dimensions of burnout (exhaustion and disengagement), and the Psychosocial Working Conditions Questionnaire (PWC), a Polish instrument used for monitoring psychosocial stress at work. The study sample consisted of 31 participants. Their average time of working in a cell and tissue bank was 13.20 years. Majority of the PWC scales and subscales scores fell in the Average range, and the OLBI results for the Disengagement and the Exhaustion scales were in the Average range. A number of correlations between the Exhaustion or Disengagement and the PWC scales and subscales were detected, majority of which fell in the Moderate range. In spite of the limited number of participants, the results of this pilot study are consistent with the burnout literature reports. Among the detected correlates of professional burnout, it is job-related support which seems to be the most important factor which may influence the efficacy of transplantation network in Poland.

  12. Liquid Microjunction Surface Sampling Probe Electrospray Mass Spectrometry for Detection of Drugs and Metabolites in Thin Tissue Sections

    Energy Technology Data Exchange (ETDEWEB)

    Van Berkel, Gary J [ORNL; Kertesz, Vilmos [ORNL; Koeplinger, Kenneth A. [Merck Research Laboratories; Vavek, Marissa [Merck Research Laboratories; Kong, Ah-Ng Tony [Rutgers University

    2008-01-01

    A self-aspirating, liquid micro-junction surface sampling probe/electrospray emitter mass spectrometry system was demonstrated for use in the direct analysis of spotted and dosed drugs and their metabolites in thin tissue sections. Proof-of-principle sampling and analysis directly from tissue without the need for sample preparation was demonstrated first by raster scanning a region on a section of rat liver onto which reserpine was spotted. The mass spectral signal from selected reaction monitoring was used to develop a chemical image of the spotted drug on the tissue. The probe was also used to selectively spot sample areas of sagittal whole mouse body tissue sections that had been dosed orally (90 mg/kg) with R,S-sulforaphane 3 hrs prior to sacrifice. Sulforaphane and its glutathione and N-acetyl cysteine conjugates were monitored with selected reaction monitoring and detected in the stomach and various other tissues from the dosed mouse. No signal for these species was observed in the tissue from a control mouse. The same dosed tissue section was used to illustrate the possibility of obtaining a line scan across the whole body section. In total these results illustrate the potential for rapid screening of the distribution of drugs and metabolites in tissue sections with the micro-liquid junction surface sampling probe/electrospray mass spectrometry approach.

  13. Multimodal Raman-fluorescence spectroscopy of formalin fixed samples is able to discriminate brain tumors from dysplastic tissue

    Science.gov (United States)

    Anand, Suresh; Cicchi, Riccardo; Giordano, Flavio; Buccoliero, Anna Maria; Pavone, Francesco Saverio

    2014-05-01

    In the recent years, there has been a considerable surge in the application of spectroscopy for disease diagnosis. Raman and fluorescence spectra provide characteristic spectral profile related to biochemical and morphological changes when tissues progress from normal state towards malignancy. Spectroscopic techniques offer the advantage of being minimally invasive compared to traditional histopathology, real time and quantitative. In biomedical optical diagnostics, freshly excised specimens are preferred for making ex-vivo spectroscopic measurements. With regard to fresh tissues, if the lab is located far away from the clinic it could pose a problem as spectral measurements have to be performed immediately after dissection. Tissue samples are usually placed in a fixative agent such as 4% formaldehyde to preserve the samples before processing them for routine histopathological studies. Fixation prevents the tissues from decomposition by arresting autolysis. In the present study, we intend to investigate the possibility of using formalin fixed samples for discrimination of brain tumours from dysplastic tissue using Raman and fluorescence spectroscopy. Formalin fixed samples were washed with phosphate buffered saline for about 5 minutes in order to remove the effects of formalin during spectroscopic measurements. In case of fluorescence spectroscopy, changes in spectral profile have been observed in the region between 550-670 nm between dysplastic and tumor samples. For Raman measurements, we found significant differences in the spectral profiles between dysplasia and tumor. In conclusion, formalin fixed samples can be potentially used for the spectroscopic discrimination of tumor against dysplastic tissue in brain samples.

  14. Gold sputtered fiducial markers for combined secondary ion mass spectrometry and MALDI imaging of tissue samples.

    Science.gov (United States)

    Ogrinc Potočnik, Nina; Škrášková, Karolina; Flinders, Bryn; Pelicon, Primož; Heeren, Ron M A

    2014-07-15

    Mass spectrometry imaging (MSI) is a label free technique capable of providing simultaneous identification and localization of biomolecules. A multimodal approach is required that allows for the study of the complexity of biological tissue samples to overcome the limitations of a single MSI technique. Secondary ion mass spectrometry (SIMS) allows for high spatial resolution imaging while matrix-assisted laser desorption (MALDI) offers a significantly wider mass range. The combination of coregistered SIMS and MALDI images results in detailed and unique biomolecular information. In this Technical Note, we describe how gold sputtered/implanted fiducial markers (FM) are created and can be used to ensure a proper overlay and coregistration of the two-dimensional images provided by the two MSI modalities.

  15. Leptine: an hormone secreted by adipose tissue. First study in Uruguayan population sample

    International Nuclear Information System (INIS)

    Pisabarro, Raul; Irrazabal, Ernesto; Recalde, Alicia; Barrios, Enrique; Arocena, Beatriz; Garcia Loriente, Jose Maria; Lorenzo Bonifazio, Juan

    1999-01-01

    The recent discovery of leptine, an hormone secreted by adipose tissue which modulates the energetic expenditure has signified a gigantic advance in studying obesity facts. In spite of a recent description of absence of leptine in humans, the obesity human model answers to leptine resistance. In this paper, we revise the actual concepts and show leptine values of a sample of 101 middle aged uruguayans, male and female, of normal weight and over weighted (table 1), correlated with corporal mass index (CMI) as an indirect measure of total body fat and waist diameter as an indirect measure of visceral fat, and hips (periferical fat). Bioimpedance studies were carried out to get the corporal composition. Results: good correlation between corporal fat and leptine, but fat distribution was not found representative. All in all, this data set confirms the correlation between leptine and total body fat mass

  16. A probe-based quantitative PCR assay for detecting Tetracapsuloides bryosalmonae in fish tissue and environmental DNA water samples

    Science.gov (United States)

    Hutchins, Patrick; Sepulveda, Adam; Martin, Renee; Hopper, Lacey

    2017-01-01

    A probe-based quantitative real-time PCR assay was developed to detect Tetracapsuloides bryosalmonae, which causes proliferative kidney disease in salmonid fish, in kidney tissue and environmental DNA (eDNA) water samples. The limits of detection and quantification were 7 and 100 DNA copies for calibration standards and T. bryosalmonae was reliably detected down to 100 copies in tissue and eDNA samples. The assay presented here is a highly sensitive and quantitative tool for detecting T. bryosalmonae with potential applications for tissue diagnostics and environmental detection.

  17. Microscopic diagnosis of sodium acetate-acetic acid-formalin-fixed stool samples for helminths and intestinal protozoa: a comparison among European reference laboratories.

    Science.gov (United States)

    Utzinger, J; Botero-Kleiven, S; Castelli, F; Chiodini, P L; Edwards, H; Köhler, N; Gulletta, M; Lebbad, M; Manser, M; Matthys, B; N'Goran, E K; Tannich, E; Vounatsou, P; Marti, H

    2010-03-01

    The present study aimed to compare the diagnostic performance of different European reference laboratories in diagnosing helminths and intestinal protozoa, using an ether-concentration method applied to sodium acetate-acetic acid-formalin (SAF)-preserved faecal samples. In total, 102 stool specimens were analysed during a cross-sectional parasitological survey in urban farming communities in Côte d'Ivoire. Five SAF-preserved faecal samples were prepared from each specimen and forwarded to the participating reference laboratories, processed and examined under a microscope adhering to a standard operating procedure (SOP). Schistosoma mansoni (cumulative prevalence: 51.0%) and hookworm (cumulative prevalence: 39.2%) were the predominant helminths. There was excellent agreement (kappa > 0.8; p protozoa were Entamoeba coli (median prevalence: 67.6%), Blastocystis hominis (median prevalence: 55.9%) and Entamoeba histolytica/Entamoeba dispar (median prevalence: 47.1%). Substantial agreement among reference laboratories was found for E. coli (kappa = 0.69), but only fair or moderate agreement was found for other Entamoeba species, Giardia intestinalis and Chilomastix mesnili. There was only poor agreement for B. hominis, Isospora belli and Trichomonas intestinalis. In conclusion, although common helminths were reliably diagnosed by European reference laboratories, there was only moderate agreement between centres for pathogenic intestinal protozoa. Continued external quality assessment and the establishment of a formal network of reference laboratories is necessary to further enhance both accuracy and uniformity in parasite diagnosis.

  18. Comparing the proteome of snap frozen, RNAlater preserved, and formalin-fixed paraffin-embedded human tissue samples

    DEFF Research Database (Denmark)

    Bennike, Tue Bjerg; Kastaniegaard, Kenneth; Padurariu, Simona

    2016-01-01

    Large biobanks exist worldwide containing formalin-fixed, paraffin-embedded samples and samples stored in RNAlater. However, the impact of tissue preservation on the result of a quantative proteome analysis remains poorly described.Human colon mucosal biopsies were extracted from the sigmoideum...

  19. Prophylactic Ozone Administration Reduces Intestinal Mucosa Injury Induced by Intestinal Ischemia-Reperfusion in the Rat

    Directory of Open Access Journals (Sweden)

    Ozkan Onal

    2015-01-01

    Full Text Available Objectives. Intestinal ischemia-reperfusion injury is associated with mucosal damage and has a high rate of mortality. Various beneficial effects of ozone have been shown. The aim of the present study was to show the effects of ozone in ischemia reperfusion model in intestine. Material and Method. Twenty eight Wistar rats were randomized into four groups with seven rats in each group. Control group was administered serum physiologic (SF intraperitoneally (ip for five days. Ozone group was administered 1 mg/kg ozone ip for five days. Ischemia Reperfusion (IR group underwent superior mesenteric artery occlusion for one hour and then reperfusion for two hours. Ozone + IR group was administered 1 mg/kg ozone ip for five days and at sixth day IR model was applied. Rats were anesthetized with ketamine∖xyzlazine and their intracardiac blood was drawn completely and they were sacrificed. Intestinal tissue samples were examined under light microscope. Levels of superoxide dismutase (SOD, catalase (CAT, glutathioneperoxidase (GSH-Px, malondyaldehide (MDA, and protein carbonyl (PCO were analyzed in tissue samples. Total oxidant status (TOS, and total antioxidant capacity (TAC were analyzed in blood samples. Data were evaluated statistically by Kruskal Wallis test. Results. In the ozone administered group, degree of intestinal injury was not different from the control group. IR caused an increase in intestinal injury score. The intestinal epithelium maintained its integrity and decrease in intestinal injury score was detected in Ozone + IR group. SOD, GSH-Px, and CAT values were high in ozone group and low in IR. TOS parameter was highest in the IR group and the TAC parameter was highest in the ozone group and lowest in the IR group. Conclusion. In the present study, IR model caused an increase in intestinal injury.In the present study, ozone administration had an effect improving IR associated tissue injury. In the present study, ozone therapy

  20. Effect of dietary organic zinc sources on growth performance, incidence of diarrhoea, serum and tissue zinc concentrations, and intestinal morphology in growing rabbits

    Directory of Open Access Journals (Sweden)

    J.Y. Yan

    2017-03-01

    Full Text Available This study was conducted to evaluate the effect of dietary organic zinc (Zn sources on growth performance, the incidence of diarrhoea, serum and tissue Zn concentration, and intestinal morphology in growing rabbits. A total of 120 New Zealand White rabbits aged 35 d and with an initial body weight of 755±15 g, were randomly divided into 4 treatment groups for a 49 d feeding trial. Dietary treatments were designed with different Zn supplements as follows: (1 Control group: 80 mg/kg Zn as ZnSO4; (2 ZnLA group: 80 mg/kg Zn as Zn lactate; (3 ZnMet group: 80 mg/kg Zn as Zn methionine; (4 ZnGly group: 80 mg/kg Zn as Zn glycine. The results showed that, when compared with rabbits fed ZnSO4, supplementation with ZnLA improved (P4. Supplementing with ZnLA increased duodenum villi height (681.63 vs. 587.14 μm, P4, except that feeding ZnMet led to higher (P4. The results indicated that supplementation with 80 mg/kg Zn as ZnLA could improve growth performance, increase liver Zn concentration and enhance duodenum morphology, while reducing the incidence of diarrhoea in growing rabbits.

  1. Localization of serotoni (5-hydroxytryptamine, 5-HT) with partial purification and characterization of a serotonin binding protein in the intestinal tissue of the nematode Ascaris suum

    Energy Technology Data Exchange (ETDEWEB)

    Martin, R.E.

    1989-01-01

    An intracellular 5-HT binding protein (SBP) from intestinal tissue was partially purified and characterized. Binding of ({sup 3}H) 5-HT to the protein appeared to be Fe{sup +2}-sensitive and maximal (20.8pmol/mg protein) at 5 {times} 10{sup {minus}4}M Fe{sup +2} and 10{sup {minus}7}M ({sup 3}H) 5-HT. There were two 5-HT binding sites present at optimum Fe{sup +2} concentrations. The Bmax values of these sites were more sensitive to Fe{sup +2} than Kd values. Sulfhydryl reducing agents, cation chelators, Fe{sup +3}, Ca{sup +2} and antagonists of 5-HT uptake and storage inhibited binding of 5-HT to SBP. Gel exclusion chromatography indicated the presence of a 45Kda SBP that in 5 {times} 10{sup {minus}5}M Fe{sup +2} may form aggregates ranging in size from approximately 80 to >1000Kda. The data indicate these in vitro aggregates may correspond to the electron-opaque patches observed in situ. Ascaris suum may provide a model system to further elucidate the physiological role of analogous serotonin binding proteins that have been identified in mammalian systems.

  2. Gene expression profiling of human breast tissue samples using SAGE-Seq.

    Science.gov (United States)

    Wu, Zhenhua Jeremy; Meyer, Clifford A; Choudhury, Sibgat; Shipitsin, Michail; Maruyama, Reo; Bessarabova, Marina; Nikolskaya, Tatiana; Sukumar, Saraswati; Schwartzman, Armin; Liu, Jun S; Polyak, Kornelia; Liu, X Shirley

    2010-12-01

    We present a powerful application of ultra high-throughput sequencing, SAGE-Seq, for the accurate quantification of normal and neoplastic mammary epithelial cell transcriptomes. We develop data analysis pipelines that allow the mapping of sense and antisense strands of mitochondrial and RefSeq genes, the normalization between libraries, and the identification of differentially expressed genes. We find that the diversity of cancer transcriptomes is significantly higher than that of normal cells. Our analysis indicates that transcript discovery plateaus at 10 million reads/sample, and suggests a minimum desired sequencing depth around five million reads. Comparison of SAGE-Seq and traditional SAGE on normal and cancerous breast tissues reveals higher sensitivity of SAGE-Seq to detect less-abundant genes, including those encoding for known breast cancer-related transcription factors and G protein-coupled receptors (GPCRs). SAGE-Seq is able to identify genes and pathways abnormally activated in breast cancer that traditional SAGE failed to call. SAGE-Seq is a powerful method for the identification of biomarkers and therapeutic targets in human disease.

  3. Grinding and polishing instead of sectioning for the tissue samples with a graft: Implications for light and electron microscopy.

    Science.gov (United States)

    Mukhamadiyarov, Rinat A; Sevostyanova, Victoria V; Shishkova, Daria K; Nokhrin, Andrey V; Sidorova, Olga D; Kutikhin, Anton G

    2016-06-01

    A broad use of the graft replacement requires a detailed investigation of the host-graft interaction, including both histological examination and electron microscopy. A high quality sectioning of the host tissue with a graft seems to be complicated; in addition, it is difficult to examine the same tissue area by both of the mentioned microscopy techniques. To solve these problems, we developed a new technique of epoxy resin embedding with the further grinding, polishing, and staining. Graft-containing tissues prepared by grinding and polishing preserved their structure; however, sectioning frequently required the explantation of the graft and led to tissue disintegration. Moreover, stained samples prepared by grinding and polishing may then be assessed by both light microscopy and backscattered scanning electron microscopy. Therefore, grinding and polishing outperform sectioning when applied to the tissues with a graft. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Intestinal Obstruction

    Science.gov (United States)

    ... the obstruction along the intestines. Treatment Suction via nasogastric tube Fluids given by vein Surgery for strangulation Sometimes ... nose and placed in the stomach (called a nasogastric tube) or into the intestine. Suction is applied to ...

  5. Normalization of gene expression measurement of tissue samples obtained by transurethral resection of bladder tumors

    Directory of Open Access Journals (Sweden)

    Pop LA

    2016-06-01

    Full Text Available Laura A Pop,1,* Valentina Pileczki,1,2,* Roxana M Cojocneanu-Petric,1 Bogdan Petrut,3,4 Cornelia Braicu,1 Ancuta M Jurj,1 Rares Buiga,5 Patriciu Achimas-Cadariu,6,7 Ioana Berindan-Neagoe1,8 1The Research Center for Functional Genomics, Biomedicine and Translational Medicine, Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-Napoca, Cluj, Romania; 2Department of Analytical Chemistry, Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-Napoca, Cluj, Romania; 3Department of Surgery II – Urology, The Oncology Institute “Prof Dr Ion Chiricuţă”, Cluj-Napoca, Cluj, Romania; 4Department of Urology, Iuliu Haţieganu University of Medicine and Pharmacy, Cluj-Napoca, Cluj, Romania; 5Department of Pathology, The Oncology Institute “Prof. Dr Ion Chiricuţă”, Cluj-Napoca, Cluj, Romania; 6Department of Surgery, The Oncology Institute “Prof Dr Ion Chiricuţă”, Cluj-Napoca, Cluj, Romania; 7Department of Surgical Oncology and Gynecological Oncology, Iuliu Haţieganu University of Medicine and Pharmacy, 8Department of Functional Genomics and Experimental Pathology, The Oncology Institute “Prof Dr Ion Chiricuţă”, Cluj-Napoca, Cluj, Romania *These authors contributed equally to this work Background: Sample processing is a crucial step for all types of genomic studies. A major challenge for researchers is to understand and predict how RNA quality affects the identification of transcriptional differences (by introducing either false-positive or false-negative errors. Nanotechnologies help improve the quality and quantity control for gene expression studies. Patients and methods: The study was performed on 14 tumor and matched normal pairs of tissue from patients with bladder urothelial carcinomas. We assessed the RNA quantity by using the NanoDrop spectrophotometer and the quality by nano-microfluidic capillary electrophoresis technology provided by Agilent 2100 Bioanalyzer. We evaluated the amplification status of three

  6. A comparison of sample preparation strategies for biological tissues and subsequent trace element analysis using LA-ICP-MS.

    Science.gov (United States)

    Bonta, Maximilian; Török, Szilvia; Hegedus, Balazs; Döme, Balazs; Limbeck, Andreas

    2017-03-01

    Laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) is one of the most commonly applied methods for lateral trace element distribution analysis in medical studies. Many improvements of the technique regarding quantification and achievable lateral resolution have been achieved in the last years. Nevertheless, sample preparation is also of major importance and the optimal sample preparation strategy still has not been defined. While conventional histology knows a number of sample pre-treatment strategies, little is known about the effect of these approaches on the lateral distributions of elements and/or their quantities in tissues. The technique of formalin fixation and paraffin embedding (FFPE) has emerged as the gold standard in tissue preparation. However, the potential use for elemental distribution studies is questionable due to a large number of sample preparation steps. In this work, LA-ICP-MS was used to examine the applicability of the FFPE sample preparation approach for elemental distribution studies. Qualitative elemental distributions as well as quantitative concentrations in cryo-cut tissues as well as FFPE samples were compared. Results showed that some metals (especially Na and K) are severely affected by the FFPE process, whereas others (e.g., Mn, Ni) are less influenced. Based on these results, a general recommendation can be given: FFPE samples are completely unsuitable for the analysis of alkaline metals. When analyzing transition metals, FFPE samples can give comparable results to snap-frozen tissues. Graphical abstract Sample preparation strategies for biological tissues are compared with regard to the elemental distributions and average trace element concentrations.

  7. Measurement of microparticle tissue factor activity in clinical samples: A summary of two tissue factor-dependent FXa generation assays.

    Science.gov (United States)

    Hisada, Yohei; Alexander, Wyeth; Kasthuri, Raj; Voorhees, Peter; Mobarrez, Fariborz; Taylor, Angela; McNamara, Coleen; Wallen, Hakan; Witkowski, Marco; Key, Nigel S; Rauch, Ursula; Mackman, Nigel

    2016-03-01

    Thrombosis is a leading cause of morbidity and mortality. Detection of a prothrombotic state using biomarkers would be of great benefit to identify patients at risk of thrombosis that would benefit from thromboprophylaxis. Tissue factor (TF) is a highly procoagulant protein that under normal conditions is not present in the blood. However, increased levels of TF in the blood in the form of microparticles (MPs) (also called extracellular vesicles) are observed under various pathological conditions. In this review, we will discuss studies that have measured MP-TF activity in a variety of diseases using two similar FXa generation assay. One of the most robust signals for MP-TF activity (16-26 fold higher than healthy controls) is observed in pancreatic cancer patients with venous thromboembolism. In this case, the TF+ MPs appear to be derived from the cancer cells. Surprisingly, cirrhosis and acute liver injury are associated with 17-fold and 38-fold increases in MP-TF activity, respectively. Based on mouse models, we speculate that the TF+ MPs are derived from hepatocytes. More modest increases are observed in patients with urinary tract infections (6-fold) and in a human endotoxemia model (9-fold) where monocytes are the likely source of the TF+ MPs. Finally, there is no increase in MP-TF activity in the majority of cardiovascular disease patients. These studies indicate that MP-TF activity may be a useful biomarker to identify patients with particular diseases that have an increased risk of thrombosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Epigenome-wide profiling of DNA methylation in paired samples of adipose tissue and blood.

    Science.gov (United States)

    Huang, Yen-Tsung; Chu, Su; Loucks, Eric B; Lin, Chien-Ling; Eaton, Charles B; Buka, Stephen L; Kelsey, Karl T

    2016-03-03

    Many epigenetic association studies have attempted to identify DNA methylation markers in blood that are able to mirror those in target tissues. Although some have suggested potential utility of surrogate epigenetic markers in blood, few studies have collected data to directly compare DNA methylation across tissues from the same individuals. Here, epigenomic data were collected from adipose tissue and blood in 143 subjects using Illumina HumanMethylation450 BeadChip array. The top axis of epigenome-wide variation differentiates adipose tissue from blood, which is confirmed internally using cross-validation and externally with independent data from the two tissues. We identified 1,285 discordant genes and 1,961 concordant genes between blood and adipose tissue. RNA expression data of the two classes of genes show consistent patterns with those observed in DNA methylation. The discordant genes are enriched in biological functions related to immune response, leukocyte activation or differentiation, and blood coagulation. We distinguish the CpG-specific correlation from the within-subject correlation and emphasize that the magnitude of within-subject correlation does not guarantee the utility of surrogate epigenetic markers. The study reinforces the critical role of DNA methylation in regulating gene expression and cellular phenotypes across tissues, and highlights the caveats of using methylation markers in blood to mirror the corresponding profile in the target tissue.

  9. Long-term effects of Ag-containing alloys on mucous tissue present in biopsy samples.

    Science.gov (United States)

    Aoyagi, Hidekazu; Katagiri, Masataka

    2004-09-01

    The aim of this study was to investigate the long-term effects of alloys containing silver (mainly Ag-Sn alloy) on oral mucous tissue. We observed biopsy tissue specimens from patients diagnosed as having amalgam tattoo and/or metal pigmentation by light and electron microscopy and electron-probe microanalysis (EPMA). In most cases, Ag-Sn alloy was present in the tissue but it could not be confirmed if the alloy originated from amalgam. Distributions of both Ag-S and Ag-Sn have typical patterns. Most Ag forms Ag2S and is stably deposited in three patterns along the collagen, basement membrane, and fibrous cells without inducing any host reaction. On the other hand, Sn forms large granules that contain Ag, S, C, N, P, and Ca, and is in soft state in the tissue. Tissue reactions to the alloy become weaker as time passes.

  10. Hippo signalling directs intestinal fate

    DEFF Research Database (Denmark)

    le Bouteiller, Marie Catherine M; Jensen, Kim Bak

    2015-01-01

    Hippo signalling has been associated with many important tissue functions including the regulation of organ size. In the intestinal epithelium differing functions have been proposed for the effectors of Hippo signalling, YAP and TAZ1. These are now shown to have a dual role in the intestinal epit...

  11. Comparison of ESR1 Mutations in Tumor Tissue and Matched Plasma Samples from Metastatic Breast Cancer Patients

    Directory of Open Access Journals (Sweden)

    Takashi Takeshita

    2017-10-01

    Full Text Available BACKGROUND: ESR1 mutation in circulating cell-free DNA (cfDNA is emerging as a noninvasive biomarker of acquired resistance to endocrine therapy, but there is a paucity of data comparing the status of ESR1 gene in cfDNA with that in its corresponding tumor tissue. The objective of this study is to validate the degree of concordance of ESR1 mutations between plasma and tumor tissue. METHODS: ESR1 ligand-binding domain mutations Y537S, Y537N, Y537C, and D538G were analyzed using droplet digital PCR in 35 patients with metastatic breast cancer (MBC (35 tumor tissue samples and 67 plasma samples. RESULTS: Of the 35 paired samples, 26 (74.3% were concordant: one patient had detectable ESR1 mutations both plasma (ESR1 Y537S/Y537N and tumor tissue (ESR1 Y537S/Y537C, and 25 had WT ESR1 alleles in both. Nine (25.7% had discordance between the plasma and tissue results: five had mutations detected only in their tumor tissue (two Y537S, one Y537C, one D538G, and one Y537S/Y537N/D538G, and four had mutations detected only in their plasma (one Y537S, one Y537N, and two Y537S/Y537N/D538G. Furthermore, longitudinal plasma samples from 19 patients were used to assess changes in the presence of ESR1 mutations during treatment. Eleven patients had cfDNA ESR1 mutations over the course of treatment. A total of eight of 11 patients with MBC with cfDNA ESR1 mutations (72.7% had the polyclonal mutations. CONCLUSION: We have shown the independent distribution of ESR1 mutations between plasma and tumor tissue in 35 patients with MBC.

  12. Effects of Digested Onion Extracts on Intestinal Gene Expression: An Interspecies Comparison Using Different Intestine Models.

    Directory of Open Access Journals (Sweden)

    Nicole J W de Wit

    Full Text Available Human intestinal tissue samples are barely accessible to study potential health benefits of nutritional compounds. Numbers of animals used in animal trials, however, need to be minimalized. Therefore, we explored the applicability of in vitro (human Caco-2 cells and ex vivo intestine models (rat precision cut intestine slices and the pig in-situ small intestinal segment perfusion (SISP technique to study the effect of food compounds. In vitro digested yellow (YOd and white onion extracts (WOd were used as model food compounds and transcriptomics was applied to obtain more insight into which extent mode of actions depend on the model. The three intestine models shared 9,140 genes which were used to compare the responses to digested onions between the models. Unsupervised clustering analysis showed that genes up- or down-regulated by WOd in human Caco-2 cells and rat intestine slices were similarly regulated by YOd, indicating comparable modes of action for the two onion species. Highly variable responses to onion were found in the pig SISP model. By focussing only on genes with significant differential expression, in combination with a fold change > 1.5, 15 genes showed similar onion-induced expression in human Caco-2 cells and rat intestine slices and 2 overlapping genes were found between the human Caco-2 and pig SISP model. Pathway analyses revealed that mainly processes related to oxidative stress, and especially the Keap1-Nrf2 pathway, were affected by onions in all three models. Our data fit with previous in vivo studies showing that the beneficial effects of onions are mostly linked to their antioxidant properties. Taken together, our data indicate that each of the in vitro and ex vivo intestine models used in this study, taking into account their limitations, can be used to determine modes of action of nutritional compounds and can thereby reduce the number of animals used in conventional nutritional intervention studies.

  13. Effects of Digested Onion Extracts on Intestinal Gene Expression: An Interspecies Comparison Using Different Intestine Models.

    Science.gov (United States)

    de Wit, Nicole J W; Hulst, Marcel; Govers, Coen; van der Meulen, Jan; van Hoef, Angeline; Stoopen, Geert; Hamers, Astrid; Hoekman, Arjan; de Vos, Ric; Bovee, Toine F H; Smits, Mari; Mes, Jurriaan J; Hendriksen, Peter J M

    2016-01-01

    Human intestinal tissue samples are barely accessible to study potential health benefits of nutritional compounds. Numbers of animals used in animal trials, however, need to be minimalized. Therefore, we explored the applicability of in vitro (human Caco-2 cells) and ex vivo intestine models (rat precision cut intestine slices and the pig in-situ small intestinal segment perfusion (SISP) technique) to study the effect of food compounds. In vitro digested yellow (YOd) and white onion extracts (WOd) were used as model food compounds and transcriptomics was applied to obtain more insight into which extent mode of actions depend on the model. The three intestine models shared 9,140 genes which were used to compare the responses to digested onions between the models. Unsupervised clustering analysis showed that genes up- or down-regulated by WOd in human Caco-2 cells and rat intestine slices were similarly regulated by YOd, indicating comparable modes of action for the two onion species. Highly variable responses to onion were found in the pig SISP model. By focussing only on genes with significant differential expression, in combination with a fold change > 1.5, 15 genes showed similar onion-induced expression in human Caco-2 cells and rat intestine slices and 2 overlapping genes were found between the human Caco-2 and pig SISP model. Pathway analyses revealed that mainly processes related to oxidative stress, and especially the Keap1-Nrf2 pathway, were affected by onions in all three models. Our data fit with previous in vivo studies showing that the beneficial effects of onions are mostly linked to their antioxidant properties. Taken together, our data indicate that each of the in vitro and ex vivo intestine models used in this study, taking into account their limitations, can be used to determine modes of action of nutritional compounds and can thereby reduce the number of animals used in conventional nutritional intervention studies.

  14. Direct experimental manipulation of intestinal cells in Ascaris suum, with minor influences on the global transcriptome.

    Science.gov (United States)

    Rosa, Bruce A; McNulty, Samantha N; Mitreva, Makedonka; Jasmer, Douglas P

    2017-04-01

    Ascaris suum provides a powerful model for studying parasitic nematodes, including individual tissues such as the intestine, an established target for anthelmintic treatments. Here, we add a valuable experimental component to our existing functional, proteomic, transcriptomic and phylogenomic studies of the Ascaris suum intestine, by developing a method to manipulate intestinal cell functions via direct delivery of experimental treatments (in this case, double-stranded (ds)RNA) to the apical intestinal membrane. We developed an intestinal perfusion method for direct, controlled delivery of dsRNA/heterogeneous small interfering (hsi) RNA into the intestinal lumen for experimentation. RNA-Seq (22 samples) was used to assess influences of the method on global intestinal gene expression. Successful mRNA-specific knockdown in intestinal cells of adult A. suum was accomplished with this new experimental method. Global transcriptional profiling confirmed that targeted transcripts were knocked down more significantly than any others, with only 12 (0.07% of all genes) or 238 (1.3%) off-target gene transcripts consistently differentially regulated by dsRNA treatment or the perfusion experimental design, respectively (after 24h). The system supports controlled, effective delivery of treatments (dsRNA/hsiRNA) to the apical intestinal membrane with relatively minor off-target effects, and builds on our experimental model to dissect A. suum intestinal cell functions with broad relevance to parasitic nematodes. Copyright © 2017 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.

  15. Resonant Mie scattering (RMieS) correction applied to FTIR images of biological tissue samples.

    Science.gov (United States)

    Bambery, Keith R; Wood, Bayden R; McNaughton, Don

    2012-01-07

    Recently a resonant Mie scattering (RMieS) correction approach has been developed and demonstrated to be effective for removing the baseline distortions that compromise the raw data in individual spectra. In this paper RMieS correction is extended to FTIR images of a tissue section from biopsy of the human cervical transformation zone and a coronal tissue section of a Wistar rat brain and compared to the uncorrected images. It is shown that applying RMieS correction to FTIR images a) removes baseline distortions from the image spectra and thus reveals previously hidden information on spatial variation of chemical contents within the tissue and b) can lead to improved automatic tissue feature classification through multivariate cluster analysis. This journal is © The Royal Society of Chemistry 2012

  16. Development and application of specific cytokine assays in tissue samples from a bottlenose dolphin with hyperinsulinemia

    Science.gov (United States)

    Chronic inflammation has been associated with insulin resistance and type 2 diabetes in humans. Postmortem hepatic and splenic tissue from a 46-year old geriatric male bottlenose dolphin (Tursiops truncatus) with insulin resistance (chronic hyperinsulinemia with hyperglycemia) , chronic = inflamma...

  17. 76 FR 24862 - Proposed Information Collection; Comment Request; Protocol for Access to Tissue Specimen Samples...

    Science.gov (United States)

    2011-05-03

    ... DEPARTMENT OF COMMERCE National Oceanic and Atmospheric Administration Proposed Information... Tissue Bank AGENCY: National Oceanic and Atmospheric Administration (NOAA), Commerce. ACTION: Notice... analyses to determine environmental trends of contaminants and other substances of interest. The NMMTB...

  18. Biomedical analysis of formalin-fixed, paraffin-embedded tissue samples: The Holy Grail for molecular diagnostics.

    Science.gov (United States)

    Donczo, Boglarka; Guttman, Andras

    2018-04-02

    More than a century ago in 1893, a revolutionary idea about fixing biological tissue specimens was introduced by Ferdinand Blum, a German physician. Since then, a plethora of fixation methods have been investigated and used. Formalin fixation with paraffin embedment became the most widely used types of fixation and preservation method, due to its proper architectural conservation of tissue structures and cellular shape. The huge collection of formalin-fixed, paraffin-embedded (FFPE) sample archives worldwide holds a large amount of unearthed information about diseases that could be the Holy Grail in contemporary biomarker research utilizing analytical omics based molecular diagnostics. The aim of this review is to critically evaluate the omics options for FFPE tissue sample analysis in the molecular diagnostics field. Copyright © 2018. Published by Elsevier B.V.

  19. Sugar and acid interconversion in tomato fruits based on biopsy sampling of locule gel and pericarp tissue

    NARCIS (Netherlands)

    Schouten, R.E.; Woltering, E.J.; Tijskens, L.M.M.

    2016-01-01

    This study deals with quantifying sugar and acids levels important for the perceived taste of tomatoes (Solanum lycopersicum). Sugar and acids levels were measured repeatedly on the same tomato using tissue samples obtained with a biopsy needle in combination with HPLC protocols. Biopsies of

  20. Development and application of specific cytokine assays in tissue samples from a bottlenose dolphin with hyperinsulinemia

    Directory of Open Access Journals (Sweden)

    Kirsten eEberle

    2013-10-01

    Full Text Available Chronic inflammation has been associated with insulin resistance and type 2 diabetes (T2D in humans. Postmortem hepatic and splenic tissue from a 46-year-old geriatric male bottlenose dolphin (Tursiops truncatus with insulin resistance (chronic hyperinsulinemia with hyperglycemia, chronic inflammation (white blood cell count greater than 12,000 cells/μL, and mild fatty liver disease was evaluated for elevated pro-inflammatory mediators. Cytokine mRNA expression in postmortem hepatic and splenic tissue, as determined by real-time PCR, included an array of cytokines: TGF-β, TNF-α, IFN-γ, IL-2, IL-4, IL-10, IL-12p40, IL-13, and IL-18. Values from this dolphin were compared to a younger reference dolphin with no known chronic metabolic perturbations or inflammation. Levels of TGF-β, TNF-α, and IL-4 were higher in the case dolphin's liver compared to that of the reference dolphin. In the case dolphin's spleen, IL-10 and IFN-γ mRNA was upregulated while IL-4 was less than reference dolphin. IL-18 and IL-13 were upregulated in both tissues. Fluorescent immunohistochemistry (IHC utilized the following antibodies: anti-porcine IL-6, anti-bovine IFN-γ, IL-4, and IL-10, anti-human TGF-β, anti-ovine IL-1β, and anti-dolphin IL-8. Fluorescent IHC in spleen from the case dolphin revealed staining of IL-4, IL-6, IL-8, and TGF-β throughout the tissue. IL-10 and IFN-γ were seen to predominate in areas surrounding the follicles of splenic tissue. This is the first characterization of cytokine levels in dolphin hepatic and splenic tissue. While there are limitations to a case study, this report of inflammatory biomarkers in tissues of a dolphin with insulin resistance and fatty liver disease are similar to those observed in human patients.

  1. Measurement of characteristic prompt gamma rays emitted from oxygen and carbon in tissue-equivalent samples during proton beam irradiation.

    Science.gov (United States)

    Polf, Jerimy C; Panthi, Rajesh; Mackin, Dennis S; McCleskey, Matt; Saastamoinen, Antti; Roeder, Brian T; Beddar, Sam

    2013-09-07

    The purpose of this work was to characterize how prompt gamma (PG) emission from tissue changes as a function of carbon and oxygen concentration, and to assess the feasibility of determining elemental concentration in tissues irradiated with proton beams. For this study, four tissue-equivalent water-sucrose samples with differing densities and concentrations of carbon, hydrogen, and oxygen were irradiated with a 48 MeV proton pencil beam. The PG spectrum emitted from each sample was measured using a high-purity germanium detector, and the absolute detection efficiency of the detector, average beam current, and delivered dose distribution were also measured. Changes to the total PG emission from (12)C (4.44 MeV) and (16)O (6.13 MeV) per incident proton and per Gray of absorbed dose were characterized as a function of carbon and oxygen concentration in the sample. The intensity of the 4.44 MeV PG emission per incident proton was found to be nearly constant for all samples regardless of their carbon concentration. However, we found that the 6.13 MeV PG emission increased linearly with the total amount (in grams) of oxygen irradiated in the sample. From the measured PG data, we determined that 1.64 × 10(7) oxygen PGs were emitted per gram of oxygen irradiated per Gray of absorbed dose delivered with a 48 MeV proton beam. These results indicate that the 6.13 MeV PG emission from (16)O is proportional to the concentration of oxygen in tissue irradiated with proton beams, showing that it is possible to determine the concentration of oxygen within tissues irradiated with proton beams by measuring (16)O PG emission.

  2. Measurement of characteristic prompt gamma rays emitted from oxygen and carbon in tissue-equivalent samples during proton beam irradiation

    International Nuclear Information System (INIS)

    Polf, Jerimy C; Panthi, Rajesh; Mackin, Dennis S; Beddar, Sam; McCleskey, Matt; Saastamoinen, Antti; Roeder, Brian T

    2013-01-01

    The purpose of this work was to characterize how prompt gamma (PG) emission from tissue changes as a function of carbon and oxygen concentration, and to assess the feasibility of determining elemental concentration in tissues irradiated with proton beams. For this study, four tissue-equivalent water–sucrose samples with differing densities and concentrations of carbon, hydrogen, and oxygen were irradiated with a 48 MeV proton pencil beam. The PG spectrum emitted from each sample was measured using a high-purity germanium detector, and the absolute detection efficiency of the detector, average beam current, and delivered dose distribution were also measured. Changes to the total PG emission from 12 C (4.44 MeV) and 16 O (6.13 MeV) per incident proton and per Gray of absorbed dose were characterized as a function of carbon and oxygen concentration in the sample. The intensity of the 4.44 MeV PG emission per incident proton was found to be nearly constant for all samples regardless of their carbon concentration. However, we found that the 6.13 MeV PG emission increased linearly with the total amount (in grams) of oxygen irradiated in the sample. From the measured PG data, we determined that 1.64 × 10 7 oxygen PGs were emitted per gram of oxygen irradiated per Gray of absorbed dose delivered with a 48 MeV proton beam. These results indicate that the 6.13 MeV PG emission from 16 O is proportional to the concentration of oxygen in tissue irradiated with proton beams, showing that it is possible to determine the concentration of oxygen within tissues irradiated with proton beams by measuring 16 O PG emission. (paper)

  3. Real time monitoring of pulsatile change in hemoglobin concentrations of cerebral tissue by a portable tissue oximeter with a 10-Hz sampling rate

    Science.gov (United States)

    Shiga, Toshikazu; Chihara, Eiichi; Tanabe, Kazuhisa; Tanaka, Yoshifumi; Yamamoto, Katsuyuki

    1998-01-01

    A portable CW tissue oximeter of a 10-Hz sampling rate was developed for examination of pulsatile components of the output signals as a mean of checking the signal reliability during long-term monitoring. Feasible studies were performed on a healthy subject. Changes in Hb and HbO2 signals of cerebral tissue were continuously measured by placing a photoprobe on the forehead during 6-hour sleep. Pulsatile changes in Hb and HbO2 were steadily observed over a whole period of the recording. The phase relation of pulsation in Hb and HbO2 was almost inverse. Not only information for reliable monitoring but also physiological parameters with respect to cerebral circulation and metabolism could be obtained by measuring the pulsatile components.

  4. HER-2 and INT-2 amplification estimated by quantitative PCR in paraffin-embedded ovarian cancer tissue samples.

    Science.gov (United States)

    Hruza, C; Dobianer, K; Beck, A; Czerwenka, K; Hanak, H; Klein, M; Leodolter, S; Medl, M; Müllauer-Ertl, S; Preiser, J

    1993-01-01

    Competitive polymerase chain reaction (PCR) systems were developed for rapid and quantitative estimation of HER-2 (c-erbB-2) and INT-2 oncogene amplification in paraffin-embedded ovarian cancer tissue samples. The beta-globin gene was used as reference and DNA from paraffin-embedded placenta tissue as single copy control. Reliability of the PCR method could be demonstrated by comparing dot blot data with PCR data of identical tumour samples. The PCR method was used to determine HER-2 and INT-2 copy numbers in 196 ovarian cancer samples. HER-2 and INT-2 were found to be amplified in 40 and 19%, respectively. In 8% HER-2 copy numbers were greater than five, but no high INT-2 copies were noted. Kaplan-Meier estimates did not reveal significant association with overall survival. Indirect correlation between HER-2 and INT-2 amplification was observed. The present PCR system is a valuable method for prospective and retrospective studies.

  5. Rapid detection of transmissible gastroenteritis virus in swine small intestine samples using real-time reverse transcription recombinase polymerase amplification.

    Science.gov (United States)

    Wang, Jinfeng; Wang, Jianchang; Zhang, Ruoxi; Liu, Libing; Shi, Ruihan; Han, Qingan; Yuan, Wanzhe

    2018-03-14

    A rapid and specific real-time reverse-transcription recombinase polymerase amplification assay (RT-RPA) was developed to detect the transmissible gastroenteritis virus (TGEV) in this study. The primers and exo probe were designed to be specific for a portion of spike (S) gene conserved in TGEV, but absent in the closely related porcine respiratory coronavirus (PRCV). The amplification was performed at 40 °C for 20 min. The assay could only detect the TGEV, and there was no cross-reaction with other pathogens tested. Using the in vitro transcribed TGEV RNA as template, the limit of detection of the developed RT-RPA was 100 copies per reaction. The assay performance was evaluated by testing 76 clinical samples by RT-RPA and a real-time RT-PCR. Fourteen samples were TGEV RNA positive in RT-RPA (18.4%, 14/76), which were also positive in the real-time RT-PCR. The diagnostic agreement between the two assays was 100% (76/76). The R 2 value of RT-RPA and real-time RT-PCR was 0.959 by linear regression analysis. The developed RT-RPA assay provides a useful alternative tool for rapid, simple and reliable detection of TGEV in resource-limited diagnostic laboratories and on-site facilities. Copyright © 2018 Elsevier B.V. All rights reserved.

  6. Concentration of organochlorines in human brain, liver, and adipose tissue autopsy samples from Greenland

    DEFF Research Database (Denmark)

    Dewailly, Éric; Mulvad, Gert; Pedersen, Henning S.

    1999-01-01

    Organochlorines are persistent lipophilic compounds that accumulate in Inuit people living in circumpolar countries. Organochlorines accumulate as a result of the Inuits' large consumption of sea mammal fat; however, available data are limited to blood lipids, milk fat, and adipose tissue. We rep...

  7. Bisulfite-Based DNA Methylation Analysis from Recent and Archived Formalin-Fixed, Paraffin Embedded Colorectal Tissue Samples.

    Science.gov (United States)

    Kalmár, Alexandra; Péterfia, Bálint; Hollósi, Péter; Wichmann, Barnabás; Bodor, András; Patai, Árpád V; Schöller, Andrea; Krenács, Tibor; Tulassay, Zsolt; Molnár, Béla

    2015-09-01

    We aimed to test the applicability of formalin-fixed and paraffin-embedded (FFPE) tissue samples for gene specific DNA methylation analysis after using two commercially available DNA isolation kits. Genomic DNA was isolated from 5 colorectal adenocarcinomas and 5 normal adjacent tissues from "recent", collected within 6 months, and "archived", collected more than 5 years ago, FFPE tissues using either High Pure FFPET DNA Isolation kit or QIAamp DNA FFPE Tissue kit. DNA methylation analysis of MAL, SFRP1 and SFRP2 genes, known to be hypermethylated in CRC, was performed using methylation-sensitive high resolution melting (MS-HRM) analysis and sequencing. QIAamp (Q) method resulted in slightly higher recovery in archived (HP: 1.22 ± 3.18 μg DNA; Q: 3.00 ± 4.04 μg DNA) and significantly (p < 0.05) higher recovery in recent samples compared to High Pure method (HP) (HP: 4.10 ± 2.91 μg DNA; Q: 11.51 ± 7.50 μg DNA). Both OD260/280 and OD260/230 ratios were lower, but still high in the High Pure isolated archived and recent samples compared to those isolated with QIAamp. Identical DNA methylation patterns were detected for all 3 genes tested by MS-HRM with both isolation kits in the recent group. However, despite of higher DNA recovery in QIAamp slightly more reproducible methylation results were obtained from High Pure isolated archived samples. Sequencing confirmed DNA hypermethylation in CRCs. In conclusion, reproducible DNA methylation patterns were obtained from recent samples using both isolation kits. However, long term storage may affect the reliability of the results leading to moderate differences between the efficiency of isolation kits.

  8. Comparison of Accuracy of Whole-Exome Sequencing with Formalin-Fixed Paraffin-Embedded and Fresh Frozen Tissue Samples.

    Directory of Open Access Journals (Sweden)

    Ensel Oh

    Full Text Available Formalin fixing with paraffin embedding (FFPE has been a standard sample preparation method for decades, and archival FFPE samples are still very useful resources. Nonetheless, the use of FFPE samples in cancer genome analysis using next-generation sequencing, which is a powerful technique for the identification of genomic alterations at the nucleotide level, has been challenging due to poor DNA quality and artificial sequence alterations. In this study, we performed whole-exome sequencing of matched frozen samples and FFPE samples of tissues from 4 cancer patients and compared the next-generation sequencing data obtained from these samples. The major differences between data obtained from the 2 types of sample were the shorter insert size and artificial base alterations in the FFPE samples. A high proportion of short inserts in the FFPE samples resulted in overlapping paired reads, which could lead to overestimation of certain variants; >20% of the inserts in the FFPE samples were double sequenced. A large number of soft clipped reads was found in the sequencing data of the FFPE samples, and about 30% of total bases were soft clipped. The artificial base alterations, C>T and G>A, were observed in FFPE samples only, and the alteration rate ranged from 200 to 1,200 per 1M bases when sequencing errors were removed. Although high-confidence mutation calls in the FFPE samples were compatible to that in the frozen samples, caution should be exercised in terms of the artifacts, especially for low-confidence calls. Despite the clearly observed artifacts, archival FFPE samples can be a good resource for discovery or validation of biomarkers in cancer research based on whole-exome sequencing.

  9. Endoscopic examination and tissue sampling of the bovine teat and udder cistern

    OpenAIRE

    Vangroenweghe, Fr; Van Den Broeck, Wim; De Ketelaere, Adelheid; van Bree, Henri; Duchateau, Luc; Burvenich, Christian

    2006-01-01

    The aim of this study was to evaluate the application of an endoscopic technique to investigate the teat and udder cisterns of the bovine mammary gland, and to biopsy tissues within the cisterns. An anesthetic protocol for application in standing animals was designed, using a combination of general and local anesthesia. Individual quarter milk production (QMP), quarter somatic cell count (SCC), and occurrence of new intramammary infection were assessed after application of the technique, and ...

  10. Micro-Raman spectroscopy of tissue samples for oral pathology follow-up monitoring

    Science.gov (United States)

    Delfino, I.; Camerlingo, C.; Zenone, F.; Perna, G.; Capozzi, V.; Cirillo, N.; Gaeta, G. M.; Lepore, M.

    2010-04-01

    An "in vitro" study of Raman spectra from oral human tissues is reported in order to the develop a diagnostic method suitable for "in vivo" oral pathology follow-up. The investigated pathology is Pemphigus Vulgaris (PV) for which new techniques for guiding and monitoring therapy would be particularly useful. Raman spectra were obtained in the wavenumber regions from 1000 to 1800 cm-1 and 2700 to 3200 cm-1 from tissues from patients at different stages of pathology (active PV, under therapy and in PV remission stage) as confirmed by histopathological and immunofluorescence analysis. Differences in the spectra depending on tissue illness stage arise in 1150-1250 cm-1 (amide III) and 1420-1450 cm-1 (CH3 deformation) regions and around 1650 cm-1 (amide I) and 2930 cm-1 (CH3 symmetric stretch). A wavelet deconvolution procedure was applied to the spectra for better discriminating among the three different stages of illness and a linear regression analysis was used to fully exploit the content of information of Raman spectra.

  11. Epo receptors are not detectable in primary human tumor tissue samples.

    Directory of Open Access Journals (Sweden)

    Steve Elliott

    Full Text Available Erythropoietin (Epo is a cytokine that binds and activates an Epo receptor (EpoR expressed on the surface of erythroid progenitor cells to promote erythropoiesis. While early studies suggested EpoR transcripts were expressed exclusively in the erythroid compartment, low-level EpoR transcripts were detected in nonhematopoietic tissues and tumor cell lines using sensitive RT-PCR methods. However due to the widespread use of nonspecific anti-EpoR antibodies there are conflicting data on EpoR protein expression. In tumor cell lines and normal human tissues examined with a specific and sensitive monoclonal antibody to human EpoR (A82, little/no EpoR protein was detected and it was not functional. In contrast, EpoR protein was reportedly detectable in a breast tumor cell line (MCF-7 and breast cancer tissues with an anti-EpoR polyclonal antibody (M-20, and functional responses to rHuEpo were reported with MCF-7 cells. In another study, a functional response was reported with the lung tumor cell line (NCI-H838 at physiological levels of rHuEpo. However, the specificity of M-20 is in question and the absence of appropriate negative controls raise questions about possible false-positive effects. Here we show that with A82, no EpoR protein was detectable in normal human and matching cancer tissues from breast, lung, colon, ovary and skin with little/no EpoR in MCF-7 and most other breast and lung tumor cell lines. We show further that M-20 provides false positive staining with tissues and it binds to a non-EpoR protein that migrates at the same size as EpoR with MCF-7 lysates. EpoR protein was detectable with NCI-H838 cells, but no rHuEpo-induced phosphorylation of AKT, STAT3, pS6RP or STAT5 was observed suggesting the EpoR was not functional. Taken together these results raise questions about the hypothesis that most tumors express high levels of functional EpoR protein.

  12. Intestine transplantation

    Directory of Open Access Journals (Sweden)

    Tadeja Pintar

    2011-02-01

    Conclusion: Intestine transplantation is reserved for patients with irreversible intestinal failure due to short gut syndrome requiring total paranteral nutrition with no possibility of discontinuation and loss of venous access for patient maintenance. In these patients complications of underlying disease and long-term total parenteral nutrition are present.

  13. A simple microfluidic platform for the partial treatment of insuspendable tissue samples with orientation control.

    Science.gov (United States)

    Fan, Anthony; Tofangchi, Alireza; De Venecia, Matthew; Saif, Taher

    2018-02-27

    Microfluidic devices have extensively been applied to study biological samples, including single cells. Exploiting laminar flows on a small scale, microfluidics allow for the selective and partial exposure of samples to various chemical treatments. Traditionally, suspendable samples are first flowed into formed microchannels and are allowed to adhere to the channel floor randomly with no control over sample placement or orientation, before being subjected to partial treatment. This severely limits the choice of samples and the extent of sample preparations. Here, we overcame this limit by reversing the sequence. We prepared the samples first on glass substrates. A patterned silicone slab was then placed on the substrate to form channels at an appropriate orientation with respect to the sample. We used liquid silicone rubber (LSR) as the base material. Its compliance (low elastic modulus) and its adhesion to glass offer the necessary seal to form the microchannels naturally. The applicability of the device was demonstrated by testing single axons of embryonic Drosophila motor neurons in vivo. A segment of the axons was subjected to drugs that inhibit myosin activities or block voltage-gated sodium ion channels. In response, the axons reduced the clustering of neuro-transmitter vesicles at the presynaptic terminal of neuromuscular junctions, or increased the calcium intake and underwent membrane hyperpolarization, respectively. Such fundamental studies cannot be carried out using conventional microfluidics.

  14. Clinical use of fungal PCR from deep tissue samples in the diagnosis of invasive fungal diseases: a retrospective observational study.

    Science.gov (United States)

    Ala-Houhala, M; Koukila-Kähkölä, P; Antikainen, J; Valve, J; Kirveskari, J; Anttila, V-J

    2018-03-01

    To assess the clinical use of panfungal PCR for diagnosis of invasive fungal diseases (IFDs). We focused on the deep tissue samples. We first described the design of panfungal PCR, which is in clinical use at Helsinki University Hospital. Next we retrospectively evaluated the results of 307 fungal PCR tests performed from 2013 to 2015. Samples were taken from normally sterile tissues and fluids. The patient population was nonselected. We classified the likelihood of IFD according to the criteria of the European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG), comparing the fungal PCR results to the likelihood of IFD along with culture and microscopy results. There were 48 positive (16%) and 259 negative (84%) PCR results. The sensitivity and specificity of PCR for diagnosing IFDs were 60.5% and 91.7%, respectively, while the negative predictive value and positive predictive value were 93.4% and 54.2%, respectively. The concordance between the PCR and the culture results was 86% and 87% between PCR and microscopy, respectively. Of the 48 patients with positive PCR results, 23 had a proven or probable IFD. Fungal PCR can be useful for diagnosing IFDs in deep tissue samples. It is beneficial to combine fungal PCR with culture and microscopy. Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  15. Simultaneous single-sample determination of NMNAT isozyme activities in mouse tissues.

    Directory of Open Access Journals (Sweden)

    Giuseppe Orsomando

    Full Text Available A novel assay procedure has been developed to allow simultaneous activity discrimination in crude tissue extracts of the three known mammalian nicotinamide mononucleotide adenylyltransferase (NMNAT, EC 2.7.7.1 isozymes. These enzymes catalyse the same key reaction for NAD biosynthesis in different cellular compartments. The present method has been optimized for NMNAT isozymes derived from Mus musculus, a species often used as a model for NAD-biosynthesis-related physiology and disorders, such as peripheral neuropathies. Suitable assay conditions were initially assessed by exploiting the metal-ion dependence of each isozyme recombinantly expressed in bacteria, and further tested after mixing them in vitro. The variable contributions of the three individual isozymes to total NAD synthesis in the complex mixture was calculated by measuring reaction rates under three selected assay conditions, generating three linear simultaneous equations that can be solved by a substitution matrix calculation. Final assay validation was achieved in a tissue extract by comparing the activity and expression levels of individual isozymes, considering their distinctive catalytic efficiencies. Furthermore, considering the key role played by NMNAT activity in preserving axon integrity and physiological function, this assay procedure was applied to both liver and brain extracts from wild-type and Wallerian degeneration slow (Wld(S mouse. Wld(S is a spontaneous mutation causing overexpression of NMNAT1 as a fusion protein, which protects injured axons through a gain-of-function. The results validate our method as a reliable determination of the contributions of the three isozymes to cellular NAD synthesis in different organelles and tissues, and in mutant animals such as Wld(S.

  16. Proteoglycan and proteome profiling of central human pulmonary fibrotic tissue utilizing miniaturized sample preparation

    DEFF Research Database (Denmark)

    Malmström, Johan; Larsen, Kristoffer; Hansson, Lennart

    2002-01-01

    -dimensional electrophoresis was interfaced to miniaturized sample preparation techniques using microcapillary extraction. Four protein groups were identified; cytoskeletal, adhesion, scavenger and metabolic proteins. These patient's proteomes showed a high degree of heterogeneity between patients but larger homogeneity...

  17. Incidence of mycobacterial infections in cats in Great Britain: estimate from feline tissue samples submitted to diagnostic laboratories.

    Science.gov (United States)

    Gunn-Moore, D A; Gaunt, C; Shaw, D J

    2013-08-01

    The aim of this study was to estimate the incidence of mycobacterial infections in cats in Great Britain (GB). This was performed using the proxy measure of feline tissue samples submitted to diagnostic laboratories in GB that were found to have histopathological changes typical of mycobacterial infection ('MYC'). Sixteen primary diagnostic laboratories were asked for information on the number of feline samples submitted in 2009, the number with MYC, the number undergoing Ziehl-Neelsen (ZN) staining and, for comparison, the number diagnosed with lymphoma. Eight laboratories provided full data for the whole year: 11,782 samples; lymphoma 3.2% (mean, 95% CI: 2.89, 3.5), MYC 1.16% (0.98; 1.37) and ZN-positive 0.31% (0.22; 0.43). Data on 1569 samples from seven laboratories that provided partial data on samples for the whole year revealed similar results, although all changes were more frequent: lymphoma 5.42% (4.35; 6.66), MYC 2.36% (1.66; 3.23) and ZN-positive 0.77% (0.40; 1.33). One laboratory only provided data for part of the year (4.5 months), reporting all three types of histopathology less frequently: 18,232 samples; lymphoma 0.2% (0.18; 0.32), MYC 0.07% (0.04; 0.12) and ZN-positive 0.05% (0.02; 0.09). The reasons for low reporting rates in this high-throughput laboratory are unclear. In total, 187 samples were reported as having MYC. Five Reference laboratories were also contacted, reporting 174 feline tissue submissions in 2009, with mycobacteria being cultured from 90. The study shows that MYC are frequently reported in tissue samples from cats in GB, being reported in ~1% of samples, with confirmation as ZN-positive in ~0.3%. Lymphoma is recognized as a common disease in cats, being seen in ~3% of samples in this study. When compared against MYC, lymphoma was reported only twice as frequently. This confirms that far from being rare, clinically significant mycobacterial infections occur commonly in cats in GB. © 2012 Blackwell Verlag GmbH.

  18. Human biological sample biobanking to support tissue biomarkers in pharmaceutical research and development.

    Science.gov (United States)

    Womack, Christopher; Mager, S Rachel

    2014-11-01

    Advances in the understanding of molecular pathology and thereby the mechanisms that could be amenable to therapeutic manipulation are the reason that pharmaceutical research and development is focused increasingly on measurement of molecular biomarkers in human biological samples. Obtaining direct or indirect access to sufficient samples that are fit for research purposes can be a major challenge. A biobanking infrastructure has a significant role in the acquisition, storage and usage of human biological samples and here we review some key requirements for establishing a biobank. These include ensuring; that appropriate governance mechanisms are in place, that samples available are appropriate and fit for the intended research purposes that the infrastructure is sustainable in the future and that use of the biobank assets meets the strategic aims of the host organisation. Finally we present a case study--the STRATUM project which has recently completed and through a collaborative approach involving six industry and public partners drawing on a network of experts, examined biobank policies, public attitudes to biobanking, donor consent, sample and data standards, technical requirements for a register and biobanking financial models, albeit from a UK perspective. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. A comparative examination of sample treatment procedures for ICAP-AES analysis of biological tissue

    Science.gov (United States)

    De Boer, J. L. M.; Maessen, F. J. M. J.

    The objective of this study was to contribute to the evaluation of existing sample preparation procedures for ICAP-AES analysis of biological material. Performance characteristics were established of current digestion procedures comprising extraction, solubilization, pressure digestion, and wet and dry ashing methods. Apart from accuracy and precision, a number of criteria of special interest for the analytical practice was applied. As a test sample served SRM bovine liver. In this material six elements were simultaneously determined. Results showed that every procedure has its defects and advantages. Hence, unambiguous recommendation of standard digestion procedures can be made only when taking into account the specific analytical problem.

  20. The presence of enterovirus, adenovirus, and parvovirus B19 in myocardial tissue samples from autopsies

    DEFF Research Database (Denmark)

    Nielsen, Trine Skov; Hansen, Jakob; Nielsen, Lars Peter

    2014-01-01

    of adenovirus, enterovirus, and parvovirus B19 (PVB) in myocardial autopsy samples from myocarditis related deaths and in non-inflamed control hearts in an effort to clarify their significance as the causes of myocarditis in a forensic material. METHODS: We collected all autopsy cases diagnosed with myocarditis...... of myocarditis. The detection of PVB in myocardial autopsy samples most likely represents a persistent infection with no or limited association with myocardial inflammation. The forensic investigation of myocardial inflammation demands a thorough examination, including special attention to non-viral causes...

  1. Whole-genome gene expression profiling of formalin-fixed, paraffin-embedded tissue samples.

    Directory of Open Access Journals (Sweden)

    Craig April

    2009-12-01

    Full Text Available We have developed a gene expression assay (Whole-Genome DASL, capable of generating whole-genome gene expression profiles from degraded samples such as formalin-fixed, paraffin-embedded (FFPE specimens.We demonstrated a similar level of sensitivity in gene detection between matched fresh-frozen (FF and FFPE samples, with the number and overlap of probes detected in the FFPE samples being approximately 88% and 95% of that in the corresponding FF samples, respectively; 74% of the differentially expressed probes overlapped between the FF and FFPE pairs. The WG-DASL assay is also able to detect 1.3-1.5 and 1.5-2 -fold changes in intact and FFPE samples, respectively. The dynamic range for the assay is approximately 3 logs. Comparing the WG-DASL assay with an in vitro transcription-based labeling method yielded fold-change correlations of R(2 approximately 0.83, while fold-change comparisons with quantitative RT-PCR assays yielded R(2 approximately 0.86 and R(2 approximately 0.55 for intact and FFPE samples, respectively. Additionally, the WG-DASL assay yielded high self-correlations (R(2>0.98 with low intact RNA inputs ranging from 1 ng to 100 ng; reproducible expression profiles were also obtained with 250 pg total RNA (R(2 approximately 0.92, with approximately 71% of the probes detected in 100 ng total RNA also detected at the 250 pg level. When FFPE samples were assayed, 1 ng total RNA yielded self-correlations of R(2 approximately 0.80, while still maintaining a correlation of R(2 approximately 0.75 with standard FFPE inputs (200 ng.Taken together, these results show that WG-DASL assay provides a reliable platform for genome-wide expression profiling in archived materials. It also possesses utility within clinical settings where only limited quantities of samples may be available (e.g. microdissected material or when minimally invasive procedures are performed (e.g. biopsied specimens.

  2. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins: Focus on sample preparation and derivatization conditions.

    Science.gov (United States)

    Weber, Daniela; Davies, Michael J; Grune, Tilman

    2015-08-01

    Protein oxidation is involved in regulatory physiological events as well as in damage to tissues and is thought to play a key role in the pathophysiology of diseases and in the aging process. Protein-bound carbonyls represent a marker of global protein oxidation, as they are generated by multiple different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples. © 2015 Published by Elsevier Ltd.

  3. Evidence of presence of Mycobacterium tuberculosis in bovine tissue samples by multiplex PCR: possible relevance to reverse zoonosis.

    Science.gov (United States)

    Mittal, M; Chakravarti, S; Sharma, V; Sanjeeth, B S; Churamani, C P; Kanwar, N S

    2014-04-01

    Bovine tuberculosis, caused by Mycobacterium bovis, remains one of the most important zoonotic health concerns worldwide. The transmission of Mycobacterium tuberculosis from humans to animals also occurs especially in countries where there is close interaction of humans with the animals. In the present study, thirty bovine lung tissue autopsy samples from an organized dairy farm located in North India were screened for the presence of Mycobacterium tuberculosis complex by smear microscopy, histopathological findings and PCR. Differential diagnosis of M. tuberculosis and M. bovis was made based on the deletion of mce-3 operon in M. bovis. The present study found eight of these samples positive for M. tuberculosis by multiplex PCR. Sequencing was performed on two PCR-positive representative samples and on annotation, and BLAST analysis confirmed the presence of gene fragment specific to Mycobacterium tuberculosis. The presence of M. tuberculosis in all the positive samples raises the possibility of human-to-cattle transmission and possible adaptation of this organism in bovine tissues. This study accentuates the importance of screening and differential diagnosis of Mycobacterium tuberculosis complex in humans and livestock for adopting effective TB control and eradication programmes. © 2014 Blackwell Verlag GmbH.

  4. Clinical procedure for colon carcinoma tissue sampling directly affects the cancer marker-capacity of VEGF family members

    International Nuclear Information System (INIS)

    Pringels, Sarah; Van Damme, Nancy; De Craene, Bram; Pattyn, Piet; Ceelen, Wim; Peeters, Marc; Grooten, Johan

    2012-01-01

    mRNA levels of members of the Vascular Endothelial Growth Factor family (VEGF-A, -B, -C, -D, Placental Growth Factor/PlGF) have been investigated as tissue-based markers of colon cancer. These studies, which used specimens obtained by surgical resection or colonoscopic biopsy, yielded contradictory results. We studied the effect of the sampling method on the marker accuracy of VEGF family members. Comparative RT-qPCR analysis was performed on healthy colon and colon carcinoma samples obtained by biopsy (n = 38) or resection (n = 39) to measure mRNA expression levels of individual VEGF family members. mRNA levels of genes encoding the eicosanoid enzymes cyclooxygenase 2 (COX2) and 5-lipoxygenase (5-LOX) and of genes encoding the hypoxia markers glucose transporter 1 (GLUT-1) and carbonic anhydrase IX (CAIX) were included as markers for cellular stress and hypoxia. Expression levels of COX2, 5-LOX, GLUT-1 and CAIX revealed the occurrence in healthy colon resection samples of hypoxic cellular stress and a concurrent increment of basal expression levels of VEGF family members. This increment abolished differential expression of VEGF-B and VEGF-C in matched carcinoma resection samples and created a surgery-induced underexpression of VEGF-D. VEGF-A and PlGF showed strong overexpression in carcinoma samples regardless of the sampling method. Sampling-induced hypoxia in resection samples but not in biopsy samples affects the marker-reliability of VEGF family members. Therefore, biopsy samples provide a more accurate report on VEGF family mRNA levels. Furthermore, this limited expression analysis proposes VEGF-A and PlGF as reliable, sampling procedure insensitive mRNA-markers for molecular diagnosis of colon cancer

  5. Attitudes towards biomedical use of tissue sample collections, consent, and biobanks among Finns

    DEFF Research Database (Denmark)

    Tupasela, Aaro Mikael; Sihvo, Sinikka; Snell, Karolna

    2010-01-01

    To ascertain the attitudes towards the use of existing diagnostic and research samples, the setting up of a national biobank, and different types of informed consent among Finns. Method: A population survey of 2,400 randomly selected Finns aged 24-65 was conducted at the beginning of 2007....

  6. Tensile rupture of medial arterial tissue studied by X-ray micro-tomography on stained samples.

    Science.gov (United States)

    Helfenstein-Didier, Clémentine; Taïnoff, Damien; Viville, Julien; Adrien, Jérôme; Maire, Éric; Badel, Pierre

    2018-02-01

    Detailed characterization of damage and rupture mechanics of arteries is one the current challenges in vascular biomechanics, which requires developing suitable experimental approaches. This paper introduces an approach using in situ tensile tests in an X-ray micro-tomography setup to observe mechanisms of damage initiation and progression in medial layers of porcine aortic samples. The technique requires the use of sodium polytungstate as a contrast agent, of which the conditions for use are detailed in this paper. Immersion of the samples during 24h in a 15g/L concentrated solution provided the best compromise for viewing musculo-elastic units in this tissue. The process of damage initiation, delamination and rupture of medial tissue under tensile loading was observed and can be described as an elementary process repeating several times until complete failure. This elementary process initiates with a sudden mode I fracture of a group of musculo-elastic units, followed by an elastic recoil of these units, causing mode II separation of these, hence a delamination plane. The presented experimental approach constitutes a basis for observation of other constituents, or for investigations on other tissues and damage mechanisms. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  7. Intestinal Ischemia

    Science.gov (United States)

    ... weight loss Intestinal ischemia Symptoms & causes Diagnosis & treatment Advertisement Mayo Clinic does not endorse companies or products. ... a Job Site Map About This Site Twitter Facebook Google YouTube Pinterest Mayo Clinic is a not- ...

  8. Micro-scaled high-throughput digestion of plant tissue samples for multi-elemental analysis

    Directory of Open Access Journals (Sweden)

    Husted Søren

    2009-09-01

    Full Text Available Abstract Background Quantitative multi-elemental analysis by inductively coupled plasma (ICP spectrometry depends on a complete digestion of solid samples. However, fast and thorough sample digestion is a challenging analytical task which constitutes a bottleneck in modern multi-elemental analysis. Additional obstacles may be that sample quantities are limited and elemental concentrations low. In such cases, digestion in small volumes with minimum dilution and contamination is required in order to obtain high accuracy data. Results We have developed a micro-scaled microwave digestion procedure and optimized it for accurate elemental profiling of plant materials (1-20 mg dry weight. A commercially available 64-position rotor with 5 ml disposable glass vials, originally designed for microwave-based parallel organic synthesis, was used as a platform for the digestion. The novel micro-scaled method was successfully validated by the use of various certified reference materials (CRM with matrices rich in starch, lipid or protein. When the micro-scaled digestion procedure was applied on single rice grains or small batches of Arabidopsis seeds (1 mg, corresponding to approximately 50 seeds, the obtained elemental profiles closely matched those obtained by conventional analysis using digestion in large volume vessels. Accumulated elemental contents derived from separate analyses of rice grain fractions (aleurone, embryo and endosperm closely matched the total content obtained by analysis of the whole rice grain. Conclusion A high-throughput micro-scaled method has been developed which enables digestion of small quantities of plant samples for subsequent elemental profiling by ICP-spectrometry. The method constitutes a valuable tool for screening of mutants and transformants. In addition, the method facilitates studies of the distribution of essential trace elements between and within plant organs which is relevant for, e.g., breeding programmes aiming at

  9. Intestinal Coccidia

    OpenAIRE

    MJ Ggaravi

    2007-01-01

    Intestinal Coccidia are a subclass of Apicomplexa phylum. Eucoccidida are facultative heteroxenous, but some of them are monoxenous. They have sexual and asexual life cycle. Some coccidia are human pathogens, for example: Cryptosporidium: Cryptosporidiums has many species that are mammalian intestinal parasites.C. Parvum specie is a human pathogenic protozoa. Cryptosporidum has circle or ellipse shapes and nearly 4-6 mm. It is transmitted in warm seasons. Oocyst is obtained insexual life cycl...

  10. Isotope Concentrations from 24-h Urine and 3-h Serum Samples Can Be Used to Measure Intestinal Magnesium Absorption in Postmenopausal Women123

    Science.gov (United States)

    Hansen, Karen E.; Nabak, Andrea C.; Johnson, Rachael Erin; Marvdashti, Sheeva; Keuler, Nicholas S.; Shafer, Martin M.; Abrams, Steven A.

    2014-01-01

    Studies suggest a link between magnesium status and osteoporosis. One barrier to more conclusive research on the potential relation is measuring intestinal magnesium absorption (MgA), which requires the use of stable isotopes and a ≥6-d stool or 3-d urine collection. We evaluated alternative methods of measuring MgA. We administered 2 stable magnesium isotopes to 15 postmenopausal women (cohort 1) aged 62 ± 8 y with a dietary magnesium intake of 345 ± 72 mg/d. Participants fasted from 1200 h to 0700 h and then consumed breakfast with ∼23 mg of oral 26Mg and ∼11 mg of i.v. 25Mg. We measured magnesium isotope concentrations in 72-h urine, spot urine (36, 48, 60, and 72 h), and spot serum (1, 3, and 5 h) samples collected after isotope dosing. We calculated MgA using the dose-corrected fraction of isotope concentrations from the 72-h urine collection. We validated new methods in 10 postmenopausal women (cohort 2) aged 59 ± 5 y with a dietary magnesium intake of 325 ± 122 mg/d. In cohort 1, MgA based on the 72-h urine collection was 0.28 ± 0.08. The 72-h MgA correlated most highly with 0–24 h urine MgA value alone (ρ = 0.95, P urine and the 3-h (ρ = 0.93, P urine and 3-h serum MgA values. We conclude that means of 0–24 h urine and 3-h serum MgA provide a reasonable estimate of 72-h MgA. However, if researchers seek to identify small changes in MgA, we recommend a 3-d urine or extended stool collection. This trial was registered at clinicaltrials.gov as NCT01593501. PMID:24500940

  11. Isotope concentrations from 24-h urine and 3-h serum samples can be used to measure intestinal magnesium absorption in postmenopausal women.

    Science.gov (United States)

    Hansen, Karen E; Nabak, Andrea C; Johnson, Rachael Erin; Marvdashti, Sheeva; Keuler, Nicholas S; Shafer, Martin M; Abrams, Steven A

    2014-04-01

    Studies suggest a link between magnesium status and osteoporosis. One barrier to more conclusive research on the potential relation is measuring intestinal magnesium absorption (MgA), which requires the use of stable isotopes and a ≥6-d stool or 3-d urine collection. We evaluated alternative methods of measuring MgA. We administered 2 stable magnesium isotopes to 15 postmenopausal women (cohort 1) aged 62 ± 8 y with a dietary magnesium intake of 345 ± 72 mg/d. Participants fasted from 1200 h to 0700 h and then consumed breakfast with ∼23 mg of oral ²⁶Mg and ∼11 mg of i.v. ²⁵Mg. We measured magnesium isotope concentrations in 72-h urine, spot urine (36, 48, 60, and 72 h), and spot serum (1, 3, and 5 h) samples collected after isotope dosing. We calculated MgA using the dose-corrected fraction of isotope concentrations from the 72-h urine collection. We validated new methods in 10 postmenopausal women (cohort 2) aged 59 ± 5 y with a dietary magnesium intake of 325 ± 122 mg/d. In cohort 1, MgA based on the 72-h urine collection was 0.28 ± 0.08. The 72-h MgA correlated most highly with 0-24 h urine MgA value alone (ρ = 0.95, P urine and the 3-h (ρ = 0.93, P urine and 3-h serum MgA values. We conclude that means of 0-24 h urine and 3-h serum MgA provide a reasonable estimate of 72-h MgA. However, if researchers seek to identify small changes in MgA, we recommend a 3-d urine or extended stool collection.

  12. Museums and disease: using tissue archive and museum samples to study pathogens.

    Science.gov (United States)

    Tsangaras, Kyriakos; Greenwood, Alex D

    2012-01-20

    Molecular studies of archival and fossil samples have traditionally focused on the nucleic acids derived from the host species. However, there has recently been an increase in ancient DNA research on the identification and characterization of infectious agents within the hosts. The study of pathogens from the past provides great opportunities for discovering the causes of historical infection events, characterizing host-microorganism co-evolution and directly investigating the evolution of specific pathogens. Several research teams have been able to isolate and characterize a variety of different bacterial, parasite and viral microorganisms. However, this emerging field is not without obstacles. The diagenetic processes that make ancient DNA research generally difficult are also impediments to ancient pathogen research and perhaps more so given that their DNA may represent an even rarer proportion of the remaining nucleic acids in a fossil sample than host DNA. However, studies performed under controlled conditions and following stringent ancient DNA protocols can and have yielded reliable and often surprising results. This article reviews the advantages, problems, and failures of ancient microbiological research. Copyright © 2011 Elsevier GmbH. All rights reserved.

  13. Advancements in mass spectrometry for biological samples: Protein chemical cross-linking and metabolite analysis of plant tissues

    Energy Technology Data Exchange (ETDEWEB)

    Klein, Adam [Iowa State Univ., Ames, IA (United States)

    2015-01-01

    This thesis presents work on advancements and applications of methodology for the analysis of biological samples using mass spectrometry. Included in this work are improvements to chemical cross-linking mass spectrometry (CXMS) for the study of protein structures and mass spectrometry imaging and quantitative analysis to study plant metabolites. Applications include using matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) to further explore metabolic heterogeneity in plant tissues and chemical interactions at the interface between plants and pests. Additional work was focused on developing liquid chromatography-mass spectrometry (LC-MS) methods to investigate metabolites associated with plant-pest interactions.

  14. Quantitative histological analysis of bovine small intestines before and after processing into natural sausage casings.

    Science.gov (United States)

    Wijnker, Joris J; Tersteeg, M H G; Berends, B R; Vernooij, J C M; Koolmees, P A

    2008-06-01

    A histological study was undertaken to determine the efficiency in the removal of the mucosa and Peyer's patches by standard processing of bovine intestines into natural sausage casings. The second objective was to calculate the quantity of lymphoid and nervous tissue per consumable sausage. For the histological analysis, intestinal samples were collected from 80 beef cattle during the slaughter process. Fresh and cleaned intestines were compared in analyzing the thickness of the intestinal wall, weight reduction during cleaning, removal of the mucosal layer, and the presence of lymphoid and neural tissue after cleaning. The obtained data indicate a weight reduction of about 50% during standard cleaning procedures, as 90% of the mucosa and 48% of the lymphoid tissue are removed. Based on the quantitative histological image analysis, it was calculated that 1 m of cleaned casings, weighing on average 64 g, contains about 2.8 g of mucosa, 0.3 g of lymphoid tissue, and 0.1 g of neural tissue. Assuming, in a worst-case scenario, that the sausage casing is ingested when consuming 200 g of sausage at one meal, this consumption includes 0.09 g of lymphoid tissue and 0.02 g of neural tissue as part of the sausage casing. These data can be included in a risk assessment on the potentialexposure of consumers to bovine spongiform encephalopathy infectivity after eating sausages in beef casings.

  15. Giardia duodenalis Infection Reduces Granulocyte Infiltration in an In Vivo Model of Bacterial Toxin-Induced Colitis and Attenuates Inflammation in Human Intestinal Tissue

    OpenAIRE

    Cotton, James A.; Motta, Jean-Paul; Schenck, L. Patrick; Hirota, Simon A.; Beck, Paul L.; Buret, Andre G.

    2014-01-01

    Giardia duodenalis (syn. G. intestinalis, G. lamblia) is a predominant cause of waterborne diarrheal disease that may lead to post-infectious functional gastrointestinal disorders. Although Giardia-infected individuals could carry as much as 106 trophozoites per centimetre of gut, their intestinal mucosa is devoid of overt signs of inflammation. Recent studies have shown that in endemic countries where bacterial infectious diseases are common, Giardia infections can protect against the develo...

  16. Normal tissue tolerance to external beam radiation therapy: Small bowel; Dose de tolerance a l'irradiation des tissus sains: intestin grele

    Energy Technology Data Exchange (ETDEWEB)

    Martin, E. [Departement de radiotherapie, centre Georges-Francois-Leclerc, 21 - Dijon (France); Pointreau, Y.; Barillot, I. [Service de radiotherapie, centre regional universitaire de cancerologie Henry-S.-Kaplan, hopital Bretonneau, CHRU de Tours, 37 - Tours (France); Roche-Forestier, S. [Centre Jean-Bernard, 72 - Le Mans (France); Barillot, I. [Universite Francois-Rabelais, centre de cancerologie Henry-S.-Kaplan, CHU de Tours, 37 - Tours (France)

    2010-07-15

    The small bowel is a hollow organ involved in the transit and absorption of food. In relation to its anatomical location, a significant amount of this organ is exposed in whole or in part to ionizing radiation in external radiotherapy during abdominal or pelvic irradiation either for primary cancers or metastasis. The acute functional changes during external beam radiation are mainly leading to diarrhea, abdominal pain and bloating. The main late side effects of irradiation of the small intestine are chronic diarrhea, malabsorption with steatorrhoea, abdominal spasms, intestinal obstruction, bleeding and fistulas. The architecture of the small intestine may be considered as parallel with a significant correlation between the irradiated volume of small bowel and the likelihood of acute toxicity, whatever the dose. The literature analysis recommends to consider the volume of small bowel receiving 15 Gy (threshold of 100 to 200 cm{sup 3}) but also 30 and 50 Gy (thresholds of 35 to 300 cm{sup 3}, depending on the level of dose considered). Modern techniques of conformal radiotherapy with modulated intensity will probably have beneficial impact on small bowel toxicity. (authors)

  17. Small Intestine Disorders

    Science.gov (United States)

    ... disease Crohn's disease Infections Intestinal cancer Intestinal obstruction Irritable bowel syndrome Ulcers, such as peptic ulcer Treatment of disorders of the small intestine depends on the cause.

  18. Sampling

    CERN Document Server

    Thompson, Steven K

    2012-01-01

    Praise for the Second Edition "This book has never had a competitor. It is the only book that takes a broad approach to sampling . . . any good personal statistics library should include a copy of this book." —Technometrics "Well-written . . . an excellent book on an important subject. Highly recommended." —Choice "An ideal reference for scientific researchers and other professionals who use sampling." —Zentralblatt Math Features new developments in the field combined with all aspects of obtaining, interpreting, and using sample data Sampling provides an up-to-date treat

  19. A new classification method for MALDI imaging mass spectrometry data acquired on formalin-fixed paraffin-embedded tissue samples.

    Science.gov (United States)

    Boskamp, Tobias; Lachmund, Delf; Oetjen, Janina; Cordero Hernandez, Yovany; Trede, Dennis; Maass, Peter; Casadonte, Rita; Kriegsmann, Jörg; Warth, Arne; Dienemann, Hendrik; Weichert, Wilko; Kriegsmann, Mark

    2017-07-01

    Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) shows a high potential for applications in histopathological diagnosis, and in particular for supporting tumor typing and subtyping. The development of such applications requires the extraction of spectral fingerprints that are relevant for the given tissue and the identification of biomarkers associated with these spectral patterns. We propose a novel data analysis method based on the extraction of characteristic spectral patterns (CSPs) that allow automated generation of classification models for spectral data. Formalin-fixed paraffin embedded (FFPE) tissue samples from N=445 patients assembled on 12 tissue microarrays were analyzed. The method was applied to discriminate primary lung and pancreatic cancer, as well as adenocarcinoma and squamous cell carcinoma of the lung. A classification accuracy of 100% and 82.8%, resp., could be achieved on core level, assessed by cross-validation. The method outperformed the more conventional classification method based on the extraction of individual m/z values in the first application, while achieving a comparable accuracy in the second. LC-MS/MS peptide identification demonstrated that the spectral features present in selected CSPs correspond to peptides relevant for the respective classification. This article is part of a Special Issue entitled: MALDI Imaging, edited by Dr. Corinna Henkel and Prof. Peter Hoffmann. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. The effect of onion exposure on gene expression profiles in intestinal Caco-2 cells

    NARCIS (Netherlands)

    Wit, de N.J.W.; Govers, C.C.F.M.; Boekschoten, M.V.; Mes, J.J.

    2016-01-01

    Background: Human intestinal tissue samples are barely accessible to study potential health benefits of nutritional compounds. Numbers of animals used in animal trials, however, need to be minimalized. Therefore, in this study we explored the applicability of an in vitro model, namely human

  1. Nonlinear dimensionality reduction of gene expression data for visualization and clustering analysis of cancer tissue samples.

    Science.gov (United States)

    Shi, Jinlong; Luo, Zhigang

    2010-08-01

    Gene expression data are the representation of nonlinear interactions among genes and environmental factors. Computing analysis of these data is expected to gain knowledge of gene functions and disease mechanisms. Clustering is a classical exploratory technique of discovering similar expression patterns and function modules. However, gene expression data are usually of high dimensions and relatively small samples, which results in the main difficulty for the application of clustering algorithms. Principal component analysis (PCA) is usually used to reduce the data dimensions for further clustering analysis. While PCA estimates the similarity between expression profiles based on the Euclidean distance, which cannot reveal the nonlinear connections between genes. This paper uses nonlinear dimensionality reduction (NDR) as a preprocessing strategy for feature selection and visualization, and then applies clustering algorithms to the reduced feature spaces. In order to estimate the effectiveness of NDR for capturing biologically relevant structures, the comparative analysis between NDR and PCA is exploited to five real cancer expression datasets. Results show that NDR can perform better than PCA in visualization and clustering analysis of complex gene expression data. Copyright 2010 Elsevier Ltd. All rights reserved.

  2. The Contributions of Human Mini-Intestines to the Study of Intestinal Physiology and Pathophysiology.

    Science.gov (United States)

    Yu, Huimin; Hasan, Nesrin M; In, Julie G; Estes, Mary K; Kovbasnjuk, Olga; Zachos, Nicholas C; Donowitz, Mark

    2017-02-10

    The lack of accessibility to normal and diseased human intestine and the inability to separate the different functional compartments of the intestine even when tissue could be obtained have held back the understanding of human intestinal physiology. Clevers and his associates identified intestinal stem cells and established conditions to grow "mini-intestines" ex vivo in differentiated and undifferentiated conditions. This pioneering work has made a new model of the human intestine available and has begun making contributions to the understanding of human intestinal transport in normal physiologic conditions and the pathophysiology of intestinal diseases. However, this model is reductionist and lacks many of the complexities of normal intestine. Consequently, it is not yet possible to predict how great the advances using this model will be for understanding human physiology and pathophysiology, nor how the model will be modified to include multiple other intestinal cell types and physical forces necessary to more closely approximate normal intestine. This review describes recent studies using mini-intestines, which have readdressed previously established models of normal intestinal transport physiology and newly examined intestinal pathophysiology. The emphasis is on studies with human enteroids grown either as three-dimensional spheroids or two-dimensional monolayers. In addition, comments are provided on mouse studies in cases when human studies have not yet been described.

  3. A method for sampling and tissue preparation of the parathyroid glands in miniature pigs for toxicity studies.

    Science.gov (United States)

    Soshin, Tomomi; Takai, Hirotake; Kato, Chie; Fujii, Etsuko; Matsuo, Saori; Ito, Tsuneo; Suzuki, Masami

    2010-04-01

    Miniature swine (minipigs) are often used in non-rodent toxicity studies. However, unlike other animal species, the parathyroid glands of minipigs are often covered with thymic tissue and are similar in color, making macroscopical identification difficult. We investigated a method for sampling and tissue preparation of the parathyroid glands using 5- to 7-month-old minipigs. The glands were identified by finding the insertion site of a branch from the carotid artery into the cervical part of the thymus. Then the glands were marked and sampled. In a preliminary study, the glands were macroscopically and microscopically detected in 3/8 animals. The glands were not identified macroscopically in 5/8 animals but were detected in 3 of these animals. In total, we succeeded in detection of the glands in 6/8 animals (75%). The method was applied in the main toxicity study, and we succeeded in 100% detection through technical advancement. The method described herein enables high rate of detection and is useful in the pathological evaluation of the parathyroid glands of minipigs.

  4. Solid sampling-graphite furnace atomic absorption spectrometry for the direct determination of boron in plant tissues

    International Nuclear Information System (INIS)

    Resano, M.; Briceno, J.; Aramendia, M.; Belarra, M.A.

    2007-01-01

    In this work, the potential of graphite furnace atomic absorption spectrometry for the direct determination of B in plant tissues has been investigated. Three certified reference materials (NIST SRM 1570a spinach leaves, NIST SRM 1573a tomato leaves and BCR CRM 679 white cabbage) were selected for this study, the goal always being to develop a fast procedure that could be robust enough to provide a satisfactory performance for all of them, without any modifications in the conditions applied. The use of a suitable chemical modifier was found to be essential for obtaining a reproducible and sufficiently sensitive signal for boron solutions. In this regard, the performance of the combination of citric acid plus W (added as a permanent modifier) was noteworthy, resulting in well-defined signal profiles, a remarkable analyte stabilization during the pyrolysis step (up to 2100 deg. C) and minimal memory effects. This mixture of modifiers provided a good performance for the direct analysis of solid samples as well, but only if a suitable temperature program, favoring the interaction between the analyte and the modifiers, was used. Thus, such a temperature program, with two pyrolysis steps and the addition of NH 4 NO 3 in order to carry out the in situ sample microdigestion, was optimized. Under these conditions, the peak areas obtained for both solid samples and aqueous standards were comparable. Finally, the analysis of the samples was carried out. In all cases, a good agreement with the certified values was obtained, while R.S.D. values ranged between 6 and 10%. It can be concluded that the method proposed shows significant advantages for the determination of this complicated element in solid samples such as the use of aqueous standards for calibration, a high sample throughput (20 min per sample), a suitable limit of detection (0.3 μg g -1 ) and reduced risk of analyte losses and contamination

  5. Effect of Da-Cheng-Qi Decoction on Pancreatitis-Associated Intestinal Dysmotility in Patients and in Rat Models

    Directory of Open Access Journals (Sweden)

    Jianlei Zhao

    2015-01-01

    Full Text Available The impairment of intestinal motility and related infectious complications are the predominant clinical phenomenon in patients with severe acute pancreatitis (SAP. We aimed to investigate the effects of Da-Cheng-Qi decoction (DCQD on the gastrointestinal injury in SAP patients and the potential mechanism involved in rats. DCQD was enema administered to 70 patients for 7 days in West China Hospital. Mortality and organ failure during admission were observed and blood samples for laboratory analysis were collected. We also experimentally examined plasma inflammatory cytokines in rat serum and carried the morphometric studies of the gut. Intestinal propulsion index and serum and tissue vasoactive intestinal peptide (VIP were also detected. Though DCQD did not affect the overall incidence of organ failure, it shortened the average time of paralytic intestinal obstruction and decreased the morbidity of infectious complications in patients with SAP. Compared with untreated rats, the DCQD lowered the levels of proinflammatory cytokine and decreased the mean pathological intestinal lesion scores. The VIP level in intestinal tissue or serum in DCQD group was obviously lowered and intestinal propulsion index was significantly improved. In conclusion, DCQD has good effect on pancreatitis-associated intestinal dysmotility in patients and in rat models.

  6. Effect of da-cheng-qi decoction on pancreatitis-associated intestinal dysmotility in patients and in rat models.

    Science.gov (United States)

    Zhao, Jianlei; Zhong, Cejun; He, Zhiyu; Chen, Guangyuan; Tang, Wenfu

    2015-01-01

    The impairment of intestinal motility and related infectious complications are the predominant clinical phenomenon in patients with severe acute pancreatitis (SAP). We aimed to investigate the effects of Da-Cheng-Qi decoction (DCQD) on the gastrointestinal injury in SAP patients and the potential mechanism involved in rats. DCQD was enema administered to 70 patients for 7 days in West China Hospital. Mortality and organ failure during admission were observed and blood samples for laboratory analysis were collected. We also experimentally examined plasma inflammatory cytokines in rat serum and carried the morphometric studies of the gut. Intestinal propulsion index and serum and tissue vasoactive intestinal peptide (VIP) were also detected. Though DCQD did not affect the overall incidence of organ failure, it shortened the average time of paralytic intestinal obstruction and decreased the morbidity of infectious complications in patients with SAP. Compared with untreated rats, the DCQD lowered the levels of proinflammatory cytokine and decreased the mean pathological intestinal lesion scores. The VIP level in intestinal tissue or serum in DCQD group was obviously lowered and intestinal propulsion index was significantly improved. In conclusion, DCQD has good effect on pancreatitis-associated intestinal dysmotility in patients and in rat models.

  7. Simulation-assisted design of microfluidic sample traps for optimal trapping and culture of non-adherent single cells, tissues, and spheroids.

    Science.gov (United States)

    Rousset, Nassim; Monet, Frédéric; Gervais, Thomas

    2017-03-21

    This work focuses on modelling design and operation of "microfluidic sample traps" (MSTs). MSTs regroup a widely used class of microdevices that incorporate wells, recesses or chambers adjacent to a channel to individually trap, culture and/or release submicroliter 3D tissue samples ranging from simple cell aggregates and spheroids, to ex vivo tissue samples and other submillimetre-scale tissue models. Numerous MST designs employing various trapping mechanisms have been proposed in the literature, spurring the development of 3D tissue models for drug discovery and personalized medicine. Yet, there lacks a general framework to optimize trapping stability, trapping time, shear stress, and sample metabolism. Herein, the effects of hydrodynamics and diffusion-reaction on tissue viability and device operation are investigated using analytical and finite element methods with systematic parametric sweeps over independent design variables chosen to correspond to the four design degrees of freedom. Combining different results, we show that, for a spherical tissue of diameter d < 500 μm, the simplest, closest to optimal trap shape is a cube of dimensions w equal to twice the tissue diameter: w = 2d. Furthermore, to sustain tissues without perfusion, available medium volume per trap needs to be 100× the tissue volume to ensure optimal metabolism for at least 24 hours.

  8. Isotope concentrations from 24-h urine and 3-h serum samples can be used to measure intestinal magnesium absorption in postmenopausal women

    Science.gov (United States)

    Studies suggest a link between magnesium status and osteoporosis. One barrier to more conclusive research on the potential relation is measuring intestinal magnesium absorption (MgA), which requires the use of stable isotopes and a >/= 6-d stool or 3-d urine collection. We evaluated alternative meth...

  9. Gene expression related to oxidative stress in the heart of mice after intestinal ischemia

    Energy Technology Data Exchange (ETDEWEB)

    Somaio Neto, Frederico; Ikejiri, Adauto Tsutomu; Bertoletto, Paulo Roberto; Chaves, José Carlos Bertoletto [Universidade Federal da Grande Dourados - UFGD, Dourados, MS (Brazil); Teruya, Roberto [Universidade Federal do Mato Grosso do Sul - UFMS, Campo Grande, MS (Brazil); Fagundes, Djalma José, E-mail: fsomaio@cardiol.br; Taha, Murched Omar [Universidade Federal de São Paulo - UNIFESP, São Paulo, SP (Brazil)

    2014-02-15

    Intestinal ischemia-reperfusion is a frequent clinical event associated to injury in distant organs, especially the heart. To investigate the gene expression of oxidative stress and antioxidant defense in the heart of inbred mice subjected to intestinal ischemia and reperfusion (IR). Twelve mice (C57BL / 6) were assigned to: IR Group (GIR) with 60 minutes of superior mesenteric artery occlusion followed by 60 minutes of reperfusion; Control Group (CG) which underwent anesthesia and laparotomy without IR procedure and was observed for 120 minutes. Intestine and heart samples were processed using the RT-qPCR / Reverse transcriptase-quantitative Polymerase Chain Reaction method for the gene expression of 84 genes related to oxidative stress and oxidative defense (Student's 't' test, p < 0.05). The intestinal tissue (GIR) was noted to have an up-regulation of 65 genes (74.71%) in comparison to normal tissue (CG), and 37 genes (44.04%) were hyper-expressed (greater than three times the threshold allowed by the algorithm). Regarding the remote effects of intestinal I/R in cardiac tissue an up-regulation of 28 genes (33.33%) was seen, but only eight genes (9.52%) were hyper-expressed three times above threshold. Four (7.14%) of these eight genes were expressed in both intestinal and cardiac tissues. Cardiomyocytes with smaller and pyknotic nuclei, rich in heterochromatin with rare nucleoli, indicating cardiac distress, were observed in the GIR. Intestinal I/R caused a statistically significant over expression of 8 genes associated with oxidative stress in remote myocardial tissue.

  10. Docosahexaenoyl serotonin emerges as most potent inhibitor of IL-17 and CCL-20 released by blood mononuclear cells from a series of N-acyl serotonins identified in human intestinal tissue.

    Science.gov (United States)

    Wang, Ya; Balvers, Michiel G J; Hendriks, Henk F J; Wilpshaar, Tessa; van Heek, Tjarda; Witkamp, Renger F; Meijerink, Jocelijn

    2017-09-01

    Fatty acid amides (FAAs), conjugates of fatty acids with ethanolamine, mono-amine neurotransmitters or amino acids are a class of molecules that display diverse functional roles in different cells and tissues. Recently we reported that one of the serotonin-fatty acid conjugates, docosahexaenoyl serotonin (DHA-5-HT), previously found in gut tissue of mouse and pig, attenuates the IL-23-IL-17 signaling axis in LPS-stimulated mice macrophages. However, its presence and effects in humans remained to be elucidated. Here, we report for the first time its identification in human intestinal (colon) tissue, along with a series of related N-acyl serotonins. Furthermore, we tested these fatty acid conjugates for their ability to inhibit the release of IL-17 and CCL-20 by stimulated human peripheral blood mononuclear cells (PBMCs). Serotonin conjugates with palmitic acid (PA-5-HT), stearic acid (SA-5-HT) and oleic acid (OA-5-HT) were detected in higher levels than arachidonoyl serotonin (AA-5-HT) and DHA-5-HT, while eicosapentaenoyl serotonin (EPA-5-HT) could not be quantified. Among these, DHA-5-HT was the most potent in inhibiting IL-17 and CCL-20, typical Th17 pro-inflammatory mediators, by Concanavalin A (ConA)-stimulated human PBMCs. These results underline the idea that DHA-5-HT is a gut-specific endogenously produced mediator with the capacity to modulate the IL-17/Th17 signaling response. Our findings may be of relevance in relation to intestinal inflammatory diseases like Crohn's disease and Ulcerative colitis. Copyright © 2017. Published by Elsevier B.V.

  11. Bile acid receptor TGR5 overexpression is associated with decreased intestinal mucosal injury and epithelial cell proliferation in obstructive jaundice.

    Science.gov (United States)

    Ji, Chen-Guang; Xie, Xiao-Li; Yin, Jie; Qi, Wei; Chen, Lei; Bai, Yun; Wang, Na; Zhao, Dong-Qiang; Jiang, Xiao-Yu; Jiang, Hui-Qing

    2017-04-01

    Bile acids stimulate intestinal epithelial proliferation in vitro. We sought to investigate the role of the bile acid receptor TGR5 in the protection of intestinal epithelial proliferation in obstructive jaundice. Intestinal tissues and serum samples were obtained from patients with malignant obstructive jaundice and from bile duct ligation (BDL) rats. Intestinal permeability and morphological changes in the intestinal mucosa were observed. The functions of TGR5 in cell proliferation in intestinal epithelial injury were determined by overexpression or knockdown studies in Caco-2 and FHs 74 Int cells pretreated with lipopolysaccharide (LPS). Internal biliary drainage was superior to external biliary drainage in recovering intestinal permeability and mucosal histology in patients with obstructive jaundice. In BDL rats, feeding of chenodeoxycholic acid (CDCA) decreased intestinal mucosa injury. The levels of PCNA, a marker of proliferation, increased in response to CDCA feeding and were paralleled by elevated TGR5 expression. CDCA upregulated TGR5 expression and promoted proliferation in Caco-2 and FHs 74 Int cells pretreated with LPS. Overexpression of TGR5 resulted in increased PCNA, cell viability, EdU incorporation, and the proportion of cells in S phase, whereas knockdown of TGR5 had the opposite effect. Our data indicate that bile acids promote intestinal epithelial cell proliferation and decrease mucosal injury by upregulating TGR5 expression in obstructive jaundice. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. A New Sample Substrate for Imaging and Correlating Organic and Trace Metal Composition in Biological Cells and Tissues

    International Nuclear Information System (INIS)

    Miller, L.; Wang, Q.; Smith, R.; Zhong, H.; Elliott, D.; Warren, J.

    2007-01-01

    Many disease processes involve alterations in the chemical makeup of tissue. Synchrotron-based infrared (IR) and X-ray fluorescence (XRF) microscopes are becoming increasingly popular tools for imaging the organic and trace metal compositions of biological materials, respectively, without the need for extrinsic labels or stains. Fourier transform infrared microspectroscopy (FTIRM) provides chemical information on the organic components of a material at a diffraction-limited spatial resolution of 2-10 μm in the mid-infrared region. The synchrotron X-ray fluorescence (SXRF) microprobe is a complementary technique used to probe trace element content in the same systems with a similar spatial resolution. However to be most beneficial, it is important to combine the results from both imaging techniques on a single sample, which requires precise overlap of the IR and X-ray images. In this work, we have developed a sample substrate containing a gold grid pattern on its surface, which can be imaged with both the IR and X-ray microscopes. The substrate consists of a low trace element glass slide that has a gold grid patterned on its surface, where the major and minor parts of the grid contain 25 and 12 nm gold, respectively. This grid pattern can be imaged with the IR microscope because the reflectivity of gold differs as a function of thickness. The pattern can also be imaged with the SXRF microprobe because the Au fluorescence intensity changes with gold thickness. The tissue sample is placed on top of the patterned substrate. The grid pattern's IR reflectivity image and the gold SXRF image are used as fiducial markers for spatially overlapping the IR and SXRF images from the tissue. Results show that IR and X-ray images can be correlated precisely, with a spatial resolution of less than one pixel (i.e., 2-3 microns). The development of this new tool will be presented along with applications to paraffin-embedded metalloprotein crystals, Alzheimer's disease, and hair

  13. Evaluation of Streck tissue fixative, a nonformalin fixative for preservation of stool samples and subsequent parasitologic examination.

    Science.gov (United States)

    Nace, E K; Steurer, F J; Eberhard, M L

    1999-12-01

    We undertook a study to evaluate Streck tissue fixative (STF) as a substitute for formalin and polyvinyl alcohol (PVA) in fecal preservation. A comparison of formalin, PVA, (mercuric chloride based), and STF was done by aliquoting fecal samples into each fixative. Stool specimens were collected in Haiti, and parasites included Cyclospora cayetanensis, Giardia intestinalis, Entamoeba coli, Iodamoeba butschlii, Endolimax nana, Ascaris lumbricoides, Trichuris trichiura, Strongyloides stercoralis, and Necator americanus. Preserved stools were examined at various predetermined times (1 week, 1 month, and 3 months) to establish the quality of the initial preservation as well as the suitability of the fixative for long-term storage. At each time point, stool samples in fixatives were examined microscopically as follows: (i) in wet mounts (with bright-field and epifluorescence microscopy), (ii) in modified acid-fast-, trichrome-, and safranin-stained smears, and (iii) with two commercial test kits. At the time points examined, morphologic features remained comparable for samples fixed with 10% formalin and STF. For comparisons of STF- and 10% formalin-fixed samples, specific findings showed that Cyclospora oocysts retained full fluorescence, modified acid-fast- and safranin-stained smears of Cryptosporidium and Cyclospora oocysts were equal in staining quality, and results were comparable in the immunofluorescence assay and enzyme immunoassay commercial kits. Stool fixed in STF and stained with trichrome showed less-than-acceptable staining quality compared with stool fixed in PVA. STF provides an excellent substitute for formalin as a fixative in routine examination of stool samples for parasites. However, modifications to the trichrome staining procedures will be necessary to improve the staining quality for protozoal cysts fixed in STF to a level comparable to that with PVA.

  14. Use of alkaline or enzymatic sample pretreatment prior to characterization of gold nanoparticles in animal tissue by single-particle ICPMS

    DEFF Research Database (Denmark)

    Löschner, Katrin; Brabrand, Myung Suk Jung; Sloth, Jens Jørgen

    2014-01-01

    concentration obtained by conventional ICPMS analysis of acid-digested tissue. The recovery of AuNPs from enzymatically digested tissue, however, was approximately four times lower. Spiking experiments of blank spleen samples with AuNPs showed that the lower recovery was caused by an inferior transport...

  15. Imaging of wood tissue by ToF-SIMS: Critical evaluation and development of sample preparation techniques

    International Nuclear Information System (INIS)

    Tokareva, E.N.; Fardim, P.; Pranovich, A.V.; Fagerholm, H.-P.; Daniel, G.; Holmbom, B.

    2007-01-01

    Tissue of Norway spruce wood was investigated by time-of-flight secondary-ion mass spectrometry (ToF-SIMS) and field-emission scanning electron microscopy (FE-SEM). Different sample preparation techniques (including sectioning and drying) traditionally used for SEM analysis were compared and critically evaluated. A high contamination of wood surfaces by polytetrafluoroethylene (PTFE) introduced onto the sample during sectioning was detected by ToF-SIMS. A new protocol was developed and images of wood sections dried in different ways were compared. Main location of Mg, Na, K and Ca, but no lignin in membranes of bordered pits in spruce section surfaces was detected by ToF-SIMS imaging if critical point drying (CPD) or acetone extraction followed by drying under nitrogen flow (AEND) was applied. No specific locations of wood components as well as low signals from metal ions (Mg and Na) were observed on the wood surfaces of freeze-dried (FD) or air-dried (AD) sections. It was found that extractive substances, presenting on surfaces of FD and AD wood section, were evenly distributed on the surfaces and interfered with ToF-SIMS analysis. Our results indicated that ToF-SIMS imaging was critically sensitive to the sample preparation technique and a strict protocol for characterization of metals, lignin and polysaccharides in wood was suggested

  16. [Authorization of pathologists for the estimation of the tumor cell percentage on tissue sample for molecular analysis purpose].

    Science.gov (United States)

    Luquain, Alexandra; Arbez-Gindre, Francine; Bedgedjian, Isabelle; Felix, Sophie; Harimenshi, Jean-Marie; Mihai, Ionela-Marcela; Monnien, Franck; Singeorzan, Cristina; Valmary-Degano, Séverine

    2016-08-01

    Before molecular analysis is performed, morphological control with an estimation of the tumour cell percentage (%TC) could have a major impact on mutation detection. Accreditation according to NF EN ISO 15189 commands an authorization through evaluation of skills. The objective of this work was to validate the empowerment of pathologists to estimate %TC in tissue sample prior to molecular analysis. The accreditation technical guidance methods in Medical biology and histopathology were taken as references. %TC was the ratio of tumour cell nuclei on all nuclei within the area selected for the DNA extraction. External evaluations quality scores were used for accuracy. In order to assess the intermediate precision, 35 %TC estimation were performed 15 days apart in 4 samples (biopsies, transparietal biopsies or surgical specimen, either fixed or frozen) by 7 pathologists. Three other cases with interference (inflammation, mucus, necrosis) were evaluated. A result was satisfactory if %TC were within ±20 % of expected percentage obtained by the average of 35 estimates. The performances were satisfactory since no estimate was made more than 20 % of the expected percentage. Low interpathologists reproducibility has been reported in the literature and can have a consequence on molecular analysis in samples with low %TC, where the value reach the analytical sensitivity thresholds of molecular techniques. The current report is an example of a step of the accreditation process, which is a challenge for pathologists' activity in the future. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  17. Exploring the role of paraoxonase-2 in bladder cancer: analyses performed on tissue samples, urines and cell cultures.

    Science.gov (United States)

    Bacchetti, Tiziana; Sartini, Davide; Pozzi, Valentina; Cacciamani, Tiziana; Ferretti, Gianna; Emanuelli, Monica

    2017-04-25

    The enzyme paraoxonase-2 (PON2) is ubiquitously expressed and exerts its antiapoptotic and antioxidative functions in several intracellular compartments.The aim of this study is to investigate the role of PON2 in bladder cancer (BC). The expression levels of PON2 in paired tumor and normal bladder tissue samples and in urinary exfoliated cells from patients affected with BC and healthy donors were evaluated. Moreover, the effect of PON2 overexpression on tumor cell proliferation and susceptibility to oxidative stress was investigated in human bladder cancer cell line T24.Our results showed that PON2 expression levels were significantly higher in BC compared with non-tumor tissue. In urinary exfoliated cells from BC patients, PON2 mRNA levels showed an inverse correlation with tumor stage (pT). Moreover, PON2 overexpression in T24 cells led to a significant increase in tumor cell proliferation and resistance to oxidative stress.The results obtained showed that PON2 could represent a molecular biomarker for bladder cancer and suggest a potential role of the enzyme as a prognostic factor for this neoplasm.

  18. Identification of proteins from cambium tissues of the chinese white poplar (populus tomentosa) sampled during the growing season

    International Nuclear Information System (INIS)

    Xie, J.; Liu, S.; Qi, Q.; Hou, Y.

    2014-01-01

    Various protein extraction methods have been used to investigate Chinese white poplar (Populus tomentosa) proteomics. However, extracting and characterizing proteins from woody plants remains a challenge. Two-dimensional gel electrophoresis is a powerful, widely used method for the analysis of complex protein mixtures extracted from biological samples. The technique separates mixtures of proteins along two dimensions, by isoelectric point and molecular weight, and can resolve thousands of different proteins. Here, we report a new application of two-dimensional gel electrophoresis to investigate the proteomics of P. tomentosa cambium tissues over the course of a growing season. Of three protein extraction methods that we compared (the Tris-phenol method, trichloroacetic acid-acetone method, and trichloroacetic acid-acetone-phenol method), trichloroacetic acid-acetone was the most efficient approach for protein extraction from cambium tissues of P. tomentosa. After extraction, the proteins were separated using two-dimensional gel electrophoresis. The protein quantities of six spots changed over the course of the growing season from February to July. Five spots were identified using matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry, and the sixth spot was identified by liquid chromatography-mass spectrometry. The proteins included enolase, class Ia chitinase, and four unnamed proteins. Our results show the best approach to proteomics in P. tomentosa and reveal trends in protein activities during a growing season in this tree species. (author)

  19. Postconditioning attenuates acute intestinal ischemia–reperfusion injury

    Directory of Open Access Journals (Sweden)

    Ilker Sengul

    2013-03-01

    Full Text Available The aim of this study was to test the hypothesis that postconditioning (POC would reduce the detrimental effects of the acute intestinal ischemia–reperfusion (I/R compared to those of the abrupt onset of reperfusion. POC has a protective effect on intestinal I/R injury by inhibiting events in the early minutes of reperfusion in rats. Twenty-four Wistar–Albino rats were subjected to the occlusion of superior mesenteric artery for 30 minutes, then reperfused for 120 minutes, and randomized to the four different modalities of POC: (1 control (no intervention; (2 POC-3 (three cycles of 10 seconds of reperfusion–reocclusion, 1 minute total intervention; (3 POC-6 (six cycles of 10 seconds of reperfusion–reocclusion, 2 minutes total intervention; and (4 sham operation (laparotomy only. The arterial blood samples [0.3 mL total creatine kinase (CK and 0.6 mL malondialdehyde (MDA] and the intestinal mucosal MDA were collected from each after reperfusion. POC, especially POC-6, was effective in attenuating postischemic pathology by decreasing the intestinal tissue MDA levels, serum total CK activity, inflammation, and total histopathological injury scores. POC exerted a protective effect on the intestinal mucosa by reducing the mesenteric oxidant generation, lipid peroxidation, and neutrophil accumulation. The six-cycle algorithm demonstrated the best protection.

  20. ALP gene expression in cDNA samples from bone tissue engineering using a HA/TCP/Chitosan scaffold

    Science.gov (United States)

    Stephanie, N.; Katarina, H.; Amir, L. R.; Gunawan, H. A.

    2017-08-01

    This study examined the potential use of hydroxyapatite (HA)/tricalcium phosphate (TCP)/Chitosan as a bone tissue engineering scaffold. The potential for using HA/TCP/chitosan as a scaffold was analyzed by measuring expression of the ALP osteogenic gene in cDNA from bone biopsies from four Macaque nemestrina. Experimental conditions included control (untreated), treatment with HA/TCP 70:30, HA/TCP 50:50, and HA/TCP/chitosan. cDNA samples were measured quantitively with Real-Time PCR (qPCR) and semi-quantitively by gel electrophoresis. There were no significant differences in ALP gene expression between treatment subjects after two weeks, but the HA/TCP/chitosan treatment gave the highest level of expression after four weeks. The scaffold using the HA/TCP/chitosan combination induced a higher level of expression of the osteogenic gene ALP than did scaffold without chitosan.

  1. Intestinal cholesterol secretion: future clinical implications

    NARCIS (Netherlands)

    Jakulj, L.; Besseling, J.; Stroes, E. S. G.; Groen, A. K.

    2013-01-01

    Together with the liver, the intestine serves as a homeostatic organ in cholesterol metabolism. Recent evidence has substantiated the pivotal role of the intestine in reverse cholesterol transport (RCT). RCT is a fundamental antiatherogenic pathway, mediating the removal of cholesterol from tissues

  2. Intestinal cholesterol secretion : future clinical implications

    NARCIS (Netherlands)

    Jakulj, L.; Besseling, J.; Stroes, E. S. G.; Groen, A. K.

    2013-01-01

    Together with the liver, the intestine serves as a homeostatic organ in cholesterol metabolism. Recent evidence has substantiated the pivotal role of the intestine in reverse cholesterol transport (RCT). RCT is a fundamental antiatherogenic pathway, mediating the removal of cholesterol from tissues

  3. Reduction of sample size requirements by bilateral versus unilateral research designs in animal models for cartilage tissue engineering.

    Science.gov (United States)

    Orth, Patrick; Zurakowski, David; Alini, Mauro; Cucchiarini, Magali; Madry, Henning

    2013-11-01

    Advanced tissue engineering approaches for articular cartilage repair in the knee joint rely on translational animal models. In these investigations, cartilage defects may be established either in one joint (unilateral design) or in both joints of the same animal (bilateral design). We hypothesized that a lower intraindividual variability following the bilateral strategy would reduce the number of required joints. Standardized osteochondral defects were created in the trochlear groove of 18 rabbits. In 12 animals, defects were produced unilaterally (unilateral design; n=12 defects), while defects were created bilaterally in 6 animals (bilateral design; n=12 defects). After 3 weeks, osteochondral repair was evaluated histologically applying an established grading system. Based on intra- and interindividual variabilities, required sample sizes for the detection of discrete differences in the histological score were determined for both study designs (α=0.05, β=0.20). Coefficients of variation (%CV) of the total histological score values were 1.9-fold increased following the unilateral design when compared with the bilateral approach (26 versus 14%CV). The resulting numbers of joints needed to treat were always higher for the unilateral design, resulting in an up to 3.9-fold increase in the required number of experimental animals. This effect was most pronounced for the detection of small-effect sizes and estimating large standard deviations. The data underline the possible benefit of bilateral study designs for the decrease of sample size requirements for certain investigations in articular cartilage research. These findings might also be transferred to other scoring systems, defect types, or translational animal models in the field of cartilage tissue engineering.

  4. CODIFI (Concordance in Diabetic Foot Ulcer Infection): a cross-sectional study of wound swab versus tissue sampling in infected diabetic foot ulcers in England.

    Science.gov (United States)

    Nelson, Andrea; Wright-Hughes, Alexandra; Backhouse, Michael Ross; Lipsky, Benjamin A; Nixon, Jane; Bhogal, Moninder S; Reynolds, Catherine; Brown, Sarah

    2018-01-31

    To determine the extent of agreement and patterns of disagreement between wound swab and tissue samples in patients with an infected diabetic foot ulcer (DFU). Multicentre, prospective, cross-sectional study. Primary and secondary care foot ulcer/diabetic outpatient clinics and hospital wards across England. Inclusion criteria: consenting patients aged ≥18 years; diabetes mellitus; suspected infected DFU. clinically inappropriate to take either sample. Wound swab obtained using Levine's technique; tissue samples collected using a sterile dermal curette or scalpel. Coprimary: reported presence, and number, of pathogens per sample; prevalence of resistance to antimicrobials among likely pathogens. Secondary: recommended change in antibiotic therapy based on blinded clinical review; adverse events; sampling costs. 400 consenting patients (79% male) from 25 centres.Most prevalent reported pathogens were Staphylococcus aureus (43.8%), Streptococcus (16.7%) and other aerobic Gram-positive cocci (70.6%). At least one potential pathogen was reported from 70.1% of wound swab and 86.1% of tissue samples. Pathogen results differed between sampling methods in 58% of patients, with more pathogens and fewer contaminants reported from tissue specimens.The majority of pathogens were reported significantly more frequently in tissue than wound swab samples (P<0.01), with equal disagreement for S. aureus and Pseudomonas aeruginosa. Blinded clinicians more often recommended a change in antibiotic regimen based on tissue compared with wound swab results (increase of 8.9%, 95% CI 2.65% to 15.3%). Ulcer pain and bleeding occurred more often after tissue collection versus wound swabs (pain: 9.3%, 1.3%; bleeding: 6.8%, 1.5%, respectively). Reports of tissue samples more frequently identified pathogens, and less frequently identified non-pathogens compared with wound swab samples. Blinded clinicians more often recommended changes in antibiotic therapy based on tissue compared with wound

  5. Biased visualization of hypoperfused tissue by computed tomography due to short imaging duration: improved classification by image down-sampling and vascular models

    Energy Technology Data Exchange (ETDEWEB)

    Mikkelsen, Irene Klaerke; Ribe, Lars Riisgaard; Bekke, Susanne Lise; Tietze, Anna; Oestergaard, Leif; Mouridsen, Kim [Aarhus University Hospital, Center of Functionally Integrative Neuroscience, Aarhus C (Denmark); Jones, P.S.; Alawneh, Josef [University of Cambridge, Department of Clinical Neurosciences, Cambridge (United Kingdom); Puig, Josep; Pedraza, Salva [Dr. Josep Trueta Girona University Hospitals, Department of Radiology, Girona Biomedical Research Institute, Girona (Spain); Gillard, Jonathan H. [University of Cambridge, Department of Radiology, Cambridge (United Kingdom); Warburton, Elisabeth A. [Cambrigde University Hospitals, Addenbrooke, Stroke Unit, Cambridge (United Kingdom); Baron, Jean-Claude [University of Cambridge, Department of Clinical Neurosciences, Cambridge (United Kingdom); Centre Hospitalier Sainte Anne, INSERM U894, Paris (France)

    2015-07-15

    Lesion detection in acute stroke by computed-tomography perfusion (CTP) can be affected by incomplete bolus coverage in veins and hypoperfused tissue, so-called bolus truncation (BT), and low contrast-to-noise ratio (CNR). We examined the BT-frequency and hypothesized that image down-sampling and a vascular model (VM) for perfusion calculation would improve normo- and hypoperfused tissue classification. CTP datasets from 40 acute stroke patients were retrospectively analysed for BT. In 16 patients with hypoperfused tissue but no BT, repeated 2-by-2 image down-sampling and uniform filtering was performed, comparing CNR to perfusion-MRI levels and tissue classification to that of unprocessed data. By simulating reduced scan duration, the minimum scan-duration at which estimated lesion volumes came within 10 % of their true volume was compared for VM and state-of-the-art algorithms. BT in veins and hypoperfused tissue was observed in 9/40 (22.5 %) and 17/40 patients (42.5 %), respectively. Down-sampling to 128 x 128 resolution yielded CNR comparable to MR data and improved tissue classification (p = 0.0069). VM reduced minimum scan duration, providing reliable maps of cerebral blood flow and mean transit time: 5 s (p = 0.03) and 7 s (p < 0.0001), respectively. BT is not uncommon in stroke CTP with 40-s scan duration. Applying image down-sampling and VM improve tissue classification. (orig.)

  6. Intestinal absorption of fluorescently labeled nanoparticles.

    Science.gov (United States)

    Simovic, Spomenka; Song, Yunmei; Nann, Thomas; Desai, Tejal A

    2015-07-01

    Characterization of intestinal absorption of nanoparticles is critical in the design of noninvasive anticancer, protein-based, and gene nanoparticle-based therapeutics. Here we demonstrate a general approach for the characterization of the intestinal absorption of nanoparticles and for understanding the mechanisms active in their processing within healthy intestinal cells. It is generally accepted that the cellular processing represents a major drawback of current nanoparticle-based therapeutic systems. In particular, endolysosomal trafficking causes degradation of therapeutic molecules such as proteins, lipids, acid-sensitive anticancer drugs, and genes. To date, investigations into nanoparticle processing within intestinal cells have studied mass transport through Caco-2 cells or everted rat intestinal sac models. We developed an approach to visualize directly the mechanisms of nanoparticle processing within intestinal tissue. These results clearly identify a mechanism by which healthy intestinal cells process nanoparticles and point to the possible use of this approach in the design of noninvasive nanoparticle-based therapies. Advances in nanomedicine have resulted in the development of new therapies for various diseases. Intestinal route of administration remains the easiest and most natural. The authors here designed experiments to explore and characterize the process of nanoparticle transport across the intestinal tissue. In so doing, further insights were gained for future drug design. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. A novel method for single sample multi-axial nanoindentation of hydrated heterogeneous tissues based on testing great white shark jaws.

    Directory of Open Access Journals (Sweden)

    Toni L Ferrara

    Full Text Available Nanomechanical testing methods that are suitable for a range of hydrated tissues are crucial for understanding biological systems. Nanoindentation of tissues can provide valuable insights into biology, tissue engineering and biomimetic design. However, testing hydrated biological samples still remains a significant challenge. Shark jaw cartilage is an ideal substrate for developing a method to test hydrated tissues because it is a unique heterogeneous composite of both mineralized (hard and non-mineralized (soft layers and possesses a jaw geometry that is challenging to test mechanically. The aim of this study is to develop a novel method for obtaining multidirectional nanomechanical properties for both layers of jaw cartilage from a single sample, taken from the great white shark (Carcharodon carcharias. A method for obtaining multidirectional data from a single sample is necessary for examining tissue mechanics in this shark because it is a protected species and hence samples may be difficult to obtain. Results show that this method maintains hydration of samples that would otherwise rapidly dehydrate. Our study is the first analysis of nanomechanical properties of great white shark jaw cartilage. Variation in nanomechanical properties were detected in different orthogonal directions for both layers of jaw cartilage in this species. The data further suggest that the mineralized layer of shark jaw cartilage is less stiff than previously posited. Our method allows multidirectional nanomechanical properties to be obtained from a single, small, hydrated heterogeneous sample. Our technique is therefore suitable for use when specimens are rare, valuable or limited in quantity, such as samples obtained from endangered species or pathological tissues. We also outline a method for tip-to-optic calibration that facilitates nanoindentation of soft biological tissues. Our technique may help address the critical need for a nanomechanical testing method

  8. Robust bioengineered 3D functional human intestinal epithelium.

    Science.gov (United States)

    Chen, Ying; Lin, Yinan; Davis, Kimberly M; Wang, Qianrui; Rnjak-Kovacina, Jelena; Li, Chunmei; Isberg, Ralph R; Kumamoto, Carol A; Mecsas, Joan; Kaplan, David L

    2015-09-16

    Intestinal functions are central to human physiology, health and disease. Options to study these functions with direct relevance to the human condition remain severely limited when using conventional cell cultures, microfluidic systems, organoids, animal surrogates or human studies. To replicate in vitro the tissue architecture and microenvironments of native intestine, we developed a 3D porous protein scaffolding system, containing a geometrically-engineered hollow lumen, with adaptability to both large and small intestines. These intestinal tissues demonstrated representative human responses by permitting continuous accumulation of mucous secretions on the epithelial surface, establishing low oxygen tension in the lumen, and interacting with gut-colonizing bacteria. The newly developed 3D intestine model enabled months-long sustained access to these intestinal functions in vitro, readily integrable with a multitude of different organ mimics and will therefore ensure a reliable ex vivo tissue system for studies in a broad context of human intestinal diseases and treatments.

  9. Next-Generation Sequencing of RNA and DNA Isolated from Paired Fresh-Frozen and Formalin-Fixed Paraffin-Embedded Samples of Human Cancer and Normal Tissue

    Science.gov (United States)

    Hedegaard, Jakob; Thorsen, Kasper; Lund, Mette Katrine; Hein, Anne-Mette K.; Hamilton-Dutoit, Stephen Jacques; Vang, Søren; Nordentoft, Iver; Birkenkamp-Demtröder, Karin; Kruhøffer, Mogens; Hager, Henrik; Knudsen, Bjarne; Andersen, Claus Lindbjerg; Sørensen, Karina Dalsgaard; Pedersen, Jakob Skou; Ørntoft, Torben Falck; Dyrskjøt, Lars

    2014-01-01

    Formalin-fixed, paraffin-embedded (FFPE) tissues are an invaluable resource for clinical research. However, nucleic acids extracted from FFPE tissues are fragmented and chemically modified making them challenging to use in molecular studies. We analysed 23 fresh-frozen (FF), 35 FFPE and 38 paired FF/FFPE specimens, representing six different human tissue types (bladder, prostate and colon carcinoma; liver and colon normal tissue; reactive tonsil) in order to examine the potential use of FFPE samples in next-generation sequencing (NGS) based retrospective and prospective clinical studies. Two methods for DNA and three methods for RNA extraction from FFPE tissues were compared and were found to affect nucleic acid quantity and quality. DNA and RNA from selected FFPE and paired FF/FFPE specimens were used for exome and transcriptome analysis. Preparations of DNA Exome-Seq libraries was more challenging (29.5% success) than that of RNA-Seq libraries, presumably because of modifications to FFPE tissue-derived DNA. Libraries could still be prepared from RNA isolated from two-decade old FFPE tissues. Data were analysed using the CLC Bio Genomics Workbench and revealed systematic differences between FF and FFPE tissue-derived nucleic acid libraries. In spite of this, pairwise analysis of DNA Exome-Seq data showed concordance for 70–80% of variants in FF and FFPE samples stored for fewer than three years. RNA-Seq data showed high correlation of expression profiles in FF/FFPE pairs (Pearson Correlations of 0.90 +/- 0.05), irrespective of storage time (up to 244 months) and tissue type. A common set of 1,494 genes was identified with expression profiles that were significantly different between paired FF and FFPE samples irrespective of tissue type. Our results are promising and suggest that NGS can be used to study FFPE specimens in both prospective and retrospective archive-based studies in which FF specimens are not available. PMID:24878701

  10. Increasing dietary oat fibre decreases the permeability of intestinal mucus.

    Science.gov (United States)

    Mackie, Alan; Rigby, Neil; Harvey, Pascale; Bajka, Balazs

    2016-10-01

    This study investigates the influence of the dietary fibre β-glucan on nutrient composition and mucus permeability. Pigs were fed a standard diet or a diet containing twice the β-glucan content for 3 days (n = 5 per group), followed by the collection of small intestinal mucus and tissue samples. Samples of the consumed diets were subjected to in vitro digestion to determine β-glucan release, nutrient profile and assessment of mucus permeability. In vitro digestion of the diets indicated that 90% of the β-glucan was released in the proximal small intestine. Measurements of intestinal mucus showed a reduction in permeability to 100 nm latex beads and also lipid from the digested enhanced β-glucan diet. The data from this study show for the first time that reducing mass transfer of bile and lipid through the intestinal mucus layer may be one way in which this decrease in bile reabsorption by soluble fibre is enabled.

  11. Immediate postconditioning during reperfusion attenuates intestinal injury.

    Science.gov (United States)

    Liu, Ke-Xuan; Li, Yun-Sheng; Huang, Wen-Qi; Chen, Shu-Qing; Wang, Zhong-Xin; Liu, Jia-Xin; Xia, Zhengyuan

    2009-05-01

    To test the hypothesis that immediate but not delayed ischemic postconditioning (IPo) during reperfusion attenuates intestinal injury, and that ischemic preconditioning (IPC) and IPo may confer synergy in intestinal protection. Prospective laboratory animal study with concurrent control. Adult Sprague-Dawley rats. Intestinal ischemia/reperfusion (II/R) injury in rats was produced by clamping superior mesenteric artery for 60 min followed by 60 min reperfusion; IPC was elicited by 10 min ischemia and 10 min reperfusion before index ischemia; IPo was performed by three cycles of 30 s reperfusion and 30 s ischemia initiated either immediately at the onset of reperfusion (IPo) or after reperfusion for 3 min (delayed-IPo). Combination of IPC and IPo was performed by combining both protocols. Intestinal ischemia/reperfusion resulted in significant intestinal injury evidenced as significant increase in Chiu's scores and wet-to-dry intestine weight ratio accompanied with increases in plasma levels of tumor necrosis factor-alpha and interleukin-6, as well as increases in the intestinal tissue lipid peroxidation product malonediadehyde and myeloperoxidase activity as compared to control animals (all P IPo or their combination (P IPo (P > 0.05). IPC and IPo showed synergistic protection compared with either protocol alone. Ischemic postconditioning reduces intestinal injury, in part, by inhibiting oxidative injury, neutrophils filtration and proinflammatory response. The early period of reperfusion is critical to intestinal protection by IPo, and intestinal protection with IPo can be enhanced by IPC.

  12. TP53 Staining in Tissue Samples of Chronic Lymphocytic Lymphoma Cases: An Immunohistochemical Survey of 51 Cases

    Directory of Open Access Journals (Sweden)

    İbrahim Kulaç

    2017-03-01

    Full Text Available Objective: Chronic lymphocytic leukemia (CLL is the most common lymphoproliferative disease in adults. The aim of this study is to find out if the extent of proliferation centers or the immunohistochemical expression of p53 is related to disease prognosis. Materials and Methods: In the scope of this study, 54 biopsy specimens from 51 patients (50 of lymph nodes; the others of spleen, tonsil, orbit, and liver diagnosed with CLL at the Hacettepe University Department of Pathology in 2000-2013 were reevaluated. The clinical and demographic data of the patients were obtained from our patient database. Biopsy samples were assessed semi-quantitatively for the percentage of proliferation center/total biopsy area (PC/TBA and an immunohistochemical study was performed on representative blocks of tissues for p53 expression. Results: When the patients were divided into two categories according to Rai stage as high and low (stages 0, 1, and 2 vs. stages 3 and 4, it was seen that patients with low Rai stage had a better prognosis than those with high stages (p=0.030. However, there was no statistically significant correlation between overall survival and PC/TBA ratio or p53 expression levels. Conclusion: In our cohort, PC/TBA ratio and immunopositivity of p53 did not show correlations with overall survival.

  13. Performance comparison of machine learning methods for prognosis of hormone receptor status in breast cancer tissue samples.

    Science.gov (United States)

    Kalinli, Adem; Sarikoc, Fatih; Akgun, Hulya; Ozturk, Figen

    2013-06-01

    We examined the classification and prognostic scoring performances of several computer methods on different feature sets to obtain objective and reproducible analysis of estrogen receptor status in breast cancer tissue samples. Radial basis function network, k-nearest neighborhood search, support vector machines, naive bayes, functional trees, and k-means clustering algorithm were applied to the test datasets. Several features were employed and the classification accuracies of each method for these features were examined. The assessment results of the methods on test images were also experimentally compared with those of two experts. According to the results of our experimental work, a combination of functional trees and the naive bayes classifier gave the best prognostic scores indicating very good kappa agreement values (κ=0.899 and κ=0.949, p<0.001) with the experts. This combination also gave the best dichotomization rate (96.3%) for assessment of estrogen receptor status. Wavelet color features provided better classification accuracy than Laws texture energy and co-occurrence matrix features. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  14. Effect of topical anaesthetics on interstitial colloid osmotic pressure in human subcutaneous tissue sampled by wick technique.

    Directory of Open Access Journals (Sweden)

    Hans Jørgen Timm Guthe

    Full Text Available To measure colloid osmotic pressure in interstitial fluid (COP(i from human subcutaneous tissue with the modified wick technique in order to determine influence of topical application of anaesthetics, dry vs. wet wick and implantation time on COP(i.In 50 healthy volunteers interstitial fluid (IF was collected by subcutaneous implantation of multi-filamentous nylon wicks. Study subjects were allocated to two groups; one for comparing COP(i obtained from dry and saline soaked wicks, and one for comparing COP(i from unanaesthetized skin, and skin after application of a eutectic mixture of local anaesthetic (EMLA®, Astra Zeneca cream. IF was sampled from the skin of the shoulders, and implantation time was 30, 60, 75, 90 and 120 min. Colloid osmotic pressure was measured with a colloid osmometer. Pain assessment during the procedure was compared for EMLA cream and no topical anaesthesia using a visual analogue scale (VAS in a subgroup of 10 subjects.There were no significant differences between COP(i obtained from dry compared to wet wicks, except that the values after 75 and 90 min. were somewhat higher for the dry wicks. Topical anaesthesia with EMLA cream did not affect COP(i values. COP(i decreased from 30 to 75 min. of implantation (23.2 ± 4.4 mmHg to 19.6 ± 2.9 mmHg, p = 0.008 and subsequently tended to increase until 120 min. EMLA cream resulted in significant lower VAS score for the procedure.COP(i from subcutaneous tissue was easily obtained and fluid harvesting was well tolerated when topical anaesthetic was used. The difference in COP(i assessed by dry and wet wicks between 75 min. and 90 min. of implantation was in accordance with previous reports. The use of topical analgesia did not influence COP(i and topical analgesia may make the wick technique more acceptable for subjects who dislike technical procedures, including children.ClinicalTrials.gov NCT01044979.

  15. Intestinal intraepithelial lymphocyte cytometric pattern is more accurate than subepithelial deposits of anti-tissue transglutaminase IgA for the diagnosis of celiac disease in lymphocytic enteritis.

    Directory of Open Access Journals (Sweden)

    Fernando Fernández-Bañares

    Full Text Available BACKGROUND & AIMS: An increase in CD3+TCRγδ+ and a decrease in CD3- intraepithelial lymphocytes (IEL is a characteristic flow cytometric pattern of celiac disease (CD with atrophy. The aim was to evaluate the usefulness of both CD IEL cytometric pattern and anti-TG2 IgA subepithelial deposit analysis (CD IF pattern for diagnosing lymphocytic enteritis due to CD. METHODS: Two-hundred and five patients (144 females who underwent duodenal biopsy for clinical suspicion of CD and positive celiac genetics were prospectively included. Fifty had villous atrophy, 70 lymphocytic enteritis, and 85 normal histology. Eight patients with non-celiac atrophy and 15 with lymphocytic enteritis secondary to Helicobacter pylori acted as control group. Duodenal biopsies were obtained to assess both CD IEL flow cytometric (complete or incomplete and IF patterns. RESULTS: Sensitivity of IF, and complete and incomplete cytometric patterns for CD diagnosis in patients with positive serology (Marsh 1+3 was 92%, 85 and 97% respectively, but only the complete cytometric pattern had 100% specificity. Twelve seropositive and 8 seronegative Marsh 1 patients had a CD diagnosis at inclusion or after gluten free-diet, respectively. CD cytometric pattern showed a better diagnostic performance than both IF pattern and serology for CD diagnosis in lymphocytic enteritis at baseline (95% vs 60% vs 60%, p = 0.039. CONCLUSIONS: Analysis of the IEL flow cytometric pattern is a fast, accurate method for identifying CD in the initial diagnostic biopsy of patients presenting with lymphocytic enteritis, even in seronegative patients, and seems to be better than anti-TG2 intestinal deposits.

  16. Intestinal pseudo-obstruction

    Science.gov (United States)

    Primary intestinal pseudo-obstruction; Acute colonic ileus; Colonic pseudo-obstruction; Idiopathic intestinal pseudo-obstruction; Ogilvie syndrome; Chronic intestinal pseudo-obstruction; Paralytic ileus - pseudo-obstruction

  17. Intestinal myiasis

    Directory of Open Access Journals (Sweden)

    U S Udgaonkar

    2012-01-01

    Full Text Available Purpose: Intestinal myiasis is a condition when the fly larvae inhabit the gastrointestinal tract and are passed out in faeces. This type of infestation results when eggs or larvae of the fly, deposited on food are inadvertently taken by man. They survive the unfavourable conditions within the gastrointestinal tract and produce disturbances, which may vary from mild to severe. The condition is not uncommon and is often misdiagnosed as pinworm infestation. Correct diagnosis by the clinical microbiologist is important to avoid unnecessary treatment. Materials and Methods: We had 7 cases of intestinal myiasis. In 2 cases the larvae were reared to adult fly in modified meat and sand medium (developed by Udgaonkar. This medium is simple and can be easily prepared in the laboratory. Results: Of the 7 larvae, 5 were Sarcophaga haemorrhoidalis, 1 Megaselia species and 1 was identified as Muscina stabulans. Conclusions: S. haemorrhoidalis was the commonest maggot involved. A high index of suspicion is required for clinical diagnosis when the patient complains of passing wriggling worms in faeces for a long period without any response to antihelminthics. The reason for long duration of illness and recurrence of infestation is baffling. The nearest to cure was colonic wash. We feel prevention is of utmost importance, which is to avoid eating food articles with easy access to flies.

  18. Diversity of human small intestinal Streptococcus and Veillonella populations

    NARCIS (Netherlands)

    Bogert, B. van den; Erkus, O.; Boekhorst, J.; Goffau, M. de; Smid, E.J.; Zoetendal, E.G.; Kleerebezem, M

    2013-01-01

    Molecular and cultivation approaches were employed to study the phylogenetic richness and temporal dynamics of Streptococcus and Veillonella populations in the small intestine. Microbial profiling of human small intestinal samples collected from four ileostomy subjects at four time points displayed

  19. Diversity of human small intestinal Streptococcus and Veillonella populations

    NARCIS (Netherlands)

    van den Bogert, Bartholomeus; Erkus, Oylum; Boekhorst, Jos; de Goffau, Marcus; Smid, Eddy J.; Zoetendal, Erwin G.; Kleerebezem, Michiel

    Molecular and cultivation approaches were employed to study the phylogenetic richness and temporal dynamics of Streptococcus and Veillonella populations in the small intestine. Microbial profiling of human small intestinal samples collected from four ileostomy subjects at four time points displayed

  20. Simultaneous, quantitative measurement of local blood flow and glucose utilization in tissue samples in normal and injured feline brain.

    Science.gov (United States)

    DeWitt, D S; Yuan, X Q; Becker, D P; Hayes, R L

    1988-01-01

    Cerebral blood flow (CBF) and local cerebral glucose utilization (LCGU) were measured using radioactive microspheres and [14C]2-deoxyglucose, respectively, in 26 brain regions in control animals (n = 8) and in animals (n = 4) sustaining low-level experimental brain injury. Examination of the initial (resting) CBF measurement in the uninjured cats revealed two subgroups with significantly (p less than 0.01) different CBF levels. In uninjured cats with normal CBF levels (33.4 +/- 1.8 ml/100 g/min) there was a close linear relationship between CBF and LCGU (n = 0.71, p less than 0.01). In contrast, the remainder of the uninjured cats exhibited abnormally high levels of CBF (72.6 +/- 9.9 ml/100 g/min) and the absence of a close relationship between CBF and LCGU (r = 0.27). One hour following low-level (2.0 atm) fluid percussion brain injury, CBF was increased and LCGU was decreased, though not significantly. The relationship between CBF and LCGU remained intact (r = 0.66, p less than 0.01) in most brain regions. However, the relationship between CBF and LCGU in the hippocampus differed significantly from the relationship between the two parameters in the rest of the brain. Thus, the use of the radioactive microsphere method for CBF measurements allows multiple measurements of CBF and permits the assessment of the status of the cerebral vasculature prior to experimental manipulations such as traumatic brain injury. In view of our current findings of an abnormal relationship between CBF and LCGU in cats with high resting CBF levels, this is an important advantage. In addition, the combination of the microsphere and 2-DG techniques within the same tissue samples allows for the investigation of the effects of traumatic injury on the important relationship between CBF and LCGU.

  1. Influence of sample preparation and reliability of automated numerical refocusing in stain-free analysis of dissected tissues with quantitative phase digital holographic microscopy

    Science.gov (United States)

    Kemper, Björn; Lenz, Philipp; Bettenworth, Dominik; Krausewitz, Philipp; Domagk, Dirk; Ketelhut, Steffi

    2015-05-01

    Digital holographic microscopy (DHM) has been demonstrated to be a versatile tool for high resolution non-destructive quantitative phase imaging of surfaces and multi-modal minimally-invasive monitoring of living cell cultures in-vitro. DHM provides quantitative monitoring of physiological processes through functional imaging and structural analysis which, for example, gives new insight into signalling of cellular water permeability and cell morphology changes due to toxins and infections. Also the analysis of dissected tissues quantitative DHM phase contrast prospects application fields by stain-free imaging and the quantification of tissue density changes. We show that DHM allows imaging of different tissue layers with high contrast in unstained tissue sections. As the investigation of fixed samples represents a very important application field in pathology, we also analyzed the influence of the sample preparation. The retrieved data demonstrate that the quality of quantitative DHM phase images of dissected tissues depends strongly on the fixing method and common staining agents. As in DHM the reconstruction is performed numerically, multi-focus imaging is achieved from a single digital hologram. Thus, we evaluated the automated refocussing feature of DHM for application on different types of dissected tissues and revealed that on moderately stained samples highly reproducible holographic autofocussing can be achieved. Finally, it is demonstrated that alterations of the spatial refractive index distribution in murine and human tissue samples represent a reliable absolute parameter that is related of different degrees of inflammation in experimental colitis and Crohn's disease. This paves the way towards the usage of DHM in digital pathology for automated histological examinations and further studies to elucidate the translational potential of quantitative phase microscopy for the clinical management of patients, e.g., with inflammatory bowel disease.

  2. Direct detection of the AR-E211 G > A gene polymorphism from blood and tissue samples without DNA isolation.

    Science.gov (United States)

    Reptova, Silvie; Trtkova, Katerina Smesny; Kolar, Zdenek

    2014-04-01

    The pathogenesis of prostate cancer (CaP) involves alterations in a gene structure of the androgen receptor (AR). The single nucleotide polymorphism AR-E211 G > A localized in exon 1 of the AR gene (G1733A) was detected using direct polymerase chain reaction and restriction digestion (PCR-RFLP) method on blood and tissue samples without prior DNA isolation. We used blood samples of patients with a diagnosis of benign prostatic hyperplasia (BPH) or CaP. From monitored group of CaP patients were selected specimen in formalin-fixed paraffin-embedded tissue blocks with morphology of BPH and CaP. The main objective of our study was to develop a method based the direct PCR-RFLP analysis from blood and tissue without prior DNA isolation for faster genotyping analysis of a large number of samples. We found no statistically significant differences in allelic % of the AR-E211 G > A polymorphism between BPH and CaP patients (p ≤ 0.8462). Genotyping of the AR-E211 G > A variant in blood was not identical with tumor tissue genotyping analysis. Significant agreement between blood and tissue AR-E211 G > A polymorphism only in non-tumor tissue focus was confirmed. Although we analyzed a limited number of the tissue samples, we suppose that a presence of the minor allele A may be associated with cancer transformation-induced changes of the modified AR gene.

  3. Intestinal Failure (Short Bowel Syndrome)

    Science.gov (United States)

    Intestinal Failure (Short Bowel Syndrome) What is intestinal failure? Intestinal failure occurs when a significant portion of the small ... intestine does. Who is at risk for intestinal failure? N Babies (usually premature) who have had surgery ...

  4. A statistical model and national data set for partioning fish-tissue mercury concentration variation between spatiotemporal and sample characteristic effects

    Science.gov (United States)

    Wente, Stephen P.

    2004-01-01

    Many Federal, Tribal, State, and local agencies monitor mercury in fish-tissue samples to identify sites with elevated fish-tissue mercury (fish-mercury) concentrations, track changes in fish-mercury concentrations over time, and produce fish-consumption advisories. Interpretation of such monitoring data commonly is impeded by difficulties in separating the effects of sample characteristics (species, tissues sampled, and sizes of fish) from the effects of spatial and temporal trends on fish-mercury concentrations. Without such a separation, variation in fish-mercury concentrations due to differences in the characteristics of samples collected over time or across space can be misattributed to temporal or spatial trends; and/or actual trends in fish-mercury concentration can be misattributed to differences in sample characteristics. This report describes a statistical model and national data set (31,813 samples) for calibrating the aforementioned statistical model that can separate spatiotemporal and sample characteristic effects in fish-mercury concentration data. This model could be useful for evaluating spatial and temporal trends in fishmercury concentrations and developing fish-consumption advisories. The observed fish-mercury concentration data and model predictions can be accessed, displayed geospatially, and downloaded via the World Wide Web (http://emmma.usgs.gov). This report and the associated web site may assist in the interpretation of large amounts of data from widespread fishmercury monitoring efforts.

  5. Partial Enteral Nutrition Preserves Elements of Gut Barrier Function, Including Innate Immunity, Intestinal Alkaline Phosphatase (IAP) Level, and Intestinal Microbiota in Mice

    OpenAIRE

    Wan, Xiao; Bi, Jingcheng; Gao, Xuejin; Tian, Feng; Wang, Xinying; Li, Ning; Li, Jieshou

    2015-01-01

    Lack of enteral nutrition (EN) during parenteral nutrition (PN) leads to higher incidence of infection because of gut barrier dysfunction. However, the effects of partial EN on intestina linnate immunity, intestinal alkaline phosphatase (IAP) and microbiota remain unclear. The mice were randomized into six groups to receive either standard chow or isocaloric and isonitrogenous nutritional support with variable partial EN to PN ratios. Five days later, the mice were sacrificed and tissue sampl...

  6. Intestinal Microbiota Influences Non-intestinal Related Autoimmune Diseases

    Directory of Open Access Journals (Sweden)

    Maria C. Opazo

    2018-03-01

    Full Text Available The human body is colonized by millions of microorganisms named microbiota that interact with our tissues in a cooperative and non-pathogenic manner. These microorganisms are present in the skin, gut, nasal, oral cavities, and genital tract. In fact, it has been described that the microbiota contributes to balancing the immune system to maintain host homeostasis. The gut is a vital organ where microbiota can influence and determine the function of cells of the immune system and contributes to preserve the wellbeing of the individual. Several articles have emphasized the connection between intestinal autoimmune diseases, such as Crohn's disease with dysbiosis or an imbalance in the microbiota composition in the gut. However, little is known about the role of the microbiota in autoimmune pathologies affecting other tissues than the intestine. This article focuses on what is known about the role that gut microbiota can play in the pathogenesis of non-intestinal autoimmune diseases, such as Grave's diseases, multiple sclerosis, type-1 diabetes, systemic lupus erythematosus, psoriasis, schizophrenia, and autism spectrum disorders. Furthermore, we discuss as to how metabolites derived from bacteria could be used as potential therapies for non-intestinal autoimmune diseases.

  7. Chronic Dietary Intake of Enniatin B in Broiler Chickens Has Low Impact on Intestinal Morphometry and Hepatic Histology, and Shows Limited Transfer to Liver Tissue

    Directory of Open Access Journals (Sweden)

    Sophie Fraeyman

    2018-01-01

    Full Text Available The Fusarium mycotoxin enniatin B (ENN B is a so-called emerging mycotoxin frequently contaminating poultry feed. To investigate the impact of chronic ENN B exposure on animal health, broiler chickens were fed either a diet naturally contaminated with ENN B (2352 µg/kg or a control diet (135 µg/kg for 2, 7, 14, or 21 days. ENN B concentrations were determined in plasma and liver using a validated ultra-high performance liquid chromatography—tandem mass spectrometry UHPLC-MS/MS method. Liver was evaluated histologically, and the villus length and crypt depth of the duodenum, jejunum, and ileum were measured. Histopathology of the livers did not reveal major abnormalities. Feeding an ENN B-contaminated diet could possibly inhibit the proliferation of enterocytes in the duodenal crypts, but did not affect villus length, crypt depth, or villus length-crypt depth ratio of the jejunum and ileum. ENN B levels in plasma and liver were significantly higher in the ENN B-fed group and ranged between <25–264 pg/mL and <0.05–0.85 ng/g, respectively. ENN B carry-over rates from feed to liver tissue were 0.005–0.014% and 0.034–0.109% in the ENN B and control group, respectively. Carry-over rates were low and indicated a limited contribution of poultry tissue-derived products to the total dietary ENN B intake for humans. The above results support the opinion of the European Food Safety Authority stating that adverse health effects from ENN B in broiler chickens are unlikely.

  8. Automated column liquid chromatographic determination of amoxicillin and cefadroxil in bovine serum and muscle tissue using on-line dialysis for sample preparation

    NARCIS (Netherlands)

    Snippe, N; van de Merbel, N C; Ruiter, F P; Steijger, O M; Lingeman, H; Brinkman, U A

    1994-01-01

    A fully automated method is described for the determination of amoxicillin and cefadroxil in bovine serum and muscle tissue. The method is based on the on-line combination of dialysis and solid-phase extraction for sample preparation, and column liquid chromatography with ultraviolet detection. In

  9. Analyzing synovial tissue samples. What can we learn about early rheumatoid arthritis, the heterogeneity of the disease, and the effects of treatment?

    NARCIS (Netherlands)

    Tak, Paul P.

    2005-01-01

    The synovium is the key target of the disease process in rheumatoid arthritis (RA). Examination of synovial tissue samples may provide insight into the events that take place in different phases of the disease and may help to decipher the mechanism of action of antirheumatic treatment. This review

  10. Color Tissue Doppler to Analyze Fetal Cardiac Time Intervals: Normal Values and Influence of Sample Gate Size.

    Science.gov (United States)

    Willruth, Arne Michael; Steinhard, Johannes; Enzensberger, Christian; Axt-Fliedner, Roland; Gembruch, Ulrich; Doelle, Astrid; Dimitriou, Ioanna; Fimmers, Rolf; Bahlmann, Franz

    2018-02-01

     To assess the time intervals of the cardiac cycle in healthy fetuses in the second and third trimester using color tissue Doppler imaging (cTDI) and to evaluate the influence of different sizes of sample gates on time interval values.  Time intervals were measured from the cTDI-derived Doppler waveform using a small and large region of interest (ROI) in healthy fetuses.  40 fetuses were included. The median gestational age at examination was 26 + 1 (range: 20 + 5 - 34 + 5) weeks. The median frame rate was 116/s (100 - 161/s) and the median heart rate 143 (range: 125 - 158) beats per minute (bpm). Using small and large ROIs, the second trimester right ventricular (RV) mean isovolumetric contraction times (ICTs) were 39.8 and 41.4 ms (p = 0.17), the mean ejection times (ETs) were 170.2 and 164.6 ms (p ICTs were 36.2 and 39.4 ms (p = 0.05), the mean ETs were 167.4 and 164.5 ms (p = 0.013), the mean IRTs were 53.9 and 57.1 ms (p = 0.05), respectively. The third trimester RV mean ICTs were 50.7 and 50.4 ms (p = 0.75), the mean ETs were 172.3 and 181.4 ms (p = 0.49), the mean IRTs were 50.2 and 54.6 ms (p = 0.03); the LV mean ICTs were 45.1 and 46.2 ms (p = 0.35), the mean ETs were 175.2 vs. 172.9 ms (p = 0.29), the mean IRTs were 47.1 and 50.0 ms (p = 0.01), respectively.  Isovolumetric time intervals can be analyzed precisely and relatively independent of ROI size. In the near future, automatic time interval measurement using ultrasound systems will be feasible and the analysis of fetal myocardial function can become part of the clinical routine. © Georg Thieme Verlag KG Stuttgart · New York.

  11. Primary human alveolar bone cells isolated from tissue samples acquired at periodontal surgeries exhibit sustained proliferation and retain osteogenic phenotype during in vitro expansion.

    Directory of Open Access Journals (Sweden)

    Darja Marolt

    Full Text Available OBJECTIVES: Bone tissue regeneration requires a source of viable, proliferative cells with osteogenic differentiation capacity. Periodontal surgeries represent an opportunity to procure small amounts of autologous tissues for primary cell isolation. Our objective was to assess the potential of human alveolar bone as a source of autologous osteogenic cells for tissue engineering and biomaterials and drug testing studies. MATERIALS AND METHODS: Alveolar bone tissue was obtained from 37 patients undergoing routine periodontal surgery. Tissue harvesting and cell isolation procedures were optimized to isolate viable cells. Primary cells were subcultured and characterized with respect to their growth characteristics, gene expression of osteogenic markers, alkaline phosphatase activity and matrix mineralization, under osteogenic stimulation. RESULTS: Alveolar bone cells were successfully isolated from 28 of the 30 samples harvested with bone forceps, and from 2 of the 5 samples obtained by bone drilling. The yield of cells in primary cultures was variable between the individual samples, but was not related to the site of tissue harvesting and the patient age. In 80% of samples (n = 5, the primary cells proliferated steadily for eight subsequent passages, reaching cumulative numbers over 10(10 cells. Analyses confirmed stable gene expression of alkaline phosphatase, osteopontin and osteocalcin in early and late cell passages. In osteogenic medium, the cells from late passages increased alkaline phosphatase activity and accumulated mineralized matrix, indicating a mature osteoblastic phenotype. CONCLUSIONS: Primary alveolar bone cells exhibited robust proliferation and retained osteogenic phenotype during in vitro expansion, suggesting that they can be used as an autologous cell source for bone regenerative therapies and various in vitro studies.

  12. Fabrication of a multi-layer three-dimensional scaffold with controlled porous micro-architecture for application in small intestine tissue engineering.

    Science.gov (United States)

    Knight, Toyin; Basu, Joydeep; Rivera, Elias A; Spencer, Thomas; Jain, Deepak; Payne, Richard

    2013-01-01

    Various methods can be employed to fabricate scaffolds with characteristics that promote cell-to-material interaction. This report examines the use of a novel technique combining compression molding with particulate leaching to create a unique multi-layered scaffold with differential porosities and pore sizes that provides a high level of control to influence cell behavior. These cell behavioral responses were primarily characterized by bridging and penetration of two cell types (epithelial and smooth muscle cells) on the scaffold in vitro. Larger pore sizes corresponded to an increase in pore penetration, and a decrease in pore bridging. In addition, smaller cells (epithelial) penetrated further into the scaffold than larger cells (smooth muscle cells). In vivo evaluation of a multi-layered scaffold was well tolerated for 75 d in a rodent model. This data shows the ability of the components of multi-layered scaffolds to influence cell behavior, and demonstrates the potential for these scaffolds to promote desired tissue outcomes in vivo.

  13. Long-term detection of seasonal influenza RNA in faeces and intestine.

    Science.gov (United States)

    Hirose, R; Daidoji, T; Naito, Y; Watanabe, Y; Arai, Y; Oda, T; Konishi, H; Yamawaki, M; Itoh, Y; Nakaya, T

    2016-09-01

    Some cases of seasonal influenza virus (human influenza A virus (IAV)/human influenza B virus (IBV)) are associated with abdominal symptoms. Although virus RNA has been detected in faeces, intestinal infection has not been clearly demonstrated. We aimed to provide evidence that IAV/IBV infects the human intestine. This prospective observational study measured virus RNA in faecal and sputum samples from 22 patients infected with IAV/IBV (19 IAV positive and three IBV positive). Nineteen patients were included in the analysis and were assigned to faecal IAV-positive and -negative groups. Virus kinetics were examined in faecal samples from an IAV-infected patient (patient 1) and an IBV-infected patient (patient 2). Finally, intestinal tissue from an IAV-diagnosed patient who developed haemorrhagic colitis and underwent colonoscopy was examined for the presence of replicating IAV (patient 3). Virus RNA was detected in faecal samples from 8/22 IAV/IBV-infected patients (36.4%). Diarrhoea occurred significantly more often in the faecal IAV-positive group (p 0.002). In patients 1 and 2, virus RNA became undetectable in sputum on days 7 and 10 after infection, respectively, but was detected in faeces for a further 2 weeks. Virus mRNA and antigens were detected in intestinal tissues (mucosal epithelium of the sigmoid colon) from patient 3. These findings suggest that IAV/IBV infects within the intestinal tract; thus, the human intestine may be an additional target organ for IAV/IBV infection. Copyright © 2016 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  14. Influence of parasite density and sample storage time on the reliability of Entamoeba histolytica-specific PCR from formalin-fixed and paraffin-embedded tissues.

    Science.gov (United States)

    Frickmann, Hagen; Tenner-Racz, Klara; Eggert, Petra; Schwarz, Norbert G; Poppert, Sven; Tannich, Egbert; Hagen, Ralf M

    2013-12-01

    We report on the reliability of polymerase chain reaction (PCR) for the detection of Entamoeba histolytica from formalin-fixed, paraffin-embedded tissue in comparison with microscopy and have determined predictors that may influence PCR results. E. histolytica-specific and Entamoeba dispar-specific real-time PCR and microscopy from adjacent histologic sections were performed using a collection of formalin-fixed, paraffin-embedded tissue specimens obtained from patients with invasive amebiasis. Specimens had been collected during the previous 4 decades. Association of sample age, parasite density, and reliability of PCR was analyzed. E. histolytica PCR was positive in 20 of 34 biopsies (58.8%); 2 of these 20 were microscopically negative for amebae in neighboring tissue sections. PCR was negative in 9 samples with visible amebae in neighboring sections and in 5 samples without visible parasites in neighboring sections. PCR was negative in all specimens that were older than 3 decades. Low parasite counts and sample ages older than 20 years were predictors for false-negative PCR results. All samples were negative for E. dispar DNA. PCR is suitable for the detection of E. histolytica in formalin-fixed, paraffin-embedded tissue samples that are younger than 2 decades and that contain intermediate to high parasite numbers. Negative results in older samples were due to progressive degradation of DNA over time as indicated by control PCRs targeting the human 18S rRNA gene. Moreover, our findings support previous suggestions that only E. histolytica but not E. dispar is responsible for invasive amebiasis.

  15. SU-F-J-193: Efficient Dose Extinction Method for Water Equivalent Path Length (WEPL) of Real Tissue Samples for Validation of CT HU to Stopping Power Conversion

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, R; Baer, E; Jee, K; Sharp, G; Flanz, J; Lu, H [Massachusetts General Hospital and Harvard Medical School, Boston, MA (United States)

    2016-06-15

    Purpose: For proton therapy, an accurate model of CT HU to relative stopping power (RSP) conversion is essential. In current practice, validation of these models relies solely on measurements of tissue substitutes with standard compositions. Validation based on real tissue samples would be much more direct and can address variations between patients. This study intends to develop an efficient and accurate system based on the concept of dose extinction to measure WEPL and retrieve RSP in biological tissue in large number of types. Methods: A broad AP proton beam delivering a spread out Bragg peak (SOBP) is used to irradiate the samples with a Matrixx detector positioned immediately below. A water tank was placed on top of the samples, with the water level controllable in sub-millimeter by a remotely controlled dosing pump. While gradually lowering the water level with beam on, the transmission dose was recorded at 1 frame/sec. The WEPL were determined as the difference between the known beam range of the delivered SOBP (80%) and the water level corresponding to 80% of measured dose profiles in time. A Gammex 467 phantom was used to test the system and various types of biological tissue was measured. Results: RSP for all Gammex inserts, expect the one made with lung-450 material (<2% error), were determined within ±0.5% error. Depends on the WEPL of investigated phantom, a measurement takes around 10 min, which can be accelerated by a faster pump. Conclusion: Based on the concept of dose extinction, a system was explored to measure WEPL efficiently and accurately for a large number of samples. This allows the validation of CT HU to stopping power conversions based on large number of samples and real tissues. It also allows the assessment of beam uncertainties due to variations over patients, which issue has never been sufficiently studied before.

  16. SU-F-J-193: Efficient Dose Extinction Method for Water Equivalent Path Length (WEPL) of Real Tissue Samples for Validation of CT HU to Stopping Power Conversion

    International Nuclear Information System (INIS)

    Zhang, R; Baer, E; Jee, K; Sharp, G; Flanz, J; Lu, H

    2016-01-01

    Purpose: For proton therapy, an accurate model of CT HU to relative stopping power (RSP) conversion is essential. In current practice, validation of these models relies solely on measurements of tissue substitutes with standard compositions. Validation based on real tissue samples would be much more direct and can address variations between patients. This study intends to develop an efficient and accurate system based on the concept of dose extinction to measure WEPL and retrieve RSP in biological tissue in large number of types. Methods: A broad AP proton beam delivering a spread out Bragg peak (SOBP) is used to irradiate the samples with a Matrixx detector positioned immediately below. A water tank was placed on top of the samples, with the water level controllable in sub-millimeter by a remotely controlled dosing pump. While gradually lowering the water level with beam on, the transmission dose was recorded at 1 frame/sec. The WEPL were determined as the difference between the known beam range of the delivered SOBP (80%) and the water level corresponding to 80% of measured dose profiles in time. A Gammex 467 phantom was used to test the system and various types of biological tissue was measured. Results: RSP for all Gammex inserts, expect the one made with lung-450 material (<2% error), were determined within ±0.5% error. Depends on the WEPL of investigated phantom, a measurement takes around 10 min, which can be accelerated by a faster pump. Conclusion: Based on the concept of dose extinction, a system was explored to measure WEPL efficiently and accurately for a large number of samples. This allows the validation of CT HU to stopping power conversions based on large number of samples and real tissues. It also allows the assessment of beam uncertainties due to variations over patients, which issue has never been sufficiently studied before.

  17. Application of Enhanced Point Estimators to a Sample of In Vivo CT-derived Facial Soft Tissue Thicknesses.

    Science.gov (United States)

    Parks, Connie L; Kyllonen, Kelsey M; Monson, Keith L

    2017-07-01

    Facial approximations based on facial soft tissue depth measurement tables often utilize the arithmetic mean as a central tendency estimator. Stephan et al. (J Forensic Sci 2013;58:1439) suggest that the shorth and 75-shormax statistics are better suited to describe the central tendency of non-normal soft tissue depth data, while also accommodating normal distributions. The shorth, 75-shormax, arithmetic mean, and other central tendency estimators were evaluated using a CT-derived facial soft tissue depth dataset. Differences between arithmetic mean and shorth mean for the tissue depths examined ranged from 0 mm to +2.3 mm (average 0.6 mm). Differences between the arithmetic mean plus one standard deviation (to approximate the same data points covered by the 75-shormax) and 75-shormax values ranged from -0.8 mm to +0.7 mm (average 0.2 mm). The results of this research suggest that few practical differences exist across the central tendency point estimators for the evaluated soft tissue depth dataset. © 2017 American Academy of Forensic Sciences.

  18. Diversity and functions of intestinal mononuclear phagocytes

    DEFF Research Database (Denmark)

    Joeris, Thorsten; Müller-Luda, K; Agace, William Winston

    2017-01-01

    The intestinal lamina propria (LP) contains a diverse array of mononuclear phagocyte (MNP) subsets, including conventional dendritic cells (cDC), monocytes and tissue-resident macrophages (mφ) that collectively play an essential role in mucosal homeostasis, infection and inflammation. In the curr......The intestinal lamina propria (LP) contains a diverse array of mononuclear phagocyte (MNP) subsets, including conventional dendritic cells (cDC), monocytes and tissue-resident macrophages (mφ) that collectively play an essential role in mucosal homeostasis, infection and inflammation....... In the current review we discuss the function of intestinal cDC and monocyte-derived MNP, highlighting how these subsets play several non-redundant roles in the regulation of intestinal immune responses. While much remains to be learnt, recent findings also underline how the various populations of MNP adapt...

  19. Identifying HIV infection in diagnostic histopathology tissue samples--the role of HIV-1 p24 immunohistochemistry in identifying clinically unsuspected HIV infection: a 3-year analysis.

    Science.gov (United States)

    Moonim, Mufaddal T; Alarcon, Lida; Freeman, Janet; Mahadeva, Ula; van der Walt, Jon D; Lucas, Sebastian B

    2010-03-01

    Because of the clinical difficulty in identifying the early stages of human immunodeficiency virus (HIV) infection, the histopathologist often has to consider the diagnosis of HIV in tissue samples from patients with no previous suspicion of HIV infection. The aim was to investigate the practicality and utility of routine HIV-1 p24 immunohistochemistry on tissue samples received at a London histopathology laboratory. Over a 3-year period, HIV-1 p24 was evaluated immunohistochemically on 123 cases. Of these, 37 (30%) showed positive expression of p24 in lesional follicular dendritic cells (FDCs). Of these 37 cases, 11 were not clinically suspected to be HIV+ and had no prior serological evidence of HIV infection. These cases represented lymph node biopsies, tonsillar and nasopharyngeal biopsies and a parotid excision. In addition to expression on FDCs, in 22 cases (60%), p24 also highlighted mononuclear cells and macrophages. p24 was also useful in confirming the presence of HIV in lymphoid tissue in non-lymphoid organs such as the lung, anus, salivary gland and brain. Immunonegativity occurred in occasional known HIV+ cases, probably related to treatment or tissue processing. This study confirms the usefulness of this technique in detecting unsuspected HIV infection in lymphoid and non-lymphoid organs on histopathological material and should be part of routine evaluation of lymph nodes and lymphoid tissue in other organs if morphological or clinical features suggest HIV infection.

  20. Structural changes of tissue samples exposed to low frequency electromagnetic field: A FT-IR absorbance study

    OpenAIRE

    Crupi, V.; Interdonato, S.; Majolino, D.; Mondello, M. R.; Pergolizzi, S.; Venuti, V.

    2004-01-01

    In the present work, we report on a preliminary Fourier Transform Infrared (FT-IR) Absorbance study performed on different kind of rat tissues, such as kidney and heart, exposed to a “non-ionizing” radiation source at low frequency, in the range typical of micro-waves (300 MHz < v < 300 GHz). The data were collected in a wide wavenumber region, from 400 cm−1 to 4000 cm−1. The comparison of the absorbance spectra in the case of the normal tissues with the irradiated ones has shown significant ...

  1. Fit for genomic and proteomic purposes: Sampling the fitness of nucleic acid and protein derivatives from formalin fixed paraffin embedded tissue.

    Directory of Open Access Journals (Sweden)

    Anna Yakovleva

    Full Text Available The demand for nucleic acid and protein derivatives from formalin-fixed paraffin-embedded (FFPE tissue has greatly increased due to advances in extraction and purification methods, making these derivatives available for numerous genomic and proteomic platforms. Previously, DNA, RNA, microRNA (miRNA, or protein derived from FFPE tissue blocks were considered "unfit" for such platforms, as the process of tissue immobilization by FFPE resulted in cross-linked, fragmented, and chemically modified macromolecules. We conducted a systematic examination of nucleic acids and proteins co-extracted from 118 FFPE blocks sampled from the AIDS and Cancer Specimen Resource (ACSR at The George Washington University after stratification by storage duration and the three most common tumor tissue types at the ACSR (adenocarcinoma, squamous cell carcinoma, and papillary carcinoma. DNA, RNA, miRNA, and protein could be co-extracted from 98% of the FFPE blocks sampled, with DNA and miRNA "fit" for diverse genomic purposes including sequencing. While RNA was the most labile of the FFPE derivatives, especially when assessed by RNA integrity number (RIN, it was still "fit" for genomic methods that use smaller sequence lengths, e.g., quantitative PCR. While more than half of the protein derivatives were fit for proteomic purposes, our analyses indicated a significant interaction effect on the absorbance values for proteins derived from FFPE, implying that storage duration may affect protein derivatives differently by tumor tissue type. The mean absorbance value for proteins derived from more recently stored FFPE was greater than protein derived from older FFPE, with the exception of adenocarcinoma tissue. Finally, the fitness of one type of derivative was weakly associated with the fitness of derivatives co-extracted from the same FFPE block. The current study used several novel quality assurance approaches and metrics to show that archival FFPE tissue blocks are a

  2. Application of FTA technology for sampling, recovery and molecular characterization of viral pathogens and virus-derived transgenes from plant tissues

    Directory of Open Access Journals (Sweden)

    Aveling Terry

    2005-05-01

    Full Text Available Abstract Background Plant viral diseases present major constraints to crop production. Effective sampling of the viruses infecting plants is required to facilitate their molecular study and is essential for the development of crop protection and improvement programs. Retaining integrity of viral pathogens within sampled plant tissues is often a limiting factor in this process, most especially when sample sizes are large and when operating in developing counties and regions remote from laboratory facilities. FTA is a paper-based system designed to fix and store nucleic acids directly from fresh tissues pressed into the treated paper. We report here the use of FTA as an effective technology for sampling and retrieval of DNA and RNA viruses from plant tissues and their subsequent molecular analysis. Results DNA and RNA viruses were successfully recovered from leaf tissues of maize, cassava, tomato and tobacco pressed into FTA® Classic Cards. Viral nucleic acids eluted from FTA cards were found to be suitable for diagnostic molecular analysis by PCR-based techniques and restriction analysis, and for cloning and nucleotide sequencing in a manner equivalent to that offered by tradition isolation methods. Efficacy of the technology was demonstrated both from sampled greenhouse-grown plants and from leaf presses taken from crop plants growing in farmer's fields in East Africa. In addition, FTA technology was shown to be suitable for recovery of viral-derived transgene sequences integrated into the plant genome. Conclusion Results demonstrate that FTA is a practical, economical and sensitive method for sampling, storage and retrieval of viral pathogens and plant genomic sequences, when working under controlled conditions and in the field. Application of this technology has the potential to significantly increase ability to bring modern analytical techniques to bear on the viral pathogens infecting crop plants.

  3. Luminal and mucosal-associated intestinal microbiota in patients with diarrhea-predominant irritable bowel syndrome

    Directory of Open Access Journals (Sweden)

    Carroll Ian M

    2010-12-01

    Full Text Available Abstract Background Recent studies have suggested a role for an altered intestinal microbiota in the pathophysiology of irritable bowel syndrome (IBS. However, no consensus has been reached regarding the association between specific enteric bacterial groups and IBS. The aim of this study was to investigate the fecal and mucosal-associated microbiota using two independent techniques in intestinal samples from diarrhea-predominant IBS (D-IBS and healthy controls. Methods Fecal and colonic mucosal biopsy samples were obtained from 10 D-IBS patients and 10 healthy controls. Colonic tissue was collected during a un-sedated un-prepped flexible sigmoidoscopy. Fecal and tissue samples were processed immediately upon collection for culture under aerobic and anaerobic conditions or frozen for further molecular analysis. DNA was extracted from all frozen samples and used to enumerate specific bacterial groups using quantitative real-time PCR (qPCR. Results Culture analysis of intestinal samples demonstrated a significant reduction in the concentration of aerobic bacteria in fecal samples from D-IBS patients when compared to healthy controls (1.4 × 107 vs. 8.4 × 108 CFUs/g feces, P = 0.002. qPCR analysis demonstrated a significant 3.6 fold increase (P = 0.02 in concentrations of fecal Lactobacillus species between D-IBS patients and healthy controls. Conclusions Our culture and molecular data indicate that quantitative differences exist in specific bacterial groups in the microbiota between D-IBS and healthy subjects.

  4. Mammalian intestinal epithelial cells in primary culture: a mini-review.

    Science.gov (United States)

    Kaeffer, Bertrand

    2002-03-01

    Epithelial cells lining the digestive tract represent a highly organized system built up by multipotent stem cells. A process of asymmetric mitosis produces a population of proliferative cells that are rapidly renewed and migrate along the crypt-villus axis, differentiating into functional mature cells before dying and exfoliating into the intestinal lumen. Isolated crypts or epithelial cells retaining high viability can be prepared within a few h after tissue sampling. After cells are cultured in serum-free media, short-term studies (16-48 h) can be conducted for endocrinology, energy metabolism, or programmed cell death. However, long-term primary culture of intestinal cells (up to 10 d) is still difficult despite progress in isolation methodologies and manipulation of the cell microenvironment. The main problem in developing primary culture is the lack of structural markers specific to the stem cell compartment. The design of a microscopic multidimensional analytic system to record the expression profiles of biomarkers all along the living intestinal crypt should improve basic knowledge of the survival and growth of adult crypt stem cells, and the selection of totipotent embryonic stem cells capable of differentiating into intestinal tissues should facilitate studies of the genomic basis of endodermal tissue differentiation.

  5. Collecting and Studying Blood and Tissue Samples From Patients With Locally Recurrent or Metastatic Prostate or Bladder/Urothelial Cancer

    Science.gov (United States)

    2017-12-04

    Healthy Control; Localized Urothelial Carcinoma of the Renal Pelvis and Ureter; Metastatic Malignant Neoplasm in the Bone; Metastatic Malignant Neoplasm in the Soft Tissues; Metastatic Urothelial Carcinoma of the Renal Pelvis and Ureter; Recurrent Bladder Carcinoma; Recurrent Prostate Carcinoma; Recurrent Urothelial Carcinoma of the Renal Pelvis and Ureter; Stage IV Bladder Cancer; Stage IV Bladder Urothelial Carcinoma; Stage IV Prostate Cancer

  6. Community and genomic analysis of the human small intestine microbiota

    NARCIS (Netherlands)

    Bogert, van den B.

    2013-01-01

    Our intestinal tract is densely populated by different microbes, collectively called microbiota, of which the majority are bacteria. Research focusing on the intestinal microbiota often use fecal samples as a representative of the bacteria that inhabit the end of the large intestine.

  7. Farewell to Animal Testing: Innovations on Human Intestinal Microphysiological Systems

    Directory of Open Access Journals (Sweden)

    Tae Hyun Kang

    2016-06-01

    Full Text Available The human intestine is a dynamic organ where the complex host-microbe interactions that orchestrate intestinal homeostasis occur. Major contributing factors associated with intestinal health and diseases include metabolically-active gut microbiota, intestinal epithelium, immune components, and rhythmical bowel movement known as peristalsis. Human intestinal disease models have been developed; however, a considerable number of existing models often fail to reproducibly predict human intestinal pathophysiology in response to biological and chemical perturbations or clinical interventions. Intestinal organoid models have provided promising cytodifferentiation and regeneration, but the lack of luminal flow and physical bowel movements seriously hamper mimicking complex host-microbe crosstalk. Here, we discuss recent advances of human intestinal microphysiological systems, such as the biomimetic human “Gut-on-a-Chip” that can employ key intestinal components, such as villus epithelium, gut microbiota, and immune components under peristalsis-like motions and flow, to reconstitute the transmural 3D lumen-capillary tissue interface. By encompassing cutting-edge tools in microfluidics, tissue engineering, and clinical microbiology, gut-on-a-chip has been leveraged not only to recapitulate organ-level intestinal functions, but also emulate the pathophysiology of intestinal disorders, such as chronic inflammation. Finally, we provide potential perspectives of the next generation microphysiological systems as a personalized platform to validate the efficacy, safety, metabolism, and therapeutic responses of new drug compounds in the preclinical stage.

  8. Two alternative DNA extraction methods to improve the detection of Mycobacterium-tuberculosis-complex members in cattle and red deer tissue samples.

    Science.gov (United States)

    Fell, Shari; Bröckl, Stephanie; Büttner, Mathias; Rettinger, Anna; Zimmermann, Pia; Straubinger, Reinhard K

    2016-09-15

    Bovine tuberculosis (bTB), which is caused by Mycobacterium bovis and M. caprae, is a notifiable animal disease in Germany. Diagnostic procedure is based on a prescribed protocol that is published in the framework of German bTB legislation. In this protocol small sample volumes are used for DNA extraction followed by real-time PCR analyses. As mycobacteria tend to concentrate in granuloma and the infected tissue in early stages of infection does not necessarily show any visible lesions, it is likely that DNA extraction from only small tissue samples (20-40 mg) of a randomly chosen spot from the organ and following PCR testing may result in false negative results. In this study two DNA extraction methods were developed to process larger sample volumes to increase the detection sensitivity of mycobacterial DNA in animal tissue. The first extraction method is based on magnetic capture, in which specific capture oligonucleotides were utilized. These nucleotides are linked to magnetic particles and capture Mycobacterium-tuberculosis-complex (MTC) DNA released from 10 to 15 g of tissue material. In a second approach remaining sediments from the magnetic capture protocol were further processed with a less complex extraction protocol that can be used in daily routine diagnostics. A total number of 100 tissue samples from 34 cattle (n = 74) and 18 red deer (n = 26) were analyzed with the developed protocols and results were compared to the prescribed protocol. All three extraction methods yield reliable results by the real-time PCR analysis. The use of larger sample volume led to a sensitivity increase of DNA detection which was shown by the decrease of Ct-values. Furthermore five samples which were tested negative or questionable by the official extraction protocol were detected positive by real time PCR when the alternative extraction methods were used. By calculating the kappa index, the three extraction protocols resulted in a moderate (0.52; protocol 1 vs 3

  9. Concentration levels of zearalenone and its metabolites in urine, muscle tissue, and liver samples of pigs fed with mycotoxin-contaminated oats.

    Science.gov (United States)

    Zöllner, Peter; Jodlbauer, Justus; Kleinova, Martina; Kahlbacher, Hermann; Kuhn, Thomas; Hochsteiner, Werner; Lindner, Wolfgang

    2002-04-24

    The content of zearalenone and its metabolites in urine and tissue samples from pigs fed zearalenone-contaminated oats was established by analytical methods combining solid-phase extraction cleanup of the samples with highly selective liquid chromatography-mass spectrometry (LC-MS)/MS detection. Investigation of the urine samples revealed that approximately 60% of zearalenone was transformed in vivo to alpha-zearalenol and its epimer beta-zearalenol in a mean ratio of 3:1. Zeranol and taleranol as further metabolites could only be detected in trace amounts. Zearalanone was identified at considerable concentrations, though only in a couple of samples. In contrast, liver samples contained predominantly alpha-zearalenol, and to a minor extent beta-zearalenol and zearalenone, with a mean ratio of alpha-/beta-zearalenol of 2.5:1, while zeranol, taleranol, or zearalanone could not be identified in any of the investigated samples. The degree of glucoronidation was established for zearalenone as 27% in urine and 62% in liver; for alpha-zearalenol as 88% in urine and 77% in liver; and for beta-zearalenol as 94% in urine and 29% in liver. Analyses of muscle tissue revealed relatively high amounts of nonglucuronidated zeranol and alpha-zearalenol together with traces of taleranol and zearalenone, indicating that the metabolism of zearalenone and its metabolites is not restricted to hepatic and gastrointestinal metabolic pathways.

  10. Intestinal Epithelial Sirtuin 1 Regulates Intestinal Inflammation During Aging in Mice by Altering the Intestinal Microbiota.

    Science.gov (United States)

    Wellman, Alicia S; Metukuri, Mallikarjuna R; Kazgan, Nevzat; Xu, Xiaojiang; Xu, Qing; Ren, Natalie S X; Czopik, Agnieszka; Shanahan, Michael T; Kang, Ashley; Chen, Willa; Azcarate-Peril, M Andrea; Gulati, Ajay S; Fargo, David C; Guarente, Leonard; Li, Xiaoling

    2017-09-01

    Intestinal epithelial homeostasis is maintained by complex interactions among epithelial cells, commensal gut microorganisms, and immune cells. Disruption of this homeostasis is associated with disorders such as inflammatory bowel disease (IBD), but the mechanisms of this process are not clear. We investigated how Sirtuin 1 (SIRT1), a conserved mammalian NAD + -dependent protein deacetylase, senses environmental stress to alter intestinal integrity. We performed studies of mice with disruption of Sirt1 specifically in the intestinal epithelium (SIRT1 iKO, villin-Cre+, Sirt1 flox/flox mice) and control mice (villin-Cre-, Sirt1 flox/flox ) on a C57BL/6 background. Acute colitis was induced in some mice by addition of 2.5% dextran sodium sulfate to drinking water for 5-9 consecutive days. Some mice were given antibiotics via their drinking water for 4 weeks to deplete their microbiota. Some mice were fed with a cholestyramine-containing diet for 7 days to sequester their bile acids. Feces were collected and proportions of microbiota were analyzed by 16S rRNA amplicon sequencing and quantitative PCR. Intestines were collected from mice and gene expression profiles were compared by microarray and quantitative PCR analyses. We compared levels of specific mRNAs between colon tissues from age-matched patients with ulcerative colitis (n=10) vs without IBD (n=8, controls). Mice with intestinal deletion of SIRT1 (SIRT1 iKO) had abnormal activation of Paneth cells starting at the age of 5-8 months, with increased activation of NF-κB, stress pathways, and spontaneous inflammation at 22-24 months of age, compared with control mice. SIRT1 iKO mice also had altered fecal microbiota starting at 4-6 months of age compared with control mice, in part because of altered bile acid metabolism. Moreover, SIRT1 iKO mice with defective gut microbiota developed more severe colitis than control mice. Intestinal tissues from patients with ulcerative colitis expressed significantly lower

  11. Predicting tissue outcome from acute stroke magnetic resonance imaging: improving model performance by optimal sampling of training data.

    Science.gov (United States)

    Jonsdottir, Kristjana Yr; Østergaard, Leif; Mouridsen, Kim

    2009-09-01

    It has been hypothesized that algorithms predicting the final outcome in acute ischemic stroke may provide future tools for identifying salvageable tissue and hence guide individualized therapy. We developed means of quantifying predictive model performance to identify model training strategies that optimize performance and reduce bias in predicted lesion volumes. We optimized predictive performance based on the area under the receiver operating curve for logistic regression and used simulated data to illustrate the effect of an unbalanced (unequal number of infarcting and surviving voxels) training set on predicted infarct risk. We then tested the performance and optimality of models based on perfusion-weighted, diffusion-weighted, and structural MRI modalities by changing the proportion of mismatch voxels in balanced training material. Predictive performance (area under the receiver operating curve) based on all brain voxels is excessively optimistic and lacks sensitivity in performance in mismatch tissue. The ratio of infarcting and noninfarcting voxels used for training predictive algorithms significantly biases tissue infarct risk estimates. Optimal training strategy is obtained using a balanced training set. We show that 60% of noninfarcted voxels consists of mismatch voxels in an optimal balanced training set for the patient data presented. An equal number of infarcting and noninfarcting voxels should be used when training predictive models. The choice of test and training sets critically affects predictive model performance and should be closely evaluated before comparisons across patient cohorts.

  12. LINE-1 hypomethylation in blood and tissue samples as an epigenetic marker for cancer risk: a systematic review and meta-analysis.

    Directory of Open Access Journals (Sweden)

    Martina Barchitta

    Full Text Available A systematic review and a meta-analysis were carried out in order to summarize the current published studies and to evaluate LINE-1 hypomethylation in blood and other tissues as an epigenetic marker for cancer risk.A systematic literature search in the Medline database, using PubMed, was conducted for epidemiological studies, published before March 2014. The random-effects model was used to estimate weighted mean differences (MDs with 95% Confidence Intervals (CIs. Furthermore, subgroup analyses were conducted by sample type (tissue or blood samples, cancer types, and by assays used to measure global DNA methylation levels. The Cochrane software package Review Manager 5.2 was used.A total of 19 unique articles on 6107 samples (2554 from cancer patients and 3553 control samples were included in the meta-analysis. LINE-1 methylation levels were significantly lower in cancer patients than in controls (MD: -6.40, 95% CI: -7.71, -5.09; p<0.001. The significant difference in methylation levels was confirmed in tissue samples (MD -7.55; 95% CI: -9.14, -65.95; p<0.001, but not in blood samples (MD: -0.26, 95% CI: -0.69, 0.17; p = 0.23. LINE-1 methylation levels were significantly lower in colorectal and gastric cancer patients than in controls (MD: -8.33; 95% CI: -10.56, -6.10; p<0.001 and MD: -5.75; 95% CI: -7.75, -3.74; p<0.001 whereas, no significant difference was observed for hepatocellular cancer.The present meta-analysis adds new evidence to the growing literature on the role of LINE-1 hypomethylation in human cancer and demonstrates that LINE-1 methylation levels were significantly lower in cancer patients than in control samples, especially in certain cancer types. This result was confirmed in tissue samples, both fresh/frozen or FFPE specimens, but not in blood. Further studies are needed to better clarify the role of LINE-1 methylation in specific subgroups, considering both cancer and sample type, and the methods of measurement.

  13. Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples

    LENUS (Irish Health Repository)

    Mee, Blanaid C.

    2011-12-29

    The Saint James\\'s Hospital Biobank was established in 2008, to develop a high-quality breast tissue BioResource, as a part of the breast cancer clinical care pathway. The aims of this work were: (1) to ascertain the quality of RNA, DNA, and protein in biobanked carcinomas and normal breast tissues, (2) to assess the efficacy of AllPrep® (Qiagen) in isolating RNA, DNA, and protein simultaneously, (3) to compare AllPrep with RNEasy® and QIAamp® (both Qiagen), and (4) to examine the effectiveness of Allprotect® (Qiagen), a new tissue stabilization medium in preserving DNA, RNA, and proteins. One hundred eleven frozen samples of carcinoma and normal breast tissue were analyzed. Tumor and normal tissue morphology were confirmed by frozen sections. Tissue type, tissue treatment (Allprotect vs. no Allprotect), extraction kit, and nucleic acid quantification were analyzed by utilizing a 4 factorial design (SPSS PASW 18 Statistics Software®). QIAamp (DNA isolation), AllPrep (DNA, RNA, and Protein isolation), and RNeasy (RNA isolation) kits were assessed and compared. Mean DNA yield and A260\\/280 values using QIAamp were 33.2 ng\\/μL and 1.86, respectively, and using AllPrep were 23.2 ng\\/μL and 1.94. Mean RNA yield and RNA Integrity Number (RIN) values with RNeasy were 73.4 ng\\/μL and 8.16, respectively, and with AllPrep were 74.8 ng\\/μL and 7.92. Allprotect-treated tissues produced higher RIN values of borderline significance (P=0.055). No discernible loss of RNA stability was detected after 6 h incubation of stabilized or nonstabilized tissues at room temperature or 4°C or in 9 freeze-thaw cycles. Allprotect requires further detailed evaluation, but we consider AllPrep to be an excellent option for the simultaneous extraction of RNA, DNA, and protein from tumor and normal breast tissues. The essential presampling procedures that maintain the diagnostic integrity of pathology specimens do not appear to compromise the quality of molecular isolates.

  14. Intestinal lineage commitment of embryonic stem cells.

    Science.gov (United States)

    Cao, Li; Gibson, Jason D; Miyamoto, Shingo; Sail, Vibhavari; Verma, Rajeev; Rosenberg, Daniel W; Nelson, Craig E; Giardina, Charles

    2011-01-01

    Generating lineage-committed intestinal stem cells from embryonic stem cells (ESCs) could provide a tractable experimental system for understanding intestinal differentiation pathways and may ultimately provide cells for regenerating damaged intestinal tissue. We tested a two-step differentiation procedure in which ESCs were first cultured with activin A to favor formation of definitive endoderm, and then treated with fibroblast-conditioned medium with or without Wnt3A. The definitive endoderm expressed a number of genes associated with gut-tube development through mouse embryonic day 8.5 (Sox17, Foxa2, and Gata4 expressed and Id2 silent). The intestinal stem cell marker Lgr5 gene was also activated in the endodermal cells, whereas the Msi1, Ephb2, and Dcamkl1 intestinal stem cell markers were not. Exposure of the endoderm to fibroblast-conditioned medium with Wnt3A resulted in the activation of Id2, the remaining intestinal stem cell markers and the later gut markers Cdx2, Fabp2, and Muc2. Interestingly, genes associated with distal gut-associated mesoderm (Foxf2, Hlx, and Hoxd8) were also simulated by Wnt3A. The two-step differentiation protocol generated gut bodies with crypt-like structures that included regions of Lgr5-expressing proliferating cells and regions of cell differentiation. These gut bodies also had a smooth muscle component and some underwent peristaltic movement. The ability of the definitive endoderm to differentiate into intestinal epithelium was supported by the vivo engraftment of these cells into mouse colonic mucosa. These findings demonstrate that definitive endoderm derived from ESCs can carry out intestinal cell differentiation pathways and may provide cells to restore damaged intestinal tissue. Copyright © 2010 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  15. Resected Human Colonic Tissue: New Model for Characterizing Adhesion of Lactic Acid Bacteria

    Science.gov (United States)

    Ouwehand, Arthur C.; Salminen, Seppo; Tölkkö, Satu; Roberts, Peter; Ovaska, Jari; Salminen, Eeva

    2002-01-01

    Adhesion to the intestinal mucosa is one of the main selection criteria for probiotic strains. The adhesion of commonly used probiotic strains to human intestinal tissue pieces and mucus was assessed. The strains tested adhered to the intestinal tissue at low levels and adhered to the intestinal mucus at higher levels. PMID:11777852

  16. Intestinal epithelial apoptosis initiates gross bowel necrosis in an experimental rat model of neonatal necrotizing enterocolitis.

    Science.gov (United States)

    Jilling, Tamas; Lu, Jing; Jackson, Michele; Caplan, Michael S

    2004-04-01

    The histopathology of necrotizing enterocolitis (NEC) is characterized by destruction of the mucosal layer in initial stages and by transmural necrosis of the intestinal wall in advanced stages of the disease. To test the hypothesis that enhanced epithelial apoptosis is an initial event underlying the gross histologic changes, we analyzed epithelial apoptosis and tissue morphology in an animal model of NEC and evaluated the effect of caspase inhibition on the incidence of experimental NEC in this model. Apoptosis was analyzed with terminal deoxynucleotidyltransferase-mediated dUTP-FITC nick end labeling (TUNEL) staining in intestinal sections and by measuring caspase 3 activity from intestinal lysates of neonatal rats subjected to formula feeding and cold/asphyxia stress (FFCAS) and from mother-fed (MF) controls. Morphologic evaluation was based on hematoxylin and eosin staining of intestinal sections. FFCAS resulted in histologic changes consistent with NEC, which were absent from MF animals. FFCAS was also associated with a significantly increased rate of nuclear DNA fragmentation in the small intestinal epithelium compared with MF. Elevated tissue caspase 3 activity confirmed the presence of apoptosis in samples with increased DNA fragmentation. Analysis of the coincidence of morphologic damage and apoptosis in corresponding tissue sections indicated that apoptosis precedes gross morphologic changes in this model. Furthermore, supplementation of formula with 8 boc-aspartyl(OMe)-fluoromethylketone, a pan-caspase inhibitor, significantly reduced the incidences of apoptosis and experimental NEC. These findings indicate that in neonatal rats FFCAS induces epithelial apoptosis that serves as an underlying cause for subsequent gross tissue necrosis.

  17. Peptidases Compartmentalized to the Ascaris suum Intestinal Lumen and Apical Intestinal Membrane

    Science.gov (United States)

    Rosa, Bruce A.

    2015-01-01

    The nematode intestine is a tissue of interest for developing new methods of therapy and control of parasitic nematodes. However, biological details of intestinal cell functions remain obscure, as do the proteins and molecular functions located on the apical intestinal membrane (AIM), and within the intestinal lumen (IL) of nematodes. Accordingly, methods were developed to gain a comprehensive identification of peptidases that function in the intestinal tract of adult female Ascaris suum. Peptidase activity was detected in multiple fractions of the A. suum intestine under pH conditions ranging from 5.0 to 8.0. Peptidase class inhibitors were used to characterize these activities. The fractions included whole lysates, membrane enriched fractions, and physiological- and 4 molar urea-perfusates of the intestinal lumen. Concanavalin A (ConA) was confirmed to bind to the AIM, and intestinal proteins affinity isolated on ConA-beads were compared to proteins from membrane and perfusate fractions by mass spectrometry. Twenty-nine predicted peptidases were identified including aspartic, cysteine, and serine peptidases, and an unexpectedly high number (16) of metallopeptidases. Many of these proteins co-localized to multiple fractions, providing independent support for localization to specific intestinal compartments, including the IL and AIM. This unique perfusion model produced the most comprehensive view of likely digestive peptidases that function in these intestinal compartments of A. suum, or any nematode. This model offers a means to directly determine functions of these proteins in the A. suum intestine and, more generally, deduce the wide array functions that exist in these cellular compartments of the nematode intestine. PMID:25569475

  18. A New Method for Endoscopic Sampling of Submucosal Tissue in the Gastrointestinal Tract: A Comparison of the Biopsy Forceps and a New Drill Instrument.

    Science.gov (United States)

    Walther, Charles; Jeremiasen, Martin; Rissler, Pehr; Johansson, Jan L M; Larsson, Marie S; Walther, Bruno S C S

    2016-12-01

    Background Sampling of submucosal lesions in the gastrointestinal tract through a flexible endoscope is a well-recognized clinical problem. One technique often used is endoscopic ultrasound-guided fine-needle aspiration, but it does not provide solid tissue biopsies with preserved architecture for histopathological evaluation. To obtain solid tissue biopsies from submucosal lesions, we have constructed a new endoscopic biopsy tool and compared it in a crossover study with the standard double cupped forceps. Methods Ten patients with endoscopically verified submucosal lesions were sampled. The endoscopist selected the position for the biopsies and used the instrument selected by randomization. After a biopsy was harvested, the endoscopist chose the next site for a biopsy and again used the instrument picked by randomization. A total of 6 biopsies, 3 with the forceps and 3 with the drill instrument, were collected in every patient. Results The drill instrument resulted in larger total size biopsies (mm 2 ; Mann-Whitney U test, P = .048) and larger submucosal part (%) of the biopsies (Mann-Whitney U test, P = .003) than the forceps. Two patients were observed because of chest pain and suspicion of bleeding in 24 hours. No therapeutic measures were necessary to be taken. Conclusion The new drill instrument for flexible endoscopy can safely deliver submucosal tissue samples from submucosal lesions in the upper gastrointestinal tract. © The Author(s) 2016.

  19. Pre-Analytical Considerations for Successful Next-Generation Sequencing (NGS: Challenges and Opportunities for Formalin-Fixed and Paraffin-Embedded Tumor Tissue (FFPE Samples

    Directory of Open Access Journals (Sweden)

    Gladys Arreaza

    2016-09-01

    Full Text Available In cancer drug discovery, it is important to investigate the genetic determinants of response or resistance to cancer therapy as well as factors that contribute to adverse events in the course of clinical trials. Despite the emergence of new technologies and the ability to measure more diverse analytes (e.g., circulating tumor cell (CTC, circulating tumor DNA (ctDNA, etc., tumor tissue is still the most common and reliable source for biomarker investigation. Because of its worldwide use and ability to preserve samples for many decades at ambient temperature, formalin-fixed, paraffin-embedded tumor tissue (FFPE is likely to be the preferred choice for tissue preservation in clinical practice for the foreseeable future. Multiple analyses are routinely performed on the same FFPE samples (such as Immunohistochemistry (IHC, in situ hybridization, RNAseq, DNAseq, TILseq, Methyl-Seq, etc.. Thus, specimen prioritization and optimization of the isolation of analytes is critical to ensure successful completion of each assay. FFPE is notorious for producing suboptimal DNA quality and low DNA yield. However, commercial vendors tend to request higher DNA sample mass than what is actually required for downstream assays, which restricts the breadth of biomarker work that can be performed. We evaluated multiple genomics service laboratories to assess the current state of NGS pre-analytical processing of FFPE. Significant differences in pre-analytical capabilities were observed. Key aspects are highlighted and recommendations are made to improve the current practice in translational research.

  20. [Effect of perioperative intestinal probiotics on intestinal flora and immune function in patients with colorectal cancer].

    Science.gov (United States)

    Zhu, Dajian; Chen, Xiaowu; Wu, Jinhao; Ju, Yongle; Feng, Jing; Lu, Guangsheng; Ouyang, Manzhao; Ren, Baojun; Li, Yong

    2012-08-01

    To investigate the effect of perioperative application of intestinal probiotics to substitute oral intestinal antimicrobial agents on intestinal flora and immune function in surgical patients with colorectal cancer. Sixty patients with colorectal cancer undergoing elective laparoscopic radical surgery were randomized to receive preoperative bowel preparation using oral intestinal antimicrobial agents (n=20) or using oral intestinal probiotics (Jinshuangqi Tablets, 2.0 g, 3 times daily) since the fifth day before the operation and at 24 h after the operation for 7 consecutive days. Upon admission and 7 days after the operation, fecal samples and fasting peripheral venous blood were collected from the patients to examine the intestinal flora and serum levels of interleukin-2 (IL-2), IgA, IgG, and IgM, NK cell activity, T lymphocytes subsets CD3(+), CD4(+), CD8(+) and CD4(+)/CD8(+) ratio. At 7 days after the operation, the patients receiving probiotics showed significantly increased counts of intestinal Bifidobacterium, Lactobacillus, and Enterococcus (Pprobiotics group compared with those in patients with conventional intestinal preparation (Pprobiotics to replace preoperative oral intestinal antimicrobial agents can effectively correct intestinal flora imbalance and improve the immune function of surgical patients with colorectal cancer.

  1. A new method to measure intestinal secretion using fluorescein isothiocyanate-inulin in small bowel of rats.

    Science.gov (United States)

    Munoz-Abraham, Armando Salim; Judeeba, Sami; Alkukhun, Abedalrazaq; Alfadda, Tariq; Patron-Lozano, Roger; Rodriguez-Davalos, Manuel I; Geibel, John P

    2015-08-01

    Small intestine ischemia can be seen in various conditions such as intestinal transplantation. To further understand the pathologic disruption in ischemia-reperfusion injury, we have developed a method to measure fluid changes in the intestinal lumen of rats. Two 10-cm rat intestine segments were procured, connected to the terminal apertures of a perfusion device, and continuously infused with 3 mL of HEPES solution (control solution) containing 50 μM of fluorescein isothiocyanate (FITC)-inulin. The perfusion device consists of concentric chambers that contain the perfused bowel segments, which are maintained at 37°C via H₂O bath. The individual chamber has four apertures as follows: two fill and/or drain the surrounding HEPES solution on the blood side of the tissue. The others provide flow of HEPES solution containing FITC-inulin through the lumens. The experimental intestine was infused with the same solution with 100 μM of Forskolin. A pump continuously circulated solutions at 6 mL/min. Samples were collected at 15-min intervals until 150 min and were measured by the nanoflourospectrometer. A mean of 6-μM decrease in the FITC-inulin concentration in the Forskolin-treated experimental intestine was observed in comparison with that in the control intestine. The FITC-inulin count dilution in the experimental intestine is a result of an increase of fluid secretion produced by the effect of Forskolin, with P values inulin to allow real-time determinations of fluid and/or electrolyte movement along the small intestine. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Effects of sodium hydrosulfide on intestinal mucosal injury in a rat model of cardiac arrest and cardiopulmonary resuscitation.

    Science.gov (United States)

    Pan, Hao; Chen, Di; Liu, Beibei; Xie, Xuemeng; Zhang, Jincheng; Yang, Guangtian

    2013-07-19

    Cardiac arrest and cardiopulmonary resuscitation (CPR) can lead to intestinal ischemia/reperfusion (I/R) injury. Increasing studies have indicated that hydrogen sulfide (H2S) is in favor of a variety of tissue I/R injury. The purpose of this study was to explore whether sodium hydrosulfide (NaHS), a H2S donor, can protect intestinal mucosa after CPR and its potential mechanisms. Male Sprague-Dawley rats were subjected to 6min cardiac arrest induced by transcutaneous electrical epicardium stimulation and then resuscitated successfully. A bolus of either NaHS (0.5mg/kg) or placebo (NaCl 0.9%) was blindly injected 1min before the start of CPR intravenously, followed by a continuous injection of NaHS (2mg/kg/h) or placebo for 3h. Intestinal and plasma samples were collected for assessments 24h after CPR. We found that NaHS can markedly alleviate cardiac arrest induced intestinal mucosal injury. Rats treated with NaHS showed a lower malondialdehyde content, higher superoxide dismutase activity and glutathione content in intestine after CPR. Increased intestinal myeloperoxidase activity was significantly decreased by NaHS after CPR. Moreover, a reduced intestinal apoptotic cells after CPR were evident when pretreated with NaHS. Further studies indicated that NaHS enhances the expression of hypoxia-inducible factor-1α (HIF-1α) in intestine after CPR. Our data demonstrated that NaHS treatment before CPR induces intestinal mucosal protection 24h post-resuscitation. The protective effects may be through oxidative stress reduction, inflammation alleviation, apoptosis inhibition and HIF-1α activation. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. A Revised Model for Dosimetry in the Human Small Intestine

    Energy Technology Data Exchange (ETDEWEB)

    John Poston; Nasir U. Bhuiyan; R. Alex Redd; Neil Parham; Jennifer Watson

    2005-02-28

    A new model for an adult human gastrointestinal tract (GIT) has been developed for use in internal dose estimations to the wall of the GIT and to the other organs and tissues of the body from radionuclides deposited in the lumenal contents of the five sections of the GIT. These sections were the esophasgus, stomach, small intestine, upper large intestine, and the lower large intestine. The wall of each section was separated from its lumenal contents.

  4. A Revised Model for Dosimetry in the Human Small Intestine

    International Nuclear Information System (INIS)

    John Poston; Bhuiyan, Nasir U.; Redd, R. Alex; Neil Parham; Jennifer Watson

    2005-01-01

    A new model for an adult human gastrointestinal tract (GIT) has been developed for use in internal dose estimations to the wall of the GIT and to the other organs and tissues of the body from radionuclides deposited in the lumenal contents of the five sections of the GIT. These sections were the esophasgus, stomach, small intestine, upper large intestine, and the lower large intestine. The wall of each section was separated from its lumenal contents

  5. Intestinal microbiome landscaping

    NARCIS (Netherlands)

    Shetty, Sudarshan A.; Hugenholtz, Floor; Lahti, Leo; Smidt, Hauke; Vos, de Willem M.

    2017-01-01

    High individuality, large complexity and limited understanding of the mechanisms underlying human intestinal microbiome function remain the major challenges for designing beneficial modulation strategies. Exemplified by the analysis of intestinal bacteria in a thousand Western adults, we discuss

  6. Nested-PCR for the detection of Mycoplasma hyopneumoniae in bronchial alveolar swabs, frozen tissues and formalin-fixed paraffin-embedded swine lung samples: comparative evaluation with immunohistochemical findings and histological features

    Directory of Open Access Journals (Sweden)

    Paula R. Almeida

    2012-08-01

    Full Text Available The diagnosis of Mycoplasma hyopneumoniae infection is often performed through histopathology, immunohistochemistry (IHC and polymerase chain reaction (PCR or a combination of these techniques. PCR can be performed on samples using several conservation methods, including swabs, frozen tissue or formalin-fixed and paraffin-embedded (FFPE tissue. However, the formalin fixation process often inhibits DNA amplification. To evaluate whether M. hyopneumoniae DNA could be recovered from FFPE tissues, 15 lungs with cranioventral consolidation lesions were collected in a slaughterhouse from swine bred in herds with respiratory disease. Bronchial swabs and fresh lung tissue were collected, and a fragment of the corresponding lung section was placed in neutral buffered formalin for 48 hours. A PCR assay was performed to compare FFPE tissue samples with samples that were only refrigerated (bronchial swabs or frozen (tissue pieces. M. hyopneumoniae was detected by PCR in all 15 samples of the swab and frozen tissue, while it was detected in only 11 of the 15 FFPE samples. Histological features of M. hyopneumoniae infection were presented in 11 cases and 7 of these samples stained positive in IHC. Concordance between the histological features and detection results was observed in 13 of the FFPE tissue samples. PCR was the most sensitive technique. Comparison of different sample conservation methods indicated that it is possible to detect M. hyopneumoniae from FFPE tissue. It is important to conduct further research using archived material because the efficiency of PCR could be compromised under these conditions.

  7. Small Intestine Cancer Treatment

    Science.gov (United States)

    ... all of an organ that contains cancer. The resection may include the small intestine and nearby organs (if the cancer has spread). The doctor may remove the section of the small intestine that contains cancer and perform an anastomosis (joining the cut ends of the intestine together). ...

  8. An assessment of computational methods for estimating purity and clonality using genomic data derived from heterogeneous tumor tissue samples.

    Science.gov (United States)

    Yadav, Vinod Kumar; De, Subhajyoti

    2015-03-01

    Solid tumor samples typically contain multiple distinct clonal populations of cancer cells, and also stromal and immune cell contamination. A majority of the cancer genomics and transcriptomics studies do not explicitly consider genetic heterogeneity and impurity, and draw inferences based on mixed populations of cells. Deconvolution of genomic data from heterogeneous samples provides a powerful tool to address this limitation. We discuss several computational tools, which enable deconvolution of genomic and transcriptomic data from heterogeneous samples. We also performed a systematic comparative assessment of these tools. If properly used, these tools have potentials to complement single-cell genomics and immunoFISH analyses, and provide novel insights into tumor heterogeneity. © The Author 2014. Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

  9. Relative sensitivity of conventional and real-time PCR assays for detection of SFG Rickettsia in blood and tissue samples from laboratory animals.

    Science.gov (United States)

    Zemtsova, Galina E; Montgomery, Merrill; Levin, Michael L

    2015-01-01

    Studies on the natural transmission cycles of zoonotic pathogens and the reservoir competence of vertebrate hosts require methods for reliable diagnosis of infection in wild and laboratory animals. Several PCR-based applications have been developed for detection of infections caused by Spotted Fever group Rickettsia spp. in a variety of animal tissues. These assays are being widely used by researchers, but they differ in their sensitivity and reliability. We compared the sensitivity of five previously published conventional PCR assays and one SYBR green-based real-time PCR assay for the detection of rickettsial DNA in blood and tissue samples from Rickettsia- infected laboratory animals (n = 87). The real-time PCR, which detected rickettsial DNA in 37.9% of samples, was the most sensitive. The next best were the semi-nested ompA assay and rpoB conventional PCR, which detected as positive 18.4% and 14.9% samples respectively. Conventional assays targeting ompB, gltA and hrtA genes have been the least sensitive. Therefore, we recommend the SYBR green-based real-time PCR as a tool for the detection of rickettsial DNA in animal samples due to its higher sensitivity when compared to more traditional assays.

  10. Genetic Characterization of Echinococcus granulosus from a Large Number of Formalin-Fixed, Paraffin-Embedded Tissue Samples of Human Isolates in Iran

    Science.gov (United States)

    Rostami, Sima; Torbaghan, Shams Shariat; Dabiri, Shahriar; Babaei, Zahra; Mohammadi, Mohammad Ali; Sharbatkhori, Mitra; Harandi, Majid Fasihi

    2015-01-01

    Cystic echinococcosis (CE), caused by the larval stage of Echinococcus granulosus, presents an important medical and veterinary problem globally, including that in Iran. Different genotypes of E. granulosus have been reported from human isolates worldwide. This study identifies the genotype of the parasite responsible for human hydatidosis in three provinces of Iran using formalin-fixed paraffin-embedded tissue samples. In this study, 200 formalin-fixed paraffin-embedded tissue samples from human CE cases were collected from Alborz, Tehran, and Kerman provinces. Polymerase chain reaction amplification and sequencing of the partial mitochondrial cytochrome c oxidase subunit 1 gene were performed for genetic characterization of the samples. Phylogenetic analysis of the isolates from this study and reference sequences of different genotypes was done using a maximum likelihood method. In total, 54.4%, 0.8%, 1%, and 40.8% of the samples were identified as the G1, G2, G3, and G6 genotypes, respectively. The findings of the current study confirm the G1 genotype (sheep strain) to be the most prevalent genotype involved in human CE cases in Iran and indicates the high prevalence of the G6 genotype with a high infectivity for humans. Furthermore, this study illustrates the first documented human CE case in Iran infected with the G2 genotype. PMID:25535316

  11. Advancing Intestinal Organoid Technology Toward Regenerative Medicine

    Directory of Open Access Journals (Sweden)

    Tetsuya Nakamura

    2018-01-01

    Full Text Available With the emergence of technologies to culture intestinal epithelial cells in vitro as various forms of intestinal organoids, there is growing interest in using such cultured intestinal cells as a source for tissue engineering and regenerative medicine. One such approach would be to combine the organoid technology with methodologies to engineer the culture environment, particularly the three-dimensional scaffold materials, to generate intestines that exquisitely recapitulate their original structures and functions. Another approach to use organoids for regenerative medicine would be to urge them to mature into functional intestines by implanting them into hosts. This process includes the tissue-engineered small intestine that uses synthetic scaffolds for tissue regeneration and direct organoid transplantation that takes advantage of submucosal tissues in the native intestines as a scaffold. Further study in these subfields could lead to the development of therapeutic options to use different types of organoids with various cell types in regenerative medicine for intestinal diseases in humans.

  12. Stem cell self-renewal in intestinal crypt

    NARCIS (Netherlands)

    Simons, B.D.; Clevers, H.

    2011-01-01

    As a rapidly cycling tissue capable of fast repair and regeneration, the intestinal epithelium has emerged as a favored model system to explore the principles of adult stem cell biology. However, until recently, the identity and characteristics of the stem cell population in both the small intestine

  13. Wnt, stem cells and cancer in the intestine.

    NARCIS (Netherlands)

    Pinto, D.; Clevers, J.C.

    2005-01-01

    The intestinal epithelium is a self-renewing tissue which represents a unique model for studying interconnected cellular processes such as proliferation, differentiation, cell migration and carcinogenesis. Although the stem cells of the intestine have not yet been physically characterized or

  14. Automated Liquid Microjunction Surface Sampling-HPLC-MS/MS Analysis of Drugs and Metabolites in Whole-Body Thin Tissue Sections

    Energy Technology Data Exchange (ETDEWEB)

    Kertesz, Vilmos [ORNL; Van Berkel, Gary J [ORNL

    2013-01-01

    A fully automated liquid extraction-based surface sampling system utilizing a commercially available autosampler coupled to high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) detection is reported. Discrete spots selected for droplet-based sampling and automated sample queue generation for both the autosampler and MS were enabled by using in-house developed software. In addition, co-registration of spatially resolved sampling position and HPLC-MS information to generate heatmaps of compounds monitored for subsequent data analysis was also available in the software. The system was evaluated with whole-body thin tissue sections from propranolol dosed rat. The hands-free operation of the system was demonstrated by creating heatmaps of the parent drug and its hydroxypropranolol glucuronide metabolites with 1 mm resolution in the areas of interest. The sample throughput was approximately 5 min/sample defined by the time needed for chromatographic separation. The spatial distributions of both the drug and its metabolites were consistent with previous studies employing other liquid extraction-based surface sampling methodologies.

  15. Increased intestinal marker absorption due to regional permeability changes and decreased intestinal transit during sepsis in the rat

    International Nuclear Information System (INIS)

    Wang, Q.; Pantzar, N.; Jeppson, B.; Westroem, B.R.; Karlsson, B.W.

    1994-01-01

    The intestinal barrier properties are impaired during inflammation and sepsis, but the mechanisms behind this are unknown and were therefore investigated during experimental sepsis in rats. The different-sized intestinal absorption markers 51 Cr-labeled ethylenediaminetetraacetic acid (EDTA) and ovalbumin were gavaged to rats made septic by intra-abdominal bacterial implantation and to sham-operated rats. Regional tissue permeability was measured in diffusion chambers, and intestinal transit was evaluated by intestinal accumulation of gavaged 51 Cr-EDTA. In comparison with the sham-operated rats, septic rats had higher 51 Cr-EDTA levels in blood and urine and showed a prolonged intestinal transit. Septic rats also had a lower tissue permeability to both markers in the small intestines but higher permeability to ovalbumin in the colon. Rats receiving morphine to decrease intestinal motility showed similar changes, with a decreased intestinal transit and increased marker absorption. Thr results suggest that the increased intestinal absorption during sepsis was due to regional permeability changes and prolonged intestinal transit. 38 refs., 4 figs., 2 tabs

  16. Intestinal radiation syndrome: sepsis and endotoxin

    International Nuclear Information System (INIS)

    Geraci, J.P.; Jackson, K.L.; Mariano, M.S.

    1985-01-01

    Rats were whole-body irradiated with 8-MeV cyclotron-produced neutrons and 137 Cs γ rays to study the role of enteric bacteria and endotoxin in the intestinal radiation syndrome. Decrease in intestinal weight was used as an index of radiation-induced breakdown of the mucosa. Neutron and γ-ray doses that were sublethal for intestinal death resulted in a dose-dependent decrease in intestinal weight, reaching minimal values 2 to 3 days after exposure, followed by recovery within 5 days after irradiation. Neutron and photon doses that caused intestinal death resulted in greater mucosal breakdown with little or no evidence of mucosal recovery. The presence of fluid in the intestine and diarrhea, but not bacteremia or endotoxemia, were related to mucosal breakdown and recovery. Neither sepsis nor endotoxin could be detected in liver samples taken at autopsy from animals which died a short time earlier from intestinal injury. These results suggest that overt sepsis and endotoxemia do not play a significant role in the intestinal radiation syndrome

  17. [Intestinal parasitic diseases in children].

    Science.gov (United States)

    Mare, Anca; Man, A; Toma, Felicia; Székely, Edit; Lôrinczi, Lilla; Sipoş, Anca

    2007-01-01

    To compare the incidence of intestinal parasitosis between children with residence in urban and rural areas: to compare the efficacy of parasitologic diagnostic methods. In our study we included two lots of children. The first lot consisted in 74 children from rural areas from which we collected 44 samples of feces and 55 samples for the "Scotch tape" test. The second lot consisted in 214 children from urban areas from which we collected 44 samples of feces. We examined each sample of feces by three different methods. The study was performed between April to June 2006. The incidence of intestinal parasitosis increases in children from urban areas towards rural areas, and in children between 5 and 10 years. Ascariasis is the most frequent disease in both urban and rural areas. By examination of each fecal sample by three different methods, the number of positive cases increased. The residence in rural areas and age between 5 to 10 years are risk factors for intestinal parasitosis. The "Scotch tape" test was more efficient in Enterobius vermicularis infection than the methods performed from feces. We recommend using at the same time three diagnostic methods for feces examination to improve the diagnostic sensibility.

  18. Viability of the vascularly perfused, recirculating rat intestine and intestine-liver preparations

    Energy Technology Data Exchange (ETDEWEB)

    Hirayama, H.; Xu, X.; Pang, K.S. (Univ. of Toronto, Ontario (Canada))

    1989-08-01

    Function and stability of vascularly perfused, recirculating in situ rat intestine (I) and intestine-liver (IL) preparations were evaluated in fasted and nonfasted rats because these techniques may be readily applied in drug metabolism studies. The rat intestine was perfused with blood medium (7.5 ml/min) via the superior mesenteric artery, with the venous outflow draining into the portal vein, which, together with hepatic arterial flow (2.5 ml/min), constituted the total blood flow (10 ml/min) to the liver. Maintenance of intestinal membrane integrity was observed. Rapid ({sup 14}C)glucose absorption against a concentration gradient and a lack of ({sup 3}H)-polyethylene glycol 4000 (PEG 4000, less than 4%) and Evans blue absorption by the recirculating I and IL preparations resulted after bolus injections of these markers into the pyloric end of the duodenum. Other indexes that revealed stable intestinal and liver functions were the following: preservation of reservoir perfusate volume, constancy in perfusion pressure, bile flow, and hemoglobin concentrations, evidence of intestinal glucose utilization and liver glucose production, and a lack of significant leakage of serum glutamic oxalic transaminase. The intestine and liver consumed oxygen at relatively constant rates, but the consumption rates for the fasted tissues (I or L) were significantly higher than those for nonfasted tissues. These results indicate that the vascularly perfused I and IL preparations were maintained in a viable and stable state for a 2-h perfusion period.

  19. Intestinal Cgi-58 deficiency reduces postprandial lipid absorption.

    Science.gov (United States)

    Xie, Ping; Guo, Feng; Ma, Yinyan; Zhu, Hongling; Wang, Freddy; Xue, Bingzhong; Shi, Hang; Yang, Jian; Yu, Liqing

    2014-01-01

    Comparative Gene Identification-58 (CGI-58), a lipid droplet (LD)-associated protein, promotes intracellular triglyceride (TG) hydrolysis in vitro. Mutations in human CGI-58 cause TG accumulation in numerous tissues including intestine. Enterocytes are thought not to store TG-rich LDs, but a fatty meal does induce temporary cytosolic accumulation of LDs. Accumulated LDs are eventually cleared out, implying existence of TG hydrolytic machinery in enterocytes. However, identities of proteins responsible for LD-TG hydrolysis remain unknown. Here we report that intestine-specific inactivation of CGI-58 in mice significantly reduces postprandial plasma TG concentrations and intestinal TG hydrolase activity, which is associated with a 4-fold increase in intestinal TG content and large cytosolic LD accumulation in absorptive enterocytes during the fasting state. Intestine-specific CGI-58 knockout mice also display mild yet significant decreases in intestinal fatty acid absorption and oxidation. Surprisingly, inactivation of CGI-58 in intestine significantly raises plasma and intestinal cholesterol, and reduces hepatic cholesterol, without altering intestinal cholesterol absorption and fecal neutral sterol excretion. In conclusion, intestinal CGI-58 is required for efficient postprandial lipoprotein-TG secretion and for maintaining hepatic and plasma lipid homeostasis. Our animal model will serve as a valuable tool to further define how intestinal fat metabolism influences the pathogenesis of metabolic disorders, such as obesity and type 2 diabetes.

  20. Intestinal Cgi-58 deficiency reduces postprandial lipid absorption.

    Directory of Open Access Journals (Sweden)

    Ping Xie

    Full Text Available Comparative Gene Identification-58 (CGI-58, a lipid droplet (LD-associated protein, promotes intracellular triglyceride (TG hydrolysis in vitro. Mutations in human CGI-58 cause TG accumulation in numerous tissues including intestine. Enterocytes are thought not to store TG-rich LDs, but a fatty meal does induce temporary cytosolic accumulation of LDs. Accumulated LDs are eventually cleared out, implying existence of TG hydrolytic machinery in enterocytes. However, identities of proteins responsible for LD-TG hydrolysis remain unknown. Here we report that intestine-specific inactivation of CGI-58 in mice significantly reduces postprandial plasma TG concentrations and intestinal TG hydrolase activity, which is associated with a 4-fold increase in intestinal TG content and large cytosolic LD accumulation in absorptive enterocytes during the fasting state. Intestine-specific CGI-58 knockout mice also display mild yet significant decreases in intestinal fatty acid absorption and oxidation. Surprisingly, inactivation of CGI-58 in intestine significantly raises plasma and intestinal cholesterol, and reduces hepatic cholesterol, without altering intestinal cholesterol absorption and fecal neutral sterol excretion. In conclusion, intestinal CGI-58 is required for efficient postprandial lipoprotein-TG secretion and for maintaining hepatic and plasma lipid homeostasis. Our animal model will serve as a valuable tool to further define how intestinal fat metabolism influences the pathogenesis of metabolic disorders, such as obesity and type 2 diabetes.

  1. Lactobacillus GG and tributyrin supplementation reduce antibiotic-induced intestinal injury.

    Science.gov (United States)

    Cresci, Gail; Nagy, Laura E; Ganapathy, Vadivel

    2013-11-01

    Antibiotic therapy negatively alters the gut microbiota. Lactobacillus GG (LGG) decreases antibiotic-associated diarrhea (AAD) symptoms, but the mechanisms are unknown. Butyrate has beneficial effects on gut health. Altered intestinal gene expression occurs in the absence of gut microbiota. We hypothesized that antibiotic-induced changes in gut microbiota reduce butyrate production, varying genes involved with gut barrier integrity and water and electrolyte absorption, lending to AAD, and that simultaneous supplementation with LGG and/or tributyrin would prevent these changes. C57BL/6 mice aged 6-8 weeks received a chow diet while divided into 8 treatment groups (± saline, ± LGG, ± tributyrin, or both). Mice received treatments orally for 7 days with ± broad-spectrum antibiotics. Water intake was recorded daily and body weight was measured. Intestine tissue samples were obtained and analyzed for expression of genes and proteins involved with water and electrolyte absorption, butyrate transport, and gut integrity via polymerase chain reaction and immunohistochemistry. Antibiotics decreased messenger RNA (mRNA) expression (butyrate transporter and receptor, Na(+)/H(+) exchanger, Cl(-)/HCO3 (-), and a water channel) and protein expression (butyrate transporter, Na(+)/H(+) exchanger, and tight junction proteins) in the intestinal tract. LGG and/or tributyrin supplementation maintained intestinal mRNA expression to that of the control animals, and tributyrin maintained intestinal protein intensity expression to that of control animals. Broad-spectrum antibiotics decrease expression of anion exchangers, butyrate transporter and receptor, and tight junction proteins in mouse intestine. Simultaneous oral supplementation with LGG and/or tributyrin minimizes these losses. Optimizing intestinal health with LGG and/or tributyrin may offer a preventative therapy for AAD.

  2. Tensile rupture of medial arterial tissue studied by X-ray micro-tomography on stained samples

    OpenAIRE

    Helfenstein-Didier, Clémentine; Taïnoff, Damien; Viville, Julien; Adrien, Jérôme; Maire, Éric; Badel, Pierre

    2017-01-01

    Detailed characterization of damage and rupture mechanics of arteries is one the current challenges in vascular biomechanics, which requires developing suitable experimental approaches. This paper introduces an approach using in situ tensile tests in an X-ray micro-tomography setup to observe mechanisms of damage initiation and progression in medial layers of porcine aortic samples. The technique requires the use of sodium polytungstate as a contrast agent, of which the conditions for use are...

  3. A rapid analytical method to quantify complex organohalogen contaminant mixtures in large samples of high lipid mammalian tissues.

    Science.gov (United States)

    Desforges, Jean-Pierre; Eulaers, Igor; Periard, Luke; Sonne, Christian; Dietz, Rune; Letcher, Robert J

    2017-06-01

    In vitro investigations of the health impact of individual chemical compounds have traditionally been used in risk assessments. However, humans and wildlife are exposed to a plethora of potentially harmful chemicals, including organohalogen contaminants (OHCs). An alternative exposure approach to individual or simple mixtures of synthetic OHCs is to isolate the complex mixture present in free-ranging wildlife, often non-destructively sampled from lipid rich adipose. High concentration stock volumes required for in vitro investigations do, however, pose a great analytical challenge to extract sufficient amounts of complex OHC cocktails. Here we describe a novel method to easily, rapidly and efficiently extract an environmentally accumulated and therefore relevant contaminant cocktail from large (10-50 g) marine mammal blubber samples. We demonstrate that lipid freeze-filtration with acetonitrile removes up to 97% of blubber lipids, with minimal effect on the efficiency of OHC recovery. Sample extracts after freeze-filtration were further processed to remove residual trace lipids via high-pressure gel permeation chromatography and solid phase extraction. Average recoveries of OHCs from triplicate analysis of killer whale (Orcinus orca), polar bear (Ursus maritimus) and pilot whale (Globicephala spp.) blubber standard reference material (NIST SRM-1945) ranged from 68 to 80%, 54-92% and 58-145%, respectively, for 13 C-enriched internal standards of six polychlorinated biphenyl congeners, 16 organochlorine pesticides and four brominated flame retardants. This approach to rapidly generate OHC mixtures shows great potential for experimental exposures using complex contaminant mixtures, research or monitoring driven contaminant quantification in biological samples, as well as the untargeted identification of emerging contaminants. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Effect of in ovo administration of an adult-derived microbiota on establishment of the intestinal microbiome in chickens.

    Science.gov (United States)

    Pedroso, Adriana A; Batal, Amy B; Lee, Margie D

    2016-05-01

    OBJECTIVE To determine effects of in ovo administration of a probiotic on development of the intestinal microbiota of 2 genetic lineages (modern and heritage) of chickens. SAMPLE 10 newly hatched chicks and 40 fertile eggs to determine intestinal microbiota at hatch, 900 fertile eggs to determine effects of probiotic on hatchability, and 1,560 chicks from treated or control eggs. PROCEDURES A probiotic competitive-exclusion product derived from adult microbiota was administered in ovo to fertile eggs of both genetic lineages. Cecal contents and tissues were collected from embryos, newly hatched chicks, and chicks. A PCR assay was used to detect bacteria present within the cecum of newly hatched chicks. Fluorescence in situ hybridization and vitality staining were used to detect viable bacteria within intestines of embryos. The intestinal microbiota was assessed by use of 16S pyrosequencing. RESULTS Microscopic evaluation of embryonic cecal contents and tissues subjected to differential staining techniques revealed viable bacteria in low numbers. Development of the intestinal microbiota of broiler chicks of both genetic lineages was enhanced by in ovo administration of adult microbiota. Although the treatment increased diversity and affected composition of the microbiota of chicks, most bacterial species present in the probiotic were transient colonizers. However, the treatment decreased the abundance of undesirable bacterial species within heritage lineage chicks. CONCLUSIONS AND CLINICAL RELEVANCE In ovo inoculation of a probiotic competitive-exclusion product derived from adult microbiota may be a viable method of managing development of the microbiota and reducing the prevalence of pathogenic bacteria in chickens.

  5. Transient inhibition of ROR-γt therapeutically limits intestinal inflammation by reducing TH17 cells and preserving ILC3

    Science.gov (United States)

    Withers, David R.; Hepworth, Matthew R.; Wang, Xinxin; Mackley, Emma C.; Halford, Emily E.; Dutton, Emma E.; Marriott, Clare L.; Brucklacher-Waldert, Verena; Veldhoen, Marc; Kelsen, Judith; Baldassano, Robert N.; Sonnenberg, Gregory F.

    2016-01-01

    RAR-related orphan receptor γt (ROR-γt) directs differentiation of pro-inflammatory T helper 17 (TH17) cells and is a potential therapeutic target in chronic autoimmune and inflammatory diseases1–3. However, ROR-γt-dependent group 3 innate lymphoid cells (ILC3s) provide essential immunity and tissue protection in the intestine4–11, suggesting that targeting ROR-γt could also result in impaired host defense to infection or enhanced tissue damage. Here, we demonstrate that transient chemical inhibition of ROR-γt in mice selectively reduces cytokine production from TH17 cells but not ILC3s in the context of intestinal infection with Citrobacter rodentium, resulting in preserved innate immunity. Transient genetic deletion of ROR-γt in mature ILC3s also did not impair cytokine responses in the steady state or during infection. Finally, pharmacologic inhibition of ROR-γt provided therapeutic benefit in mouse models of intestinal inflammation, and reduced the frequencies of TH17 cells but not ILC3s isolated from primary intestinal samples of individuals with inflammatory bowel disease (IBD). Collectively, these results reveal differential requirements for ROR-γt in the maintenance of TH17 cell versus ILC3 responses, and suggest that transient inhibition of ROR-γt is a safe and effective therapeutic approach during intestinal inflammation. PMID:26878233

  6. Effect of sample preparation techniques on the concentrations and distributions of elements in biological tissues using µSRXRF: a comparative study

    International Nuclear Information System (INIS)

    Al-Ebraheem, A; Dao, E; Desouza, E; McNeill, F E; Farquharson, M J; Li, C; Wainman, B C

    2015-01-01

    Routine tissue sample preparation using chemical fixatives is known to preserve the morphology of the tissue being studied. A competitive method, cryofixation followed by freeze drying, involves no chemical agents and maintains the biological function of the tissue. The possible effects of both sample preparation techniques in terms of the distribution of bio-metals (calcium (Ca), copper (Cu) zinc (Zn), and iron (Fe) specifically) in human skin tissue samples was investigated. Micro synchrotron radiation x-ray fluorescence (μSRXRF) was used to map bio-metal distribution in epidermal and dermal layers of human skin samples from various locations of the body that have been prepared using both techniques. For Ca, Cu and Zn, there were statistically significant differences between the epidermis and dermis using the freeze drying technique (p = 0.02, p < 0.01, and p < 0.01, respectively). Also using the formalin fixed, paraffin embedded technique the levels of Ca, Cu and Zn, were significantly different between the epidermis and dermis layers (p = 0.03, p < 0.01, and p < 0.01, respectively). However, the difference in levels of Fe between the epidermis and dermis was unclear and further analysis was required. The epidermis was further divided into two sub-layers, one mainly composed of the stratum corneum and the other deeper layer, the stratum basale. It was found that the difference between the distribution of Fe in the two epidermal layers using the freeze drying technique resulted in a statistically significant difference (p = 0.012). This same region also showed a difference in Fe using the formalin fixed, paraffin embedded technique (p < 0.01). The formalin fixed, paraffin embedded technique also showed a difference between the deeper epidermal layer and the dermis (p < 0.01). It can be concluded that studies involving Ca, Cu and Zn might show similar results using both sample preparation techniques, however studies involving Fe would need more

  7. Tissue and serum samples of patients with papillary thyroid cancer with and without benign background demonstrate different altered expression of proteins

    Directory of Open Access Journals (Sweden)

    Mardiaty Iryani Abdullah

    2016-09-01

    Full Text Available Background Papillary thyroid cancer (PTC is mainly diagnosed using fine-needle aspiration biopsy. This most common form of well-differentiated thyroid cancer occurs with or without a background of benign thyroid goiter (BTG. Methods In the present study, a gel-based proteomics analysis was performed to analyse the expression of proteins in tissue and serum samples of PTC patients with (PTCb; n = 6 and without a history of BTG (PTCa; n = 8 relative to patients with BTG (n = 20. This was followed by confirmation of the levels of proteins which showed significant altered abundances of more than two-fold difference (p < 0.01 in the tissue and serum samples of the same subjects using ELISA. Results The data of our study showed that PTCa and PTCb distinguish themselves from BTG in the types of tissue and serum proteins of altered abundance. While higher levels of alpha-1 antitrypsin (A1AT and heat shock 70 kDa protein were associated with PTCa, lower levels of A1AT, protein disulfide isomerase and ubiquitin-conjugating enzyme E2 N seemed apparent in the PTCb. In case of the serum proteins, higher abundances of A1AT and alpha 1-beta glycoprotein were detected in PTCa, while PTCb was associated with enhanced apolipoprotein A-IV and alpha 2-HS glycoprotein (AHSG. The different altered expression of tissue and serum A1AT as well as serum AHSG between PTCa and PTCb patients were also validated by ELISA. Discussion The distinctive altered abundances of the tissue and serum proteins form preliminary indications that PTCa and PTCb are two distinct cancers of the thyroid that are etiologically and mechanistically different although it is currently not possible to rule out that they may also be due other reasons such as the different stages of the malignant disease. These proteins stand to have a potential use as tissue or serum biomarkers to discriminate the three different thyroid neoplasms although this requires further validation in clinically

  8. Amelioration of Chemotherapy-Induced Intestinal Mucositis by Orally Administered Probiotics in a Mouse Model

    Science.gov (United States)

    Jiang, Chun-Bin; Cheng, Mei-Lien; Liu, Chia-Yuan; Chang, Szu-Wen; Chiang Chiau, Jen-Shiu; Lee, Hung-Chang

    2015-01-01

    Background and Aims Intestinal mucositis is a frequently encountered side effect in oncology patients undergoing chemotherapy. No well-established or up to date therapeutic strategies are available. To study a novel way to alleviate mucositis, we investigate the effects and safety of probiotic supplementation in ameliorating 5-FU-induced intestinal mucositis in a mouse model. Methods Seventy-two mice were injected saline or 5-Fluorouracil (5-FU) intraperitoneally daily. Mice were either orally administrated daily saline, probiotic suspension of Lactobacillus casei variety rhamnosus (Lcr35) or Lactobacillus acidophilus and Bifidobacterium bifidum (LaBi). Diarrhea score, pro-inflammatory cytokines serum levels, intestinal villus height and crypt depth and total RNA from tissue were assessed. Samples of blood, liver and spleen tissues were assessed for translocation. Results Marked diarrhea developed in the 5-FU groups but was attenuated after oral Lcr35 and LaBi administrations. Diarrhea scores decreased significantly from 2.64 to 1.45 and 0.80, respectively (Pprobiotics administration. We also found TNF-α, IL-1β and IL-6 mRNA expressions were up-regulated in intestinal mucositis tissues following 5-FU treatment (TNF-α: 4.35 vs. 1.18, IL-1β: 2.29 vs. 1.07, IL-6: 1.49 vs. 1.02) and that probiotics treatment suppressed this up-regulation (Pprobiotics Lcr35 and LaBi can ameliorate chemotherapy-induced intestinal mucositis in a mouse model. This suggests probiotics may serve as an alternative therapeutic strategy for the prevention or management of chemotherapy-induced mucositis in the future. PMID:26406888

  9. Evaluation of reference genes for gene expression in red-tailed phascogale (Phascogale calura liver, lung, small intestine and spleen

    Directory of Open Access Journals (Sweden)

    Oselyne T.W. Ong

    2016-10-01

    Full Text Available Background Reference genes serve an important role as an endogenous control/standard for data normalisation in gene expression studies. Although reference genes have recently been suggested for marsupials, independent analysis of reference genes on different immune tissues is yet to be tested. Therefore, an assessment of reference genes is needed for the selection of stable, expressed genes across different marsupial tissues. Methods The study was conducted on red-tailed phascogales (Phascogale calura using five juvenile and five adult males. The stability of five reference genes (glyceraldehyde-3-phosphate dehydrogenase, GAPDH; β-actin, ACTB; 18S rRNA, 18S; 28S rRNA, 28S; and ribosomal protein L13A, RPL13A was investigated using SYBR Green and analysed with the geNorm application available in qBasePLUS software. Results Gene stability for juvenile and adult tissue samples combined show that GAPDH was most stable in liver and lung tissue, and 18S in small intestine and spleen. While all reference genes were suitable for small intestine and spleen tissues, all reference genes except 28S were stable for lung and only 18S and 28S were stable for liver tissue. Separating the two age groups, we found that two different reference genes were considered stable in juveniles (ACTB and GAPDH and adults (18S and 28S, and RPL13A was not stable for juvenile small intestine tissue. Except for 28S, all reference genes were stable in juvenile and adult lungs, and all five reference genes were stable in spleen tissue. Discussion Based on expression stability, ACTB and GAPDH are suitable for all tissues when studying the expression of marsupials in two age groups, except for adult liver tissues. The expression stability between juvenile and adult liver tissue was most unstable, as the stable reference genes for juveniles and adults were different. Juvenile and adult lung, small intestine and spleen share similar stable reference genes, except for small intestine

  10. Dual-priming oligonucleotide-based multiplex PCR using tissue samples from the rapid urease test kit for the detection of Helicobacter pylori in bleeding peptic ulcers.

    Science.gov (United States)

    Chung, Woo Chul; Jeon, Eun Jung; Oh, Jung Hwan; Park, Jae Myung; Kim, Tae Ho; Cheung, Dae Young; Kim, Byung Wook; Kim, Sung Soo; Kim, Jin Il

    2016-08-01

    In patients with peptic ulcer bleeding (PUB), diagnostic tests for Helicobacter pylori (H. pylori) infection have low sensitivity. The aim of our study was to investigate the diagnostic yield of dual-priming oligonucleotide-based multiplex (DPO)-PCR using tissue samples from the rapid urease test (RUT, CLO(®)test) kit in patients with PUB. We prospectively enrolled patients with PUB. During second-look endoscopy, gastric biopsy specimens for histology and RUT were obtained from a total of 170 patients. DPO-PCR tests were performed on tissue samples obtained from the CLO(®)test kit. If testing for H. pylori was negative, endoscopy with re-biopsy was performed 8 weeks after the bleeding episode. H. pylori-associated bleeding was confirmed in 64.1% (109/170) of the patients. At the bleeding episode, the diagnostic sensitivities of RUT, histology, and DPO-PCR test were 47.7% (52/109), 71.6% (78/109) and 97.2% (106/109), respectively (p<0.01). The specificity of the DPO-PCR test was 91.8% (56/61). The positive predictive value (PPV) of the DPO-PCR test was 95.5% (106/111), and its negative predictive value (NPV) was 94.9% (56/59). In patients with PUB, the DPO-PCR test could be a useful diagnostic tool for H. pylori infection. Particularly given a negative RUT result, subsequent DPO-PCR testing of tissue samples from the CLO(®)test kit could be of considerable benefit. Copyright © 2016 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.

  11. A novel method for sample preparation of fresh lung cancer tissue for proteomics analysis by tumor cell enrichment and removal of blood contaminants

    Directory of Open Access Journals (Sweden)

    Orre Lotta

    2010-02-01

    was an effective removal of contaminants from red blood cells and plasma proteins resulting in larger proteome coverage compared to the direct lysis of frozen samples. This sample preparation method may be successfully implemented for the discovery of lung cancer biomarkers on tissue samples using mass spectrometry-based proteomics.

  12. Zonulin, a newly discovered modulator of intestinal permeability, and its expression in coeliac disease.

    Science.gov (United States)

    Fasano, A; Not, T; Wang, W; Uzzau, S; Berti, I; Tommasini, A; Goldblum, S E

    2000-04-29

    We identified zonulin, a novel human protein analogue to the Vibrio cholerae derived Zonula occludens toxin, which induces tight junction disassembly and a subsequent increase in intestinal permeability in non-human primate intestinal epithelia. Zonulin expression was raised in intestinal tissues during the acute phase of coeliac disease, a clinical condition in which tight junctions are opened and permeability is increased.

  13. Pathogen invasion changes the intestinal microbiota composition and induces innate immune responses in the zebrafish intestine.

    Science.gov (United States)

    Yang, Hui-Ting; Zou, Song-Song; Zhai, Li-Juan; Wang, Yao; Zhang, Fu-Miao; An, Li-Guo; Yang, Gui-Wen

    2017-12-01

    Numerous bacteria are harbored in the animal digestive tract and are impacted by several factors. Intestinal microbiota homeostasis is critical for maintaining the health of an organism. However, how pathogen invasion affects the microbiota composition has not been fully clarified. The mechanisms for preventing invasion by pathogenic microorganisms are yet to be elucidated. Zebrafish is a useful model for developmental biology, and studies in this organism have gradually become focused on intestinal immunity. In this study, we analyzed the microbiota of normal cultivated and infected zebrafish intestines, the aquarium water and feed samples. We found that the predominant bacteria in the zebrafish intestine belonged to Gammaproteobacteria (67%) and that feed and environment merely influenced intestinal microbiota composition only partially. Intestinal microbiota changed after a pathogenic bacterial challenge. At the genus level, the abundance of some pathogenic intestinal bacteria increased, and these genera included Halomonas (50%), Pelagibacterium (3.6%), Aeromonas (2.6%), Nesterenkonia (1%), Chryseobacterium (3.4‰), Mesorhizobium (1.4‰), Vibrio (1‰), Mycoplasma (0.7‰) and Methylobacterium (0.6‰) in IAh group. However, the abundance of some beneficial intestinal bacteria decreased, and these genera included Nitratireductor (0.8‰), Enterococcus (0.8‰), Brevundimonas (0.7‰), Lactococcus (0.7‰) and Lactobacillus (0.4‰). Additionally, we investigated the innate immune responses after infection. ROS levels in intestine increased in the early stages after a challenge and recovered subsequently. The mRNA levels of antimicrobial peptide genes lectin, hepcidin and defensin1, were upregulated in the intestine after pathogen infection. These results suggested that the invasion of pathogen could change the intestinal microbiota composition and induce intestinal innate immune responses in zebrafish. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. A lab-on-a-chip system integrating tissue sample preparation and multiplex RT-qPCR for gene expression analysis in point-of-care hepatotoxicity assessment.

    Science.gov (United States)

    Lim, Geok Soon; Chang, Joseph S; Lei, Zhang; Wu, Ruige; Wang, Zhiping; Cui, Kemi; Wong, Stephen

    2015-10-21

    A truly practical lab-on-a-chip (LOC) system for point-of-care testing (POCT) hepatotoxicity assessment necessitates the embodiment of full-automation, ease-of-use and "sample-in-answer-out" diagnostic capabilities. To date, the reported microfluidic devices for POCT hepatotoxicity assessment remain rudimentary as they largely embody only semi-quantitative or single sample/gene detection capabilities. In this paper, we describe, for the first time, an integrated LOC system that is somewhat close to a practical POCT hepatotoxicity assessment device - it embodies both tissue sample preparation and multiplex real-time RT-PCR. It features semi-automation, is relatively easy to use, and has "sample-in-answer-out" capabilities for multiplex gene expression analysis. Our tissue sample preparation module incorporating both a microhomogenizer and surface-treated paramagnetic microbeads yielded high purity mRNA extracts, considerably better than manual means of extraction. A primer preloading surface treatment procedure and the single-loading inlet on our multiplex real-time RT-PCR module simplify off-chip handling procedures for ease-of-use. To demonstrate the efficacy of our LOC system for POCT hepatotoxicity assessment, we perform a preclinical animal study with the administration of cyclophosphamide, followed by gene expression analysis of two critical protein biomarkers for liver function tests, aspartate transaminase (AST) and alanine transaminase (ALT). Our experimental results depict normalized fold changes of 1.62 and 1.31 for AST and ALT, respectively, illustrating up-regulations in their expression levels and hence validating their selection as critical genes of interest. In short, we illustrate the feasibility of multiplex gene expression analysis in an integrated LOC system as a viable POCT means for hepatotoxicity assessment.

  15. Sensitive Mie scattering immunoagglutination assay of porcine reproductive and respiratory syndrome virus (PRRSV) from lung tissue samples in a microfluidic chip.

    Science.gov (United States)

    Song, Jae-Young; Lee, Chang-Hee; Choi, Eun-Jin; Kim, Keesung; Yoon, Jeong-Yeol

    2011-12-01

    A microfluidic immunosensor utilizing Mie scattering immunoaggultination assay was developed for rapid and sensitive detection of porcine reproductive and respiratory syndrome virus (PRRSV) from lung tissue samples of domesticated pigs. Antibodies against PRRSV were conjugated to the surface of highly carboxylated polystyrene microparticles (diameter=920nm) and mixed with the diluted PRRSV tissue samples in a Y-shaped microchannel. Antibody-antigen binding induced microparticle immunoagglutination, which was detected by measuring the forward 45° light scattering of 380nm incident beam using microcallipered, proximity fiber optics. For comparison, multi-well experiments were also performed using the same optical detection setup. The detection limit was determined to be 10(-3)TCID(50)ml(-1) for PRRSV dissolved in PBS, while those of previous RT-PCR studies for PRRSV were 10(1)TCID(50)ml(-1) (conventional assays) or Mie scattering simulations were able to predict the shape of the PRRSV standard curve, indicating that any non-linearity of the standard curve can be interpreted purely as an optical phenomenon. Each assay took less than 5min. A strong correlation could be found between RT-PCR and this method for the lung tissue samples, even though their respective detection mechanisms are different fundamentally (nucleic acids for RT-PCR and virus antigens for light scattering immunoagglutination assay). Several different dilution factors were also tested for tissue samples, and 1/10 and 1/100 were found to be usable. If the microfluidic chips are used only once (i.e. without re-using them), both superior sensitivity and satisfactory specificity can be demonstrated. Specificity studies revealed the presence of Type II PRRSV and non-presence of Type I PRRSV and that the microfluidic chip assay could detect Type II North American strain of PRRSV for the animals tested. This work demonstrates the potential of the Mie scattering immunoassay on a microfluidic chip towards

  16. Locations of gut-associated lymphoid tissue in the 3-month-old chicken: a review.

    Science.gov (United States)

    Casteleyn, C; Doom, M; Lambrechts, E; Van den Broeck, W; Simoens, P; Cornillie, P

    2010-06-01

    The lymphoid tissue that is associated with the intestinal tract, the so-called gut-associated lymphoid tissue (GALT), is well developed in the chicken. Depending on the location, it is present as aggregations of lymphoid cells, or organized in lymphoid follicles and tonsils. From proximal to distal, the intestinal tract contains a pharyngeal tonsil, diffuse lymphoid tissue and lymphoid follicles in the cervical and thoracic parts of the oesophagus, an oesophageal tonsil, diffuse lymphoid tissue in the proventriculus, a pyloric tonsil, Peyer's patches, Meckel's diverticulum, two caecal tonsils, diffuse lymphoid tissue in the rectum, the bursa of Fabricius, and diffuse lymphoid tissue in the wall of the proctodeum. The lymphoid tissues are frequently covered by a lympho-epithelium that is infiltrated by lymphoid cells. Such an epithelium often contains M or microfold cells, which are specialized in antigen sampling and transport antigens to the underlying lymphoid tissue. A solid knowledge of the avian GALT could contribute to the development of vaccines to be administered orally. Additionally, immune stimulation via pre- and probiotics is based on the presence of a well-developed intestinal immune system.

  17. Temporary intestinal ischemia for radiation protection

    International Nuclear Information System (INIS)

    Lote, K.

    1983-01-01

    The most important determinant of cellular radiosensivity is the tissue oxygen content at the time of irradiation. The purpose of the present experimental work was to assess a new iscemia-inducing method in order to reduce normal tissue radiation damage during radiotherapy. Temporary ischemia was induced in a cat small intestine by degraded starch microspheres. Regional arterial and tissue blod flow immediately fell by 85% with subsequent normalization within 26 minutes after microsphere injection. No tendency of small vessel thrombosis caused by starch sphere embolization in combination with previous or current intestinal irradiation was detected. Starch sphere remenants were rapidly engulfed by, and persisted within tissue macrophages for 14 days without causing intestinal inflammatory reactions. In vitro studies showed that human platelets neither adhered to nor were aggregated by starch microspheres. The new method, wich occlude arteriolar vessels distal to the mesentric arterial arcades and thus largely excludes collateral blood flow, seems suited to provide effictive and selective feline small intestinal hypoxic radiation protection. This conclusion may also be valid in man

  18. Intestinal Stem Cell Niche: The Extracellular Matrix and Cellular Components

    Directory of Open Access Journals (Sweden)

    Laween Meran

    2017-01-01

    Full Text Available The intestinal epithelium comprises a monolayer of polarised columnar cells organised along the crypt-villus axis. Intestinal stem cells reside at the base of crypts and are constantly nourished by their surrounding niche for maintenance, self-renewal, and differentiation. The cellular microenvironment including the adjacent Paneth cells, stromal cells, smooth muscle cells, and neural cells as well as the extracellular matrix together constitute the intestinal stem cell niche. A dynamic regulatory network exists among the epithelium, stromal cells, and the matrix via complex signal transduction to maintain tissue homeostasis. Dysregulation of these biological or mechanical signals could potentially lead to intestinal injury and disease. In this review, we discuss the role of different intestinal stem cell niche components and dissect the interaction between dynamic matrix factors and regulatory signalling during intestinal stem cell homeostasis.

  19. The neonatal intestinal vasculature: contributing factors to necrotizing enterocolitis.

    Science.gov (United States)

    Nankervis, Craig A; Giannone, Peter J; Reber, Kristina M

    2008-04-01

    Based on the demonstration of coagulation necrosis, it is clear that intestinal ischemia plays a role in the pathogenesis of necrotizing enterocolitis (NEC). Intestinal vascular resistance is determined by a dynamic balance between vasoconstrictive and vasodilatory inputs. In the newborn, this balance heavily favors vasodilation secondary to the copious production of endothelium-derived nitric oxide (NO), a circumstance which serves to ensure adequate blood flow and thus oxygen delivery to the rapidly growing intestine. Endothelial cell injury could shift this balance in favor of endothelin (ET)-1-mediated vasoconstriction, leading to intestinal ischemia and tissue injury. Evidence obtained from animal models and from human tissue collected from infants with NEC implicates NO and ET-1 dysregulation in the pathogenesis of NEC. Strategies focused on maintaining the delicate balance favoring vasodilation in the newborn intestinal circulation may prove to be useful in the prevention and treatment of NEC.

  20. The use of laser microdissection in the identification of suitable reference genes for normalization of quantitative real-time PCR in human FFPE epithelial ovarian tissue samples.

    Directory of Open Access Journals (Sweden)

    Jing Cai

    Full Text Available Quantitative real-time PCR (qPCR is a powerful and reproducible method of gene expression analysis in which expression levels are quantified by normalization against reference genes. Therefore, to investigate the potential biomarkers and therapeutic targets for epithelial ovarian cancer by qPCR, it is critical to identify stable reference genes. In this study, twelve housekeeping genes (ACTB, GAPDH, 18S rRNA, GUSB, PPIA, PBGD, PUM1, TBP, HRPT1, RPLP0, RPL13A, and B2M were analyzed in 50 ovarian samples from normal, benign, borderline, and malignant tissues. For reliable results, laser microdissection (LMD, an effective technique used to prepare homogeneous starting material, was utilized to precisely excise target tissues or cells. One-way analysis of variance (ANOVA and nonparametric (Kruskal-Wallis tests were used to compare the expression differences. NormFinder and geNorm software were employed to further validate the suitability and stability of the candidate genes. Results showed that epithelial cells occupied a small percentage of the normal ovary indeed. The expression of ACTB, PPIA, RPL13A, RPLP0, and TBP were stable independent of the disease progression. In addition, NormFinder and geNorm identified the most stable combination (ACTB, PPIA, RPLP0, and TBP and the relatively unstable reference gene GAPDH from the twelve commonly used housekeeping genes. Our results highlight the use of homogeneous ovarian tissues and multiple-reference normalization strategy, e.g. the combination of ACTB, PPIA, RPLP0, and TBP, for qPCR in epithelial ovarian tissues, whereas GAPDH, the most commonly used reference gene, is not recommended, especially as a single reference gene.

  1. The use of laser microdissection in the identification of suitable reference genes for normalization of quantitative real-time PCR in human FFPE epithelial ovarian tissue samples.

    Science.gov (United States)

    Cai, Jing; Li, Tao; Huang, Bangxing; Cheng, Henghui; Ding, Hui; Dong, Weihong; Xiao, Man; Liu, Ling; Wang, Zehua

    2014-01-01

    Quantitative real-time PCR (qPCR) is a powerful and reproducible method of gene expression analysis in which expression levels are quantified by normalization against reference genes. Therefore, to investigate the potential biomarkers and therapeutic targets for epithelial ovarian cancer by qPCR, it is critical to identify stable reference genes. In this study, twelve housekeeping genes (ACTB, GAPDH, 18S rRNA, GUSB, PPIA, PBGD, PUM1, TBP, HRPT1, RPLP0, RPL13A, and B2M) were analyzed in 50 ovarian samples from normal, benign, borderline, and malignant tissues. For reliable results, laser microdissection (LMD), an effective technique used to prepare homogeneous starting material, was utilized to precisely excise target tissues or cells. One-way analysis of variance (ANOVA) and nonparametric (Kruskal-Wallis) tests were used to compare the expression differences. NormFinder and geNorm software were employed to further validate the suitability and stability of the candidate genes. Results showed that epithelial cells occupied a small percentage of the normal ovary indeed. The expression of ACTB, PPIA, RPL13A, RPLP0, and TBP were stable independent of the disease progression. In addition, NormFinder and geNorm identified the most stable combination (ACTB, PPIA, RPLP0, and TBP) and the relatively unstable reference gene GAPDH from the twelve commonly used housekeeping genes. Our results highlight the use of homogeneous ovarian tissues and multiple-reference normalization strategy, e.g. the combination of ACTB, PPIA, RPLP0, and TBP, for qPCR in epithelial ovarian tissues, whereas GAPDH, the most commonly used reference gene, is not recommended, especially as a single reference gene.

  2. Evaluation of Two Enzyme-Linked Immunosorbent Assay Kits for Chikungunya Virus IgM Using Samples from Deceased Organ and Tissue Donors.

    Science.gov (United States)

    Prince, Harry E; Altrich, Michelle L; Nowicki, Marek J

    2016-10-01

    The identification of nearly 3,500 cases of chikungunya virus (CHIKV) infection in U.S. residents returning in 2014 and 2015 from areas in which it is endemic has raised concerns within the transplant community that, should recently infected individuals become organ and/or tissue donors, CHIKV would be transmitted to transplant recipients. Thus, tests designed to detect recent CHIKV infection among U.S. organ and tissue donors may become necessary in the future. Accordingly, we evaluated 2 enzyme-linked immunosorbent assays (ELISAs) for CHIKV IgM readily available in the United States using 1,000 deidentified serum or plasma specimens collected from donors between November 2014 and March 2015. The Euroimmun indirect ELISA identified 38 reactive specimens; however, all 38 were negative for CHIKV IgG and IgM in immunofluorescence assays (IFAs) conducted at a reference laboratory and, thus, were falsely reactive in the Euroimmun CHIKV IgM assay. The InBios IgM-capture ELISA identified 26 reactive samples, and one was still reactive (index ≥ 1.00) when retested using the InBios kit with a background subtraction modification to identify false reactivity. This reactive specimen was CHIKV IgM negative but IgG positive by IFAs at two reference laboratories; plaque reduction neutralization testing (PRNT) demonstrated CHIKV-specific reactivity. The IgG and PRNT findings strongly suggest that the InBios CHIKV IgM-reactive result represents true reactivity, even though the IgM IFA result was negative. If testing organ/tissue donors for CHIKV IgM becomes necessary, the limitations of the currently available CHIKV IgM ELISAs and options for their optimization must be understood to avoid organ/tissue wastage due to falsely reactive results. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  3. 1300 nm and 890 nm OCT images of oral cancer tissue engineered models and biopsy samples offer complimentary performance (Conference Presentation)

    Science.gov (United States)

    Boadi, Joseph; Byers, Robert A.; Fernandes, Jon; Mittar, Shweta; Hearnden, Vanessa; Lu, Zenghai; MacNeil, Sheila; Thornhill, Martin; Murdoch, Craig; Hunter, Keith D.; McKechnie, Alasdair; Matcher, Stephen J.

    2016-02-01

    OCT has demonstrated great potential to non-invasively detect oral epithelial cancers, potentially guiding biopsy and surgical resection. On non-ophthalmic tissues the preferred illumination wavelength is 1300 nm. Previous studies on skin have shown that useful image data can also be obtained at shorter wavelengths, with systems at 1060 nm and 820 nm offering reduced depth penetration but higher contrast. Here we apply a similar comparison to tissue engineered models of oral cancer and also to human biopsy samples, generally finding a similar trend. 1300 nm multi-beam OCT (Michelson Diagnostics EX1301) visualises stromal structures and surface keratin more clearly, providing useful image contrast down to around 1 mm. This system was compared with an ultra-high resolution home-built system operating at 890 nm (2.5 micron resolution vs 7.5 micron axial resolution for the EX1301). The UHR system reveals epithelial features more clearly, especially in the DOK pre-invasive cell line model and the biopsy samples. The relative effects of center wavelength vs axial resolution in generating the differential, wavelength-dependent contrast are assessed and the OCT biopsy images are compared with contemporary histology.

  4. Intestinal cytochromes P450 regulating the intestinal microbiota and its probiotic profile

    Directory of Open Access Journals (Sweden)

    Eugenia Elefterios Venizelos Bezirtzoglou

    2012-09-01

    Full Text Available Cytochromes P450 (CYPs enzymes metabolize a large variety of xenobiotic substances. In this vein, a plethora of studies were conducted to investigate their role, as cytochromes are located in both liver and intestinal tissues. The P450 profile of the human intestine has not been fully characterized. Human intestine serves primarily as an absorptive organ for nutrients, although it has also the ability to metabolize drugs. CYPs are responsible for the majority of phase I drug metabolism reactions. CYP3A represents the major intestinal CYP (80% followed by CYP2C9. CYP1A is expressed at high level in the duodenum, together with less abundant levels of CYP2C8-10 and CYP2D6. Cytochromes present a genetic polymorphism intra- or interindividual and intra- or interethnic. Changes in the pharmacokinetic profile of the drug are associated with increased toxicity due to reduced metabolism, altered efficacy of the drug, increased production of toxic metabolites, and adverse drug interaction. The high metabolic capacity of the intestinal flora is due to its enormous pool of enzymes, which catalyzes reactions in phase I and phase II drug metabolism. Compromised intestinal barrier conditions, when rupture of the intestinal integrity occurs, could increase passive paracellular absorption. It is clear that high microbial intestinal charge following intestinal disturbances, ageing, environment, or food-associated ailments leads to the microbial metabolism of a drug before absorption. The effect of certain bacteria having a benefic action on the intestinal ecosystem has been largely discussed during the past few years by many authors. The aim of the probiotic approach is to repair the deficiencies in the gut flora and establish a protective effect. There is a tentative multifactorial association of the CYP (P450 cytochrome role in the different diseases states, environmental toxic effects or chemical exposures and nutritional status.

  5. Precise simultaneous quantification of methadone and cocaine in rat serum and brain tissue samples following their successive i.p. administration.

    Science.gov (United States)

    Nakhla, David S; Hussein, Lobna A; Magdy, N; Abdallah, Inas A; Hassan, Hazem E

    2017-03-24

    A sensitive high-performance liquid chromatography (HPLC) assay with dual UV detection has been developed and validated for the simultaneous quantification of methadone and cocaine in rat serum and brain tissue samples. Liquid-liquid extraction using hexanes was applied for samples extraction with Levo-Tetrahydropalmatine (L-THP) as the internal standard. Chromatographic separation of the analytes was achieved on a reversed-phase Waters Symmetry ® C 18 column (150mm×4.6mm, 5μm). A gradient elution was employed with a mobile phase consisting of 5mM potassium phosphate containing 0.1% triethylamine (pH=6.5) (A) and acetonitrile (B) with a flow rate of 1mL/min. UV detection was employed at 215nm and 235nm for the determination of methadone and cocaine, respectively. The calibration curves were linear over the range of 0.05-10μg/mL for both methadone and cocaine. The assay was validated according to FDA guidelines for bioanalytical method validation and results were satisfactory and met FDA criteria. Inter-day accuracy values of serum and brain samples ranged from 96.97 to 105.59% while intra-day accuracy values ranged from 91.49 to 111.92%. Stability assays showed that both methadone and cocaine were stable during sample storage, preparation, and analytical procedures. The method was successfully used to analyze biological samples obtained from a drug- drug interaction pharmacokinetics (PK) study conducted in rats to investigate the effect of methadone on cocaine PK. Our method not only can be used for bioanalysis of samples obtained from rats but also can potentially be applied to human biological serum samples to monitor compliance to methadone maintenance therapy (MMT) and to detect possible cocaine-methadone co-abuse. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Intestinal growth and function of broiler chicks fed sorghum based ...

    African Journals Online (AJOL)

    Margret Rukuni

    Weekly measurements of body weight (LW) and feed intake (FI) for each replicate of the treatments were taken. At days 7, 14, 21, 42 and 56 two chicks from each replicate were killed after starving them for 12 hours to limit intestinal throughput. To ensure clean histology samples, the liver and intestinal samples were ...

  7. Gastric and intestinal surgery.

    Science.gov (United States)

    Fossum, Theresa W; Hedlund, Cheryl S

    2003-09-01

    Gastric surgery is commonly performed to remove foreign bodies and correct gastric dilatation-volvulus and is less commonly performed to treat gastric ulceration or erosion, neoplasia, and benign gastric outflow obstruction. Intestinal surgery, although commonly performed by veterinarians, should never be considered routine. The most common procedures of the small intestinal tract performed in dogs and cats include enterotomy and resection/anastomosis. Surgery of the large intestine is indicated for lesions causing obstruction, perforations, colonic inertia, or chronic inflammation.

  8. Development of sheep primordial follicles encapsulated in alginate or in ovarian tissue in fresh and vitrified samples.

    Science.gov (United States)

    Sadeghnia, Samaneh; Akhondi, Mohammad Mehdi; Hossein, Ghamartaj; Mobini, Sahba; Hosseini, Laleh; Naderi, Mohammad Mehdi; Boroujeni, Sara Borjian; Sarvari, Ali; Behzadi, Bahareh; Shirazi, Abolfazl

    2016-04-01

    In vitro follicle growth is a promising strategy for female fertility preservation. This study was conducted to compare the development of ovine follicles either isolated or in the context of ovarian cortical pieces after short term (8 days) three-dimensional culture in fresh and vitrified samples. Four different experiments were conducted; I) culture of ovarian cortical pieces encapsulated in 0.5% and 1% alginate and without alginate encapsulation (CP-0.5%, CP-1% and CP, respectively), II) culture of isolated primordial and primary follicles encapsulated in 1% and 2% alginate (IF-1% and IF-2%, respectively), III) culture of fresh and vitrified-warmed cortical pieces (F-CP and Vit-CP, respectively), and IV) culture of fresh and vitrified-warmed encapsulated isolated follicles (F-IF and Vit-IF, respectively). The number of secondary follicles after culture was negatively influenced by encapsulation of ovarian cortical pieces (6.3 ± 3.3 and 10.6 ± 0.9 vs 21.5 ± 2.3 in CP-0.5% and CP-1% vs CP, respectively). The diameter of follicles in IF-2% was higher than IF-1% (54.06 ± 2 vs 41.9 ± 1.5) and no significant difference in follicular viability was observed between the two groups. The proportions of different follicular types and their viability after culture in vitrified-warmed cortical pieces were comparable with fresh ones. The viability of vitrified-warmed isolated follicles was lower than fresh counterparts. The growth rate of fresh follicles was higher than vitrified-warmed follicles after culture (47.9 ± 1 vs 44.6 ± 1). In conclusion, while encapsulation of ovarian cortical pieces decreased the follicles' development, it could better support the growth of isolated follicles. Moreover, the viability and growth rate of isolated-encapsulated follicles was decreased by vitrification. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Intestinal parasites and tuberculosis

    Directory of Open Access Journals (Sweden)

    Anuar Alonso Cedeño-Burbano

    2017-10-01

    Conclusions: The available evidence was insufficient to affirm that intestinal parasites predispose to developing tuberculous. The studies carried out so far have found statistically insignificant results.

  10. Extra-intestinal calcium handling contributes to normal serum calcium levels when intestinal calcium absorption is suboptimal.

    Science.gov (United States)

    Lieben, Liesbet; Verlinden, Lieve; Masuyama, Ritsuko; Torrekens, Sophie; Moermans, Karen; Schoonjans, Luc; Carmeliet, Peter; Carmeliet, Geert

    2015-12-01

    The active form of vitamin D, 1,25(OH)2D, is a crucial regulator of calcium homeostasis, especially through stimulation of intestinal calcium transport. Lack of intestinal vitamin D receptor (VDR) signaling does however not result in hypocalcemia, because the increased 1,25(OH)2D levels stimulate calcium handling in extra-intestinal tissues. Systemic VDR deficiency, on the other hand, results in hypocalcemia because calcium handling is impaired not only in the intestine, but also in kidney and bone. It remains however unclear whether low intestinal VDR activity, as observed during aging, is sufficient for intestinal calcium transport and for mineral and bone homeostasis. To this end, we generated mice that expressed the Vdr exclusively in the gut, but at reduced levels. We found that ~15% of intestinal VDR expression greatly prevented the Vdr null phenotype in young-adult mice, including the severe hypocalcemia. Serum calcium levels were, however, in the low-normal range, which may be due to the suboptimal intestinal calcium absorption, renal calcium loss, insufficient increase in bone resorption and normal calcium incorporation in the bone matrix. In conclusion, our results indicate that low intestinal VDR levels improve intestinal calcium absorption compared to Vdr null mice, but also show that 1,25(OH)2D-mediated fine-tuning of renal calcium reabsorption and bone mineralization and resorption is required to maintain fully normal serum calcium levels. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Enteric glial cells and their role in the intestinal epithelial barrier

    OpenAIRE

    Yu, Yan-Bo; Li, Yan-Qing

    2014-01-01

    The intestinal epithelium constitutes a physical and functional barrier between the external environment and the host organism. It is formed by a continuous monolayer of intestinal epithelial cells maintained together by intercellular junctional complex, limiting access of pathogens, toxins and xenobiotics to host tissues. Once this barrier integrity is disrupted, inflammatory disorders and tissue injury are initiated and perpetuated. Beneath the intestinal epithelial cells lies a population ...

  12. adhesive intestinal obstruction

    African Journals Online (AJOL)

    2006-06-01

    Jun 1, 2006 ... ABSTRACT. Background: Adhesions after abdominal and pelvic surgery are a major cause of intestinal obstruction in the western world and the pathology is steadily gaining prominence in our practice. Objective: To determine the magnitude of adhesive intestinal obstruction; to determine the types.

  13. A Method to Correlate mRNA Expression Datasets Obtained from Fresh Frozen and Formalin-Fixed, Paraffin-Embedded Tissue Samples: A Matter of Thresholds.

    Directory of Open Access Journals (Sweden)

    Dana A M Mustafa

    Full Text Available Gene expression profiling of tumors is a successful tool for the discovery of new cancer biomarkers and potential targets for the development of new therapeutic strategies. Reliable profiling is preferably performed on fresh frozen (FF tissues in which the quality of nucleic acids is better preserved than in formalin-fixed paraffin-embedded (FFPE material. However, since snap-freezing of biopsy materials is often not part of daily routine in pathology laboratories, one may have to rely on archival FFPE material. Procedures to retrieve the RNAs from FFPE materials have been developed and therefore, datasets obtained from FFPE and FF materials need to be made compatible to ensure reliable comparisons are possible.To develop an efficient method to compare gene expression profiles obtained from FFPE and FF samples using the same platform.Twenty-six FFPE-FF sample pairs of the same tumors representing various cancer types, and two FFPE-FF sample pairs of breast cancer cell lines, were included. Total RNA was extracted and gene expression profiling was carried out using Illumina's Whole-Genome cDNA-mediated Annealing, Selection, extension and Ligation (WG-DASL V3 arrays, enabling the simultaneous detection of 24,526 mRNA transcripts. A sample exclusion criterion was created based on the expression of 11 stably expressed reference genes. Pearson correlation at the probe level was calculated for paired FFPE-FF, and three cut-off values were chosen. Spearman correlation coefficients between the matched FFPE and FF samples were calculated for three probe lists with varying levels of significance and compared to the correlation based on all measured probes. Unsupervised hierarchical cluster analysis was performed to verify performance of the included probe lists to compare matched FPPE-FF samples.Twenty-seven FFPE-FF pairs passed the sample exclusion criterion. From the profiles of 27 FFPE and FF matched samples, the best correlating probes were identified

  14. Intestinal Barrier and Behavior.

    Science.gov (United States)

    Julio-Pieper, M; Bravo, J A

    2016-01-01

    The intestinal barrier function contributes to gut homeostasis by modulating absorption of water, electrolytes, and nutrients from the lumen into the circulation while restricting the passage of noxious luminal substances and microorganisms. Chronic conditions such as rheumatoid arthritis, inflammatory bowel disease, and celiac disease are associated to intestinal barrier dysfunction. Here, the hypothesis is that a leaky intestinal wall allowing for indiscriminate passage of intraluminal compounds to the vascular compartment could in turn lead to systemic inflammation. An increasing number of studies are now investigating the association between gut permeability and CNS disorders, under the premise that translocation of intestinal luminal contents could affect CNS function, either directly or indirectly. Still, it is unknown whether disruption of intestinal barrier is a causative agent or a consequence in these situations. Here, we discuss the latest evidence pointing to an association between increased gut permeability and disrupted behavioral responses. © 2016 Elsevier Inc. All rights reserved.

  15. CDP-choline reduces severity of intestinal injury in a neonatal rat model of necrotizing enterocolitis.

    Science.gov (United States)

    Cetinkaya, Merih; Cansev, Mehmet; Cekmez, Ferhat; Tayman, Cuneyt; Canpolat, Fuat Emre; Kafa, Ilker M; Uysal, Sema; Tunc, Turan; Sarici, S Umit

    2013-07-01

    Cytidine 5'-diphosphocholine (CDP-choline) is an endogenous intermediate in the biosynthesis of phosphatidylcholine, a contributor to the mucosal defense of the intestine. The aim of this study was to evaluate the possible cytoprotective effect of CDP-choline treatment on intestinal cell damage, membrane phospholipid content, inflammation, and apoptosis in a neonatal rat model of necrotizing enterocolitis (NEC). We divided a total of 30 newborn pups into three groups: control, NEC, and NEC + CDP-choline. We induced NEC by enteral formula feeding, exposure to hypoxia-hyperoxia, and cold stress. We administered CDP-choline intraperitoneally at 300 mg/kg/d for 3 d starting from the first day of life. We evaluated apoptosis macroscopically and histopathologically in combination with proinflammatory cytokines in the gut samples. Moreover, we determined membrane phospholipid levels as well as activities of xanthine oxidase, superoxide dismutase, glutathione peroxidase, and myeloperoxidase enzymes and the malondialdehyde content of intestinal tissue. Mean clinical sickness score, macroscopic gut assessment score, and intestinal injury score were significantly improved, whereas mean apoptosis score and caspase-3 levels were significantly reduced in pups in the NEC + CDP-choline group compared with the NEC group. Tissue proinflammatory cytokine (interleukin-1β, interleukin-6, and tumor necrosis factor-α) levels as well as tissue malondialdehyde content and myeloperoxidase activities were reduced, whereas glutathione peroxidase and superoxide dismutase activities were preserved in the NEC + CDP-choline group. In addition, NEC damage reduced intestinal tissue membrane phospholipids, whereas CDP-choline significantly enhanced total phospholipid and phosphatidylcholine levels. Long-term follow-up in additional experiments revealed increased body weight, decreased clinical sickness scores, and enhanced survival in CDP-choline-receiving versus saline-receiving pups with NEC

  16. Usefulness of pulse-wave doppler tissue sampling and dobutamine stress echocardiography for identification of false positive inferior wall defects in SPECT

    International Nuclear Information System (INIS)

    Altinmakas, S.; Dagdeviren, B.; Turkmen, M.; Gursurer, M.; Say, B.; Tezel, T.; Ersek, B.

    2000-01-01

    False positive inferior wall perfusion defects restrict the accuracy of SPECT in diagnosis of coronary artery disease (CAD). Pulse-Wave Tissue Doppler (PWTD) has been recently proposed to assess regional wall motion velocities. The objectives of this study were to evaluate the presence of CAD by using PWTD during dobutamine stress echocardiography (DSE) in patients with an inferior perfusion defect detected by SPECT and compare PWTD parameters of normal cases with patients who had inferior perfusion defect and CAD. Sixty-five patients (mean age 58±8 years, 30 men) with a normal LV systolic function at rest according to echocardiographic evaluation with an inferior ischemia determined by SPECT and a control group (CG) of 34 normal cases (mean age 56±7 years, 16 men) were included in this study. All patients underwent a standard DSE (up to 40 μg/kg/min with additional atropine during sub-maximum heart rate responses). Pulse-wave Doppler tissue sampling of inferior wall was performed in the apical 2-chamber view at rest and stress. The coronary angiography was performed within 24 hours. The results were evaluated for the prediction of significant right coronary artery (RCA) and/or left circumflex coronary artery (CX) with narrowing (≥50% diameter stenosis, assessed by quantitative coronary angiography). It was observed that the peak stress mean E/A ratio was lower in patients with CAD when compared to patients without CAD (0.78±0.2 versus 1.29±0.11 p<0.0001). Also the peak stress E/A ratio of normal cases was significantly higher than patients who had CAD (1.19±0.3 versus 0.78±0.2 p<0.0001). When the cut off point for the E/A ratio was determined as 1, the sensitivity and specificity of dobutamine stress PWTD E/A were 89% and 86%, respectively. The peak stress E/A ratio was higher than 1 in all patients with a false positive perfusion defect. Systolic S velocity increase during DSE was significantly lower in patients with CAD (54%±17 versus 99%±24 p=0

  17. Promoter Region Hypermethylation and mRNA Expression of MGMT and p16 Genes in Tissue and Blood Samples of Human Premalignant Oral Lesions and Oral Squamous Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Vikram Bhatia

    2014-01-01

    Full Text Available Promoter methylation and relative gene expression of O6-methyguanine-DNA-methyltransferase (MGMT and p16 genes were examined in tissue and blood samples of patients with premalignant oral lesions (PMOLs and oral squamous cell carcinoma (OSCC. Methylation-specific PCR and reverse transcriptase PCR were performed in 146 tissue and blood samples from controls and patients with PMOLs and OSCC. In PMOL group, significant promoter methylation of MGMT and p16 genes was observed in 59% (P=0.0010 and 57% (P=0.0016 of tissue samples, respectively, and 39% (P=0.0135 and 33% (P=0.0074 of blood samples, respectively. Promoter methylation of both genes was more frequent in patients with OSCC, that is, 76% (P=0.0001 and 82% (P=0.0001 in tissue and 57% (P=0.0002 and 70% (P=0.0001 in blood, respectively. Significant downregulation of MGMT and p16 mRNA expression was observed in both tissue and blood samples from patients with PMOLs and OSCC. Hypermethylation-induced transcriptional silencing of MGMT and p16 genes in both precancer and cancer suggests important role of these changes in progression of premalignant state to malignancy. Results support use of blood as potential surrogate to tissue samples for screening or diagnosing PMOLs and early OSCC.

  18. DNA Sampling Hook

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The DNA Sampling Hook is a significant improvement on a method of obtaining a tissue sample from a live fish in situ from an aquatic environment. A tissue sample...

  19. Scintigraphic visualization of bacterial translocation in experimental strangulated intestinal obstruction

    International Nuclear Information System (INIS)

    Galeev, Yu.M.; Popov, M.V.; Salato, O.V.; Lishmanov, Yu.B.; Grigorev, E.G.; Aparcin, K.A.

    2009-01-01

    The purpose of this study was to obtain scintigraphic images depicting translocation of 99m Tc-labelled Escherichia coli bacteria through the intestinal barrier and to quantify this process using methods of nuclear medicine. Thirty male Wistar rats (including 20 rats with modelled strangulated intestinal obstruction and 10 healthy rats) were used for bacterial scintigraphy. 99m Tc-labelled E. coli bacteria ( 99m Ts-E. coli) with an activity of 7.4-11.1 MBq were administered into a section of the small intestine. Scintigraphic visualization of bacterial translocation into organs and tissues of laboratory animals was recorded in dynamic (240 min) and static (15 min) modes. The number of labelled bacteria, which migrated through the intestinal barrier, was quantified by calculating the translocation index (TI). Control indicated no translocation of 99m Ts-E. coli administered into the intestine through the parietes of the small intestine's distal part in healthy animals. Animals with strangulated obstruction demonstrated different migration strength and routes of labelled bacteria from strangulated and superior to strangulation sections of the small intestine. 99m Ts-E. coli migrated from the strangulated loop into the peritoneal cavity later causing systemic bacteraemia through peritoneal resorption. The section of the small intestine, which was superior to the strangulation, demonstrated migration of labelled bacteria first into the portal and then into the systemic circulation. The strangulated section of the small intestine was the main source of bacteria dissemination since the number of labelled bacteria, which migrated from this section significantly, exceeded that of the area superior to the strangulation section of the small intestine (p = 0.0003). Bacterial scintigraphy demonstrated the possibility of visualizing migration routes of labelled bacteria and quantifying their translocation through the intestinal barrier. This approach to study bacterial

  20. Mycotoxins and the intestine

    Directory of Open Access Journals (Sweden)

    Leon Broom

    2015-12-01

    Full Text Available Fungal biochemical pathways can yield various compounds that are not considered to be necessary for their growth and are thus referred to as secondary metabolites. These compounds have been found to have wide ranging biological effects and include potent poisons (mycotoxins. Mycotoxins invariably contaminate crops and (thus animal feeds. The intestine is the key link between ingested mycotoxins and their detrimental effects on the animal. Effects on the intestine, or intestinal environment, and immune system have been reported with various mycotoxins. These effects are almost certainly occurring across species. Most, if not all, of the reported effects of mycotoxins are negative in terms of intestinal health, for example, decreased intestinal cell viability, reductions in short chain fatty acid (SCFA concentrations and elimination of beneficial bacteria, increased expression of genes involved in promoting inflammation and counteracting oxidative stress. This challenge to intestinal health will predispose the animal to intestinal (and systemic infections and impair efficient digestion and absorption of nutrients, with the associated effect on animal productivity.

  1. Molecular characterization of Staphylococcus aureus isolates from skin and soft tissue infections samples and healthy carriers in the Central Slovenia region.

    Science.gov (United States)

    Svent-Kucina, Natasa; Pirs, Mateja; Kofol, Romina; Blagus, Rok; Smrke, Dragica Maja; Bilban, Marjan; Seme, Katja

    2016-04-01

    Staphylococcus aureus is among the most important human pathogens. It is associated with different infections and is a major cause of skin and soft tissue infections (SSTIs). The aim of our study was to compare S. aureus isolates associated with SSTIs with isolates obtained from healthy carriers in the Central Slovenia region in terms of antimicrobial susceptibility, genetic diversity by clonal complex (CC)/sequence type, spa type, and by toxin gene profiling. In total, 274 S. aureus isolates were collected prospectively by culturing wound samples from 461 SSTI patients and nasal samples from 451 healthy carriers. We have demonstrated high heterogeneity in terms of CCs and spa type in both groups of isolates. The main clone among SSTI strains was Panton-Valentine leukocidin gene (pvl) positive CC121, whereas the main clone among carrier strains was CC45 carrying a large range of toxin genes. The main spa type in both groups was t091. Pvl was more frequently present in SSTI strains (31.2% SSTI vs 3.6% carrier strains) and staphylococcal enterotoxin C was more frequently present in carrier strains (1.6% SSTI vs 17.0% carrier strains). We have also demonstrated that methicillin-resistant S. aureus was a rare cause (2.8%) of SSTIs in our region. © 2016 APMIS. Published by John Wiley & Sons Ltd.

  2. [Chronic gastritis and intestinal metaplasia].

    Science.gov (United States)

    Castillo, T; Navarrete, J; Celestina, A

    1989-01-01

    Much has been written about gastric mucosae behavior and the occurrence of intestinal metaplasia. The aim of this paper is to learn something more about these matters in peruvian population. We selected 100 patients with endoscopically no localized lesions between 30 to 70 years of age. We took 8 samples of gastric mucosae in each patient which were carefully examined for the presence of inflammatory changes, settle the line type between antral and fundic mucosae and the frequency of intestinal metaplasia finding. The results showed disagreement between endoscopic and histological findings, so we conclude it is better to diagnose chronic gastritis on the basis of histological parameters. The line between antral and fundic mucosae was of the close type one found in 87% of all cases and it advanced proximally with increasing age. Intestinal metaplasia was present in 46% of the whole number of patients and the rate of occurrence increased in 50% over 50 years age. These findings will let us compare future investigations of gastric mucosae behavior with localized benign or malign lesions.

  3. Macrophages in intestinal homeostasis and inflammation

    Science.gov (United States)

    Bain, Calum C; Mowat, Allan McI

    2014-01-01

    The intestine contains the largest pool of macrophages in the body which are essential for maintaining mucosal homeostasis in the face of the microbiota and the constant need for epithelial renewal but are also important components of protective immunity and are involved in the pathology of inflammatory bowel disease (IBD). However, defining the biological roles of intestinal macrophages has been impeded by problems in defining the phenotype and origins of different populations of myeloid cells in the mucosa. Here, we discuss how multiple parameters can be used in combination to discriminate between functionally distinct myeloid cells and discuss the roles of macrophages during homeostasis and how these may change when inflammation ensues. We also discuss the evidence that intestinal macrophages do not fit the current paradigm that tissue-resident macrophages are derived from embryonic precursors that self-renew in situ, but require constant replenishment by blood monocytes. We describe our recent work demonstrating that classical monocytes constantly enter the intestinal mucosa and how the environment dictates their subsequent fate. We believe that understanding the factors that drive intestinal macrophage development in the steady state and how these may change in response to pathogens or inflammation could provide important insights into the treatment of IBD. PMID:24942685

  4. Glutamine and Its Effects on the Intestine

    Directory of Open Access Journals (Sweden)

    Paul E Hardy

    1991-01-01

    Full Text Available Glutamine, an amino acid, is the principal energy substrate for small intestinal cells. It also acts as a nitrogen carrier through its amide nitrogen. Arterial glutamine is supported by net synthesis in skeletal muscle. Glutamine is rapidly metabolized by the intestine, whether supplied from the lumen or from the arterial circulation. Intestinal uptake of glutamine increases after trauma and operative stress. The consumption of glutamine by the gut may in large part be dependent on mucosal glutaminase activity and on enterocyte glutamine transport. Glutaminc has been shown to improve gut morphology and outcome in animal models of encerocolitis. It may play a similar role in aiding repair of human intestinal injury in persons with sufficient glutamine in their diet compared to those who arc glutamine deficient. Glutamine may have a positive effect on the immune function of the intestinal mucosal-associated lymphoid tissue. Glutamine is not currently available in nutritional preparations for routine clinical use, yet it has recently been shown to benefit maintenance of nitrogen balance in humans. Due to the instability and low solubility of glutamine, dipeptides have been studied. L-alanyl-L-glutamine seems to be the most promising glutamine precursor for parenteral use in humans, as it is safe and rapidly hydrolyzed in vivo to release free glutamine. The exact role of glutamine as a therapeutic agent to promote intetitinal well-being has yet to be determined. However, preliminary evidence suggests that glutaminc will be helpful in a variety of clinical scenarios.

  5. Elevation of HO-1 Expression Mitigates Intestinal Ischemia-Reperfusion Injury and Restores Tight Junction Function in a Rat Liver Transplantation Model

    Directory of Open Access Journals (Sweden)

    Xinjin Chi

    2015-01-01

    Full Text Available Aims. This study was aimed at investigating whether elevation of heme oxygenase-1 (HO-1 expression could lead to restoring intestinal tight junction (TJ function in a rat liver transplantation model. Methods. Intestinal mucosa injury was induced by orthotopic autologous liver transplantation (OALT on male Sprague-Dawley rats. Hemin (a potent HO-1 activator and zinc-protoporphyrin (ZnPP, a HO-1 competitive inhibitor, were separately administered in selected groups before OALT. The serum and intestinal mucosa samples were collected at 8 hours after the operation for analysis. Results. Hemin pretreatment significantly reduced the inflammation and oxidative stress in the mucosal tissue after OALT by elevating HO-1 protein expression, while ZnPP pretreatment aggravated the OALT mucosa injury. Meanwhile, the restriction on the expression of tight junction proteins zonula occludens-1 and occludin was removed after hemin pretreatment. These molecular events led to significant improvement on intestinal barrier function, which was proved to be through increasing nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2 and reducing nuclear translocation of nuclear factor kappa-B (NF-κB in intestinal injured mucosa. Summary. Our study demonstrated that elevation of HO-1 expression reduced the OALT-induced intestinal mucosa injury and TJ dysfunction. The HO-1 protective function was likely mediated through its effects of anti-inflammation and antioxidative stress.

  6. Elevation of HO-1 Expression Mitigates Intestinal Ischemia-Reperfusion Injury and Restores Tight Junction Function in a Rat Liver Transplantation Model

    Science.gov (United States)

    Chi, Xinjin; Yao, Weifeng; Xia, Hua; Jin, Yi; Li, Xi; Cai, Jun; Hei, Ziqing

    2015-01-01

    Aims. This study was aimed at investigating whether elevation of heme oxygenase-1 (HO-1) expression could lead to restoring intestinal tight junction (TJ) function in a rat liver transplantation model. Methods. Intestinal mucosa injury was induced by orthotopic autologous liver transplantation (OALT) on male Sprague-Dawley rats. Hemin (a potent HO-1 activator) and zinc-protoporphyrin (ZnPP, a HO-1 competitive inhibitor), were separately administered in selected groups before OALT. The serum and intestinal mucosa samples were collected at 8 hours after the operation for analysis. Results. Hemin pretreatment significantly reduced the inflammation and oxidative stress in the mucosal tissue after OALT by elevating HO-1 protein expression, while ZnPP pretreatment aggravated the OALT mucosa injury. Meanwhile, the restriction on the expression of tight junction proteins zonula occludens-1 and occludin was removed after hemin pretreatment. These molecular events led to significant improvement on intestinal barrier function, which was proved to be through increasing nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and reducing nuclear translocation of nuclear factor kappa-B (NF-κB) in intestinal injured mucosa. Summary. Our study demonstrated that elevation of HO-1 expression reduced the OALT-induced intestinal mucosa injury and TJ dysfunction. The HO-1 protective function was likely mediated through its effects of anti-inflammation and antioxidative stress. PMID:26064429

  7. Transient inhibition of ROR-γt therapeutically limits intestinal inflammation by reducing TH17 cells and preserving group 3 innate lymphoid cells.

    Science.gov (United States)

    Withers, David R; Hepworth, Matthew R; Wang, Xinxin; Mackley, Emma C; Halford, Emily E; Dutton, Emma E; Marriott, Clare L; Brucklacher-Waldert, Verena; Veldhoen, Marc; Kelsen, Judith; Baldassano, Robert N; Sonnenberg, Gregory F

    2016-03-01

    RAR-related orphan receptor-γt (ROR-γt) directs differentiation of proinflammatory T helper 17 (TH17) cells and is a potential therapeutic target in chronic autoimmune and inflammatory diseases. However, ROR-γt-dependent group 3 innate lymphoid cells ILC3s provide essential immunity and tissue protection in the intestine, suggesting that targeting ROR-γt could also result in impaired host defense after infection or enhanced tissue damage. Here, we demonstrate that transient chemical inhibition of ROR-γt in mice selectively reduces cytokine production from TH17 but not ILCs in the context of intestinal infection with Citrobacter rodentium, resulting in preserved innate immunity. Temporal deletion of Rorc (encoding ROR-γt) in mature ILCs also did not impair cytokine response in the steady state or during infection. Finally, pharmacologic inhibition of ROR-γt provided therapeutic benefit in mouse models of intestinal inflammation and reduced the frequency of TH17 cells but not ILCs isolated from primary intestinal samples of individuals with inflammatory bowel disease (IBD). Collectively, these results reveal differential requirements for ROR-γt in the maintenance of TH17 cell and ILC3 responses and suggest that transient inhibition of ROR-γt is a safe and effective therapeutic approach during intestinal inflammation.

  8. Intestinal solute carriers

    DEFF Research Database (Denmark)

    Steffansen, Bente; Nielsen, Carsten Uhd; Brodin, Birger

    2004-01-01

    A large amount of absorptive intestinal membrane transporters play an important part in absorption and distribution of several nutrients, drugs and prodrugs. The present paper gives a general overview on intestinal solute carriers as well as on trends and strategies for targeting drugs and...... membrane transporters in the small intestine in order to increase oral bioavailabilities of drug or prodrug, the major influence on in vivo pharmacokinetics is suggested to be dose-dependent increase in bioavailability as well as prolonged blood circulation due to large capacity facilitated absorption...

  9. Intestinal solute carriers

    DEFF Research Database (Denmark)

    Steffansen, Bente; Nielsen, Carsten Uhd; Brodin, Birger

    2004-01-01

    membrane transporters in the small intestine in order to increase oral bioavailabilities of drug or prodrug, the major influence on in vivo pharmacokinetics is suggested to be dose-dependent increase in bioavailability as well as prolonged blood circulation due to large capacity facilitated absorption......A large amount of absorptive intestinal membrane transporters play an important part in absorption and distribution of several nutrients, drugs and prodrugs. The present paper gives a general overview on intestinal solute carriers as well as on trends and strategies for targeting drugs and...

  10. Wnt signaling in the intestinal epithelium: from endoderm to cancer.

    NARCIS (Netherlands)

    Gregorieff, A.; Clevers, J.C.

    2005-01-01

    The Wnt pathway controls cell fate during embryonic development. It also persists as a key regulator of homeostasis in adult self-renewing tissues. In these tissues, mutational deregulation of the Wnt cascade is closely associated with malignant transformation. The intestinal epithelium represents

  11. Intestinal response to myeloablative chemotherapy in piglets

    DEFF Research Database (Denmark)

    Pontoppidan, Peter Erik Lotko; Shen, René Liang; Petersen, Bodil L

    2014-01-01

    /kg) and bone marrow was collected on day 11. Histology of bone marrow samples showed total aplasia after treatment A. Using this treatment in study 2, Bu-Cy pigs showed lowered spleen and intestinal weights and variable clinical signs of dehydration, sepsis, and pneumonia at tissue collection. Oral mucositis......, bilirubin, alkaline phosphatase), except that blood iron levels were higher in Bu-Cy pigs. We conclude that a myeloablative Bu-Cy regimen to piglets results in clinical signs comparable to those seen in pediatric patients subjected to myeloablative treatment prior to HSCT. Piglets may be used as a model...... investigated in three-day-old pigs (Landrace × Yorkshire × Duroc, n = 6). Pigs were given one of three different dose combinations of Bu and Cy (A: 4 days Bu, 2 × 1.6 mg/kg plus 2 days Cy, 60 mg/kg; B: 4 days Bu, 2 × 0.8 mg/kg plus 2 days Cy, 30 mg/kg; C: 2 days Bu at 2 × 1.6 mg/kg plus 1 day Cy, 60 mg...

  12. Trace element concentration distributions in breast, lung and colon tissues

    Energy Technology Data Exchange (ETDEWEB)

    Majewska, Urszula [Institute of Physics, Swietokrzyska Academy, Swietokrzyska 15, 25-406 Kielce (Poland); Banas, Dariusz [Institute of Physics, Swietokrzyska Academy, Swietokrzyska 15, 25-406 Kielce (Poland); Braziewicz, Janusz [Institute of Physics, Swietokrzyska Academy, Swietokrzyska 15, 25-406 Kielce (Poland); Gozdz, Stanislaw [Holycross Cancer Centre, Artwinskiego 3, 25-734 Kielce (Poland); Kubala-Kukus, Aldona [Institute of Physics, Swietokrzyska Academy, Swietokrzyska 15, 25-406 Kielce (Poland); Kucharzewski, Marek [Department of General Surgery, Silesian Medical Academy, Batorego 15, 41-902 Bytom (Poland)

    2007-07-07

    The concentrations of Fe, Cu, Zn and Se in cancerous and benign tissues of breast, lung and intestine (colon) have been determined. In the cases when the element concentration has not been determined in all samples the Kaplan-Meier method has been used for the reconstruction of the original concentration distributions and estimation of the true mean concentrations and medians. Finally, the log-rank test has been applied to compare the elemental concentration distributions between cancerous and benign tissues of the same organ, between cancerous tissues and between benign tissues taken from different organs. Comparing benign and malignant neoplastic tissues, statistically significant differences have been found between Fe and Se concentration distributions of breast as well as for Cu and Zn in the case of lung tissues and in the case of colon tissues for Zn. The concentrations of all elements have been found to be statistically different in cancer tissues as well as in benign ones when comparing the different organs, i.e. groups 'breast-colon' and 'breast-lung'. Concentrations of Fe and Cu have been found to be statistically different in lung and colon cancerous tissues. For benign tissues of lung and colon a statistically significant difference has been found only for Zn.

  13. Protective effect of salvianolic acid B against intestinal ischemia ...

    African Journals Online (AJOL)

    Keywords: Salvianolic acid B, Intestinal Ischemia-reperfusion, Antioxidants, Inflammation, Intestinal ... Rats were sacrificed by cervical decapitation under pentabarbotal sodium at the dose of 50 mg/kg via i.p after IIRI induction. Blood samples were collected in a heparinized .... lipid peroxidation [17] and hence the levels of.

  14. STUDY ON INTESTINAL PROTOZOA IN SEVEN VILLAGES OF BANDARABASS

    OpenAIRE

    J.Sheiban; M.Rezaian

    1981-01-01

    A survey was carried out in seven villages around Bandarabass, on Persion Gulf, to determine the prevalence of the intestinal protozoa infections. Out of the 835 stool specimens examined, 676 samples were positive with single and multiple infections. The most prevalent of intestinal protozoa in Bandarabass were Entamoeba histolytica, Entamoeba coli, Giardia lambia and Iodamoeba butschlii.

  15. Asymptomatic intestinal protozoa in school age children in Pategi ...

    African Journals Online (AJOL)

    Asymptomatic intestinal protozoa in school age children in Pategi, Pategi LGA of Kwara state, Nigeria. ... Results: One hundred and ninety seven (26.3%) of the samples were positive for intestinal protozoan parasite. The distribution of the parasites ... Key words: Asymptomatic, amoebiasis, giardiasis, rural area, children.

  16. (13)Carbon and (15)nitrogen isotopes in autopsy liver tissue samples from Greenlandic Inuit and Danes: consumption of marine versus terrestrial food

    DEFF Research Database (Denmark)

    Milman, N.; Laursen, J.; Mulvad, G.

    2010-01-01

    in liver tissue from Greenlandic Inuit and Danes. Subjects/Methods: Normal liver tissue was obtained at autopsy in 1992-1994 from 60 Inuit with a median age of 61 years (range 25-83) and in 1986 from 15 ethnic Danes with a median age of 84 years (range 66-93). By sieving, liver tissue was separated...

  17. Tissue versus mechanical valve replacement: Short term outcome among a sample of Egyptian patients with rheumatic mitral valve disease in Minia Governorate

    Directory of Open Access Journals (Sweden)

    Faisal A. Mourad

    2016-12-01

    Conclusion: Tissue mitral valve offers excellent early postoperative results and less complication rate than mechanical mitral valve. The EOA is significantly bigger in the tissue mitral valve in sizes 27–29 thus offering less patient prosthesis mismatch. Tissue valves are suitable for populations with lower socioeconomic status as Minia Governorate.

  18. Innovative methods to study human intestinal drug metabolism in vitro : Precision-cut slices compared with Ussing chamber preparations

    NARCIS (Netherlands)

    van de Kerkhof, Esther G.; Ungell, Anna-Lena B.; Sjoberg, Asa K.; de Jager, Marina H.; Hilgendorf, Constanze; de Graaf, Inge A. M.; Groothuis, Geny M. M.

    2006-01-01

    Predictive in vitro methods to investigate drug metabolism in the human intestine using intact tissue are of high importance. Therefore, we studied the metabolic activity of human small intestinal and colon slices and compared it with the metabolic activity of the same human intestinal segments

  19. Ectopic intestinal glands after segmental small bowel irradiation in the cat

    International Nuclear Information System (INIS)

    Rubio, C.A.; Eriksson, B.; Johnsson, L.

    1983-01-01

    Following segmental irradiation of the small bowel, 5 of 64 cats demonstrated ectopic intestinal glands in the submucosal tissue. In addition, one of these 5 cats had foci of abnormal glands in the muscularis mucosae. In 2 of the 5 animals, cellular polymorphism, nucleolar irregularity and loss of cellular polarity were present in irradiation-induced ectopic intestinal glands. The review of the literature indicates that intestinal irradiation may induce intestinal adenocarcinomas with metastatic growth. The possibility that ectopic intestinal glands are precancerous lesions in the irradiated cat is discussed. (Auth.)

  20. Multifunctions of dietary polyphenols in the regulation of intestinal inflammation

    Directory of Open Access Journals (Sweden)

    Makoto Shimizu

    2017-01-01

    Full Text Available Food for specified health use is a type of functional food approved by the Japanese government, with more than 1250 products in 10 health-claim categories being approved as of April 2016. Polyphenols are currently used as functional ingredients in seven of the 10 categories. Although they have not yet been used for the food-for-specified-health-use category of “gut health promotion,” polyphenols are expected to contribute to the future development of gut-modulating food. Intestinal functions include digestion/absorption, acting as a barrier, recognition of external factors, and signal transduction. Owing to incessant exposure to external stress factors including food substances, bacteria, and environmental chemicals, intestines are always inflammatory to some extent, which may cause damage to and dysfunction of intestinal tissues depending on the situation. We identified food factors that could suppress immoderate inflammation in the intestines. In addition to certain amino acids and peptides, polyphenols such as chlorogenic acid and isoflavones were found to suppress inflammation in intestinal cells. Intestinal inflammation is caused by various factors in diverse mechanisms. Recent studies revealed that activation of pattern recognition receptors, such as Toll-like receptors and nucleotide-binding oligomerization domain proteins, in epithelial cells triggers intestinal inflammation. Intracellular receptors or signaling molecules controlling the intestinal detoxification system are also involved in the regulation of inflammation. Differentiation of regulatory T cells by activating a transcription factor Foxp-3 is known to suppress intestinal inflammation. A variety of phytochemicals including polyphenols modulate these receptors and signaling molecules, and are thus anti-inflammatory. Polyphenols affect epigenetic changes occurring in intestinal tissues by interacting with the enzymes responsible for DNA methylation and histone acetylation

  1. Enteric Virome Sensing-Its Role in Intestinal Homeostasis and Immunity.

    Science.gov (United States)

    Metzger, Rebecca N; Krug, Anne B; Eisenächer, Katharina

    2018-03-23

    Pattern recognition receptors (PRRs) sensing commensal microorganisms in the intestine induce tightly controlled tonic signaling in the intestinal mucosa, which is required to maintain intestinal barrier integrity and immune homeostasis. At the same time, PRR signaling pathways rapidly trigger the innate immune defense against invasive pathogens in the intestine. Intestinal epithelial cells and mononuclear phagocytes in the intestine and the gut-associated lymphoid tissues are critically involved in sensing components of the microbiome and regulating immune responses in the intestine to sustain immune tolerance against harmless antigens and to prevent inflammation. These processes have been mostly investigated in the context of the bacterial components of the microbiome so far. The impact of viruses residing in the intestine and the virus sensors, which are activated by these enteric viruses, on intestinal homeostasis and inflammation is just beginning to be unraveled. In this review, we will summarize recent findings indicating an important role of the enteric virome for intestinal homeostasis as well as pathology when the immune system fails to control the enteric virome. We will provide an overview of the virus sensors and signaling pathways, operative in the intestine and the mononuclear phagocyte subsets, which can sense viruses and shape the intestinal immune response. We will discuss how these might interact with resident enteric viruses directly or in context with the bacterial microbiome to affect intestinal homeostasis.

  2. Enteric Virome Sensing—Its Role in Intestinal Homeostasis and Immunity

    Directory of Open Access Journals (Sweden)

    Rebecca N. Metzger

    2018-03-01

    Full Text Available Pattern recognition receptors (PRRs sensing commensal microorganisms in the intestine induce tightly controlled tonic signaling in the intestinal mucosa, which is required to maintain intestinal barrier integrity and immune homeostasis. At the same time, PRR signaling pathways rapidly trigger the innate immune defense against invasive pathogens in the intestine. Intestinal epithelial cells and mononuclear phagocytes in the intestine and the gut-associated lymphoid tissues are critically involved in sensing components of the microbiome and regulating immune responses in the intestine to sustain immune tolerance against harmless antigens and to prevent inflammation. These processes have been mostly investigated in the context of the bacterial components of the microbiome so far. The impact of viruses residing in the intestine and the virus sensors, which are activated by these enteric viruses, on intestinal homeostasis and inflammation is just beginning to be unraveled. In this review, we will summarize recent findings indicating an important role of the enteric virome for intestinal homeostasis as well as pathology when the immune system fails to control the enteric virome. We will provide an overview of the virus sensors and signaling pathways, operative in the intestine and the mononuclear phagocyte subsets, which can sense viruses and shape the intestinal immune response. We will discuss how these might interact with resident enteric viruses directly or in context with the bacterial microbiome to affect intestinal homeostasis.

  3. Intestinal parasitic infections in renal transplant recipients.

    Science.gov (United States)

    Azami, Mehdi; Sharifi, Mehran; Hejazi, Sayed Hossein; Tazhibi, Mehdi

    2010-01-01

    The impact of intestinal parasitic infection in renal transplant recipients requires careful consideration in the developing world. However, there have been very few studies addressing this issue in Iran. This study was conducted to determine the prevalence of intestinal parasitic infections in renal transplant recipients in Iran. Stool specimens from renal transplant recipients and control groups were obtained between June 2006 and January 2007. The samples screened for intestinal parasitic infections using direct smear, formalin-ether sedimentation, Sheather's flotation and modified Ziehl-Neelsen staining methods. Out of 150 renal transplant recipients, 33.3% (50), and out of 225 control group, 20% (45) were infected with one or more type of intestinal parasites. The parasites detected among patients included Entamoeba coli (10.6%), Endolimax nana (8.7%), Giardia lamblia (7.4%), Blastocystis spp. (4.7%), Iodamoeba butschlii (0.7%), Chilomastix mesnili (0.7%) and Ascaris lumbricoides (0.7%). Multiple infections were more common among renal transplant recipients group (p < 0.05). This study highlights the importance of testing for intestinal parasites among Iranian renal transplant recipients. Routine examinations of stool samples for parasites would significantly benefit the renal transplant recipients by contributing to reduce severe infections.

  4. Intestinal parasitic infections in renal transplant recipients

    Directory of Open Access Journals (Sweden)

    Mehdi Azami

    Full Text Available The impact of intestinal parasitic infection in renal transplant recipients requires careful consideration in the developing world. However, there have been very few studies addressing this issue in Iran. This study was conducted to determine the prevalence of intestinal parasitic infections in renal transplant recipients in Iran. Stool specimens from renal transplant recipients and control groups were obtained between June 2006 and January 2007. The samples screened for intestinal parasitic infections using direct smear, formalin-ether sedimentation, Sheather's flotation and modified Ziehl-Neelsen staining methods. Out of 150 renal transplant recipients, 33.3% (50, and out of 225 control group, 20% (45 were infected with one or more type of intestinal parasites. The parasites detected among patients included Entamoeba coli (10.6%, Endolimax nana (8.7%, Giardia lamblia (7.4%, Blastocystis spp. (4.7%, Iodamoeba butschlii (0.7%, Chilomastix mesnili (0.7% and Ascaris lumbricoides (0.7%. Multiple infections were more common among renal transplant recipients group (p < 0.05. This study highlights the importance of testing for intestinal parasites among Iranian renal transplant recipients. Routine examinations of stool samples for parasites would significantly benefit the renal transplant recipients by contributing to reduce severe infections.

  5. Stem cell-based growth, regeneration, and remodeling of the planarian intestine

    Science.gov (United States)

    Forsthoefel, David J.; Park, Amanda E.; Newmark, Phillip A.

    2011-01-01

    Although some animals are capable of regenerating organs, the mechanisms by which this is achieved are poorly understood. In planarians, pluripotent somatic stem cells called neoblasts supply new cells for growth, replenish tissues in response to cellular turnover, and regenerate tissues after injury. For most tissues and organs, however, the spatiotemporal dynamics of stem cell differentiation and the fate of tissue that existed prior to injury have not been characterized systematically. Utilizing in vivo imaging and bromodeoxyuridine pulse-chase experiments, we have analyzed growth and regeneration of the planarian intestine, the organ responsible for digestion and nutrient distribution. During growth, we observe that new gut branches are added along the entire anteroposterior axis. We find that new enterocytes differentiate throughout the intestine rather than in specific growth zones, suggesting that branching morphogenesis is achieved primarily by remodeling of differentiated intestinal tissues. During regeneration, we also demonstrate a previously unappreciated degree of intestinal remodeling, in which pre-existing posterior gut tissue contributes extensively to the newly formed anterior gut, and vice versa. By contrast to growing animals, differentiation of new intestinal cells occurs at preferential locations, including within newly generated tissue (the blastema), and along pre-existing intestinal branches undergoing remodeling. Our results indicate that growth and regeneration of the planarian intestine are achieved by coordinated differentiation of stem cells and the remodeling of pre-existing tissues. Elucidation of the mechanisms by which these processes are integrated will be critical for understanding organogenesis in a post-embryonic context. PMID:21664348

  6. Repairing organs: lessons from intestine and liver

    OpenAIRE

    Gehart Helmuth; Clevers Hans

    2015-01-01

    The concept of organ regeneration has fascinated humanity from ancient mythology to modern science fiction. Recent advances offer the potential to soon bring such technology within the grasp of clinical medicine. Rapidly expanding insights into the intrinsic repair processes of the intestine and liver have uncovered significant plasticity in epithelial tissues. Harnessing this knowledge researchers have recently created culture systems that enable the expansion of stem cells into transplantab...

  7. Lymphoma Caused by Intestinal Microbiota

    Directory of Open Access Journals (Sweden)

    Mitsuko L. Yamamoto

    2014-09-01

    Full Text Available The intestinal microbiota and gut immune system must constantly communicate to maintain a balance between tolerance and activation: on the one hand, our immune system should protect us from pathogenic microbes and on the other hand, most of the millions of microbes in and on our body are innocuous symbionts and some can even be beneficial. Since there is such a close interaction between the immune system and the intestinal microbiota, it is not surprising that some lymphomas such as mucosal-associated lymphoid tissue (MALT lymphoma have been shown to be caused by the presence of certain bacteria. Animal models played an important role in establishing causation and mechanism of bacteria-induced MALT lymphoma. In this review we discuss different ways that animal models have been applied to establish a link between the gut microbiota and lymphoma and how animal models have helped to elucidate mechanisms of microbiota-induced lymphoma. While there are not a plethora of studies demonstrating a connection between microbiota and lymphoma development, we believe that animal models are a system which can be exploited in the future to enhance our understanding of causation and improve prognosis and treatment of lymphoma.

  8. Processes and procedures for a worldwide biological samples distribution; product assurance and logistic activities to support the mice drawer system tissue sharing event

    Science.gov (United States)

    Benassai, Mario; Cotronei, Vittorio

    The Mice Drawer System (MDS) is a scientific payload developed by the Italian Space Agency (ASI), it hosted 6 mice on the International Space Station (ISS) and re-entered on ground on November 28, 2009 with the STS 129 at KSC. Linked to the MDS experiment, a Tissue Sharing Program (TSP), was developed in order to make available to 16 Payload Investigators (PI) (located in USA, Canada, EU -Italy, Belgium and Germany -and Japan) the biological samples coming from the mice. ALTEC SpA (a PPP owned by ASI, TAS-I and local institutions) was responsible to support the logistics aspects of the MDS samples for the first MDS mission, in the frame of Italian Space Agency (ASI) OSMA program (OSteoporosis and Muscle Atrophy). The TSP resulted in a complex scenario, as ASI, progressively, extended the original OSMA Team also to researchers from other ASI programs and from other Agencies (ESA, NASA, JAXA). The science coordination was performed by the University of Genova (UNIGE). ALTEC has managed all the logistic process with the support of a specialized freight forwarder agent during the whole shipping operation phases. ALTEC formalized all the steps from the handover of samples by the dissection Team to the packaging and shipping process in a dedicated procedure. ALTEC approached all the work in a structured way, performing: A study of the aspects connected to international shipments of biological samples. A coopera-tive work with UNIGE/ASI /PIs to identify all the needs of the various researchers and their compatibility. A complete revision and integration of shipment requirements (addresses, tem-peratures, samples, materials and so on). A complete definition of the final shipment scenario in terms of boxes, content, refrigerant and requirements. A formal approach to identification and selection of the most suited and specialized Freight Forwarder. A clear identification of all the processes from sample dissection by PI Team, sample processing, freezing, tube preparation

  9. Oral pathology follow-up by means of micro-Raman spectroscopy on tissue and blood serum samples: an application of wavelet and multivariate data analysis

    Science.gov (United States)

    Delfino, I.; Camerlingo, C.; Zenone, F.; Perna, G.; Capozzi, V.; Cirillo, N.; Gaeta, G. M.; De Mol, E.; Lepore, M.

    2009-02-01

    Pemphigus vulgaris (PV) is a potentially fatal autoimmune disease that cause blistering of the skin and oral cavity. It is characterized by disruption of cell-cell adhesion within the suprabasal layers of epithelium, a phenomenon termed acantholysis Patients with PV develop IgG autoantibodies against normal constituents of the intercellular substance of keratinocytes. The mechanisms by which such autoantibodies induce blisters are not clearly understood. The qualitative analysis of such effects provides important clues in the search for a specific diagnosis, and the quantitative analysis of biochemical abnormalities is important in measuring the extent of the disease process, designing therapy and evaluating the efficacy of treatment. Improved diagnostic techniques could permit the recognition of more subtle forms of disease and reveal incipient lesions clinically unapparent, so that progression of potentially severe forms could be reversed with appropriate treatment. In this paper, we report the results of our micro-Raman spectroscopy study on tissue and blood serum samples from ill, recovered and under therapy PV patients. The complexity of the differences among their characteristic Raman spectra has required a specific strategy to obtain reliable information on the illness stage of the patients For this purpose, wavelet techniques and advanced multivariate analysis methods have been developed and applied to the experimental Raman spectra. Promising results have been obtained.

  10. Intestinal dendritic cells in the regulation of mucosal immunity

    DEFF Research Database (Denmark)

    Bekiaris, Vasileios; Persson, Emma K.; Agace, William Winston

    2014-01-01

    The intestine presents a huge surface area to the outside environment, a property that is of critical importance for its key functions in nutrient digestion, absorption, and waste disposal. As such, the intestine is constantly exposed to dietary and microbial-derived foreign antigens, to which im...... of the role these subsets play in the regulation of intestinal immune homeostasis and inflammation will help to define novel strategies for the treatment of intestinal pathologies and contribute to improved rational design of mucosal vaccines....... immune cells within the mucosa must suitably respond to maintain intestinal integrity, while also providing the ability to mount effective immune responses to potential pathogens. Dendritic cells (DCs) are sentinel immune cells that play a central role in the initiation and differentiation of adaptive...... immune responses. In the intestinal mucosa, DCs are located diffusely throughout the intestinal lamina propria, within gut-associated lymphoid tissues, including Peyer's patches and smaller lymphoid aggregates, as well as in intestinal-draining lymph nodes, including mesenteric lymph nodes...

  11. Related radiation effects on the intestine and their treatment

    International Nuclear Information System (INIS)

    Bardychev, M.S.; Kurpeshcheva, A.K.; Kaplan, M.A.

    1978-01-01

    Late radiation injuries of the intestine are frequent after radiation therapy of malignant tumours of female genitalia and some other tumours due to which the intestine gets into the irradiation field. On the basis of the analysis of 80 patients with late radiation injuries of intestine which developed at remote terms after radiation therapy of cervix uteri cancer and corpus uteri (65 patients) and other tumours, peculiarities of the clinical course and treatment of radiation enterocolitis, rectosigmoidites and rectites are discussed. In 39 patients these injuries were concomitant with late radiation injuries of the skin and subcutaneous soft tissues. The clinical course of radiation unjuries of the intestine was defined by the character of the pathological process in the intestine and was more sharply marked in patients suffering from radiation enterocolites. It was established that one of the pathogenetic mechanisms of late radiation injuries of the intestine was a disorder of the absorption function of the intestine. Local treatment of radiation injuries of the intestine should be combined with a general one the important component of which is a parenteral diet

  12. Protein synthesis and intestinal flora in piglets

    International Nuclear Information System (INIS)

    Namioka, Shigeo

    1980-01-01

    Utilization of non-protein nitrogen (NPN) by the flora in piglet colon was studied by administration of 15 N-urea and 15 N-ammonium salt to aseptic piglets and to SPF piglets which had been acclimatized to a clean environment after settling of intestinal flora. Administration of 15 N-urea did not result in 15 N uptake by any tissue-constituting protein at any site of the aseptic piglets, almost all 15 N being excreted into the urine. In contrast, the tissue and skeletal muscle of the SPF piglets showed incorporated 15 N from urea. Urea was converted, by urease of the intestinal flora, into NH 3 , which was absorbed from the mucosa of the intestinal tract to reach the liver where it was synthesized into glutamic acid, followed by conversion into various amino acids. 15 N-ammonium administration produced a significant amount of 15 N even in the tissue protein of the aseptic piglets. After NPN administration, the liver protein-constituting amino acid fraction showed 15 N-labeling of almost all essential, as well as non-essential amino acids. Culture of colonic flora with 15 N-urea revealed 15 N-labeling of all amino acids that constituted bacterial cells, indicating the presence of urea recycling mediated by bacterial urease in single rumen animals.(Chiba, N.)

  13. Hemolytic porcine intestinal Escherichia coli without virulence-associated genes typical of intestinal pathogenic E. coli.

    Science.gov (United States)

    Schierack, Peter; Weinreich, Joerg; Ewers, Christa; Tachu, Babila; Nicholson, Bryon; Barth, Stefanie

    2011-12-01

    Testing 1,666 fecal or intestinal samples from healthy and diarrheic pigs, we obtained hemolytic Escherichia coli isolates from 593 samples. Focusing on hemolytic E. coli isolates without virulence-associated genes (VAGs) typical for enteropathogens, we found that such isolates carried a broad variety of VAGs typical for extraintestinal pathogenic E. coli.

  14. characterization of intestinal microbiota in celiac children

    African Journals Online (AJOL)

    F. Lahcene1*, A. Tir Touil Meddah1, K. Bouziane-Nedjadi2, B. Meddah1, A. Leke3

    2016-09-01

    Sep 1, 2016 ... In this study, 13 Samples of intestinal biopsy, ... Research Article. Journal of Fundamental and Applied Sciences is licensed under a Creative Commons Attribution-NonCommercial 4.0. International License. ... disease are caused by several factors among, the ingestion of gluten, and it has been suggested.

  15. Schistosomiasis and intestinal helminth infections in Sengerema

    African Journals Online (AJOL)

    Abstract: Schistosomiasis and intestinal helminth infections are common health problems among the people of Lake. Victoria basin particularly in Mwanza ... these infections is largely lacking and that biomedical studies give only a partial ... sampling procedure was such that in each sub-village. ' Correspondence: Joseph R.

  16. intestinal helminthiasis among malnourished school age children

    African Journals Online (AJOL)

    Government Area to determine the prevalence rate of intestinal hjelminth infection among malnourished school children. Stool samples and finger prick blood .... in both male and female school children compared with a marked depreciation clue to the .... -rnalnutrition among India children. He submitted that control of such ...

  17. Oral cholera vaccination promotes homing of IgA+ memory B cells to the large intestine and the respiratory tract.

    Science.gov (United States)

    van Splunter, M; van Hoffen, E; Floris-Vollenbroek, E G; Timmerman, H; de Bos, E Lucas-van; Meijer, B; Ulfman, L H; Witteman, B; Wells, J M; Brugman, S; Savelkoul, H F J; van Neerven, R J J

    2018-02-21

    Oral cholera vaccination is used to induce immune responses in the intestines to protect against cholera infection. However, oral vaccination may also affect immune responses in other mucosal tissues. To study this, tissue-specific homing potential and kinetics of B-cell responses were characterized after oral cholera vaccination. Healthy adult volunteers received two doses of Dukoral® and blood, saliva, nasal wash, and fecal samples were collected over time to detect vaccine-specific antibodies. Additionally, homing potential of lymphocytes to small intestine, colon, airways, skin, and periphery was measured by expression of Integrin β1 and β7, CCR9, CCR10, CCR7, and CLA. After vaccination, antibody responses to cholera toxin B (CTB) and Dukoral® were detected in serum and nasal wash. CTB-specific memory B cells in peripheral blood and tissue homing profiles of memory B cells peaked at day 18. IgA + memory B cells expressed markers that enable homing to the airways and colon, while IgA - memory B cells primarily expressed small-intestine-homing markers. These data show that oral cholera vaccination has a differential effect on immune responses in various mucosal sites, including the respiratory tract.

  18. Repairing organs: lessons from intestine and liver.

    Science.gov (United States)

    Gehart, Helmuth; Clevers, Hans

    2015-06-01

    The concept of organ regeneration has fascinated humanity from ancient mythology to modern science fiction. Recent advances offer the potential to soon bring such technology within the grasp of clinical medicine. Rapidly expanding insights into the intrinsic repair processes of the intestine and liver have uncovered significant plasticity in epithelial tissues. Harnessing this knowledge, researchers have recently created culture systems that enable the expansion of stem cells into transplantable tissue in vitro. Here we discuss how the growing tool set of stem cell biology can bring organ repair from fictitious narrative to medical practice. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Coagulation of sheep intestinal and prefemoral lymph.

    Science.gov (United States)

    Hanley, C A; Johnston, M G; Nelson, W

    1988-06-01

    We have determined the most suitable method for the automated analysis of the clotting parameters in sheep intestinal and prefemoral lymph as defined by the Activated Partial Thromboplastin Times (APTT; measure of intrinsic coagulation pathway) and the Prothrombin Times (PT; measure of extrinsic coagulation pathway). As opposed to optical density systems, the use of a Fibro-System Fibrometer was found to provide the most consistent assessment of coagulation with the endpoint being the time to fibrin strand formation. We measured APTT in sheep intestinal and prefemoral lymph of 59.78 +/- 7.69 seconds and 51.03 +/- 10.49 seconds respectively. These values were more prolonged than those obtained from sheep blood plasma but only in the case of intestinal lymph were the differences significant (p less than 0.025). Human blood APTT values were significantly less than both sheep blood (p less than 0.05) and sheep intestinal (p less than 0.001) and prefemoral lymph (p less than 0.01). PT values were found to be 21.56 +/- 1.14 seconds in intestinal and 22.00 +/- 1.88 seconds in prefemoral lymph. These values were also significantly greater than those obtained from sheep blood (both p less than 0.001). Human blood PTs were significantly less than both sheep blood (p less than 0.001) and intestinal and prefemoral lymph (both p less than 0.001). Measurement of APTT and PT values in intestinal lymph and PT determinations in prefemoral lymph were not affected by storage in the refrigerator or freezer. There was some indication that APTT values in prefemoral samples were susceptible to storage artifacts; however, the differences in coagulation times were not significant.

  20. Relationships between intestinal parasitosis and handedness.

    Science.gov (United States)

    Uslu, Hakan; Dane, Senol; Uyanik, M Hamidullah; Ayyildiz, Ahmet

    2010-07-01

    The aim of the study was to investigate if there is a possible relation between intestinal parasitosis and handedness in patients with suspected intestinal parasitosis. Hand preference was assessed on the Edinburgh Handedness Inventory. Stool samples were examined microscopically for the presence of parasite. In the present study right-handers had many more helminth infections and left-handers had many more protozoon infections. Lower rate of helminth infections in the present study, and higher asthma incidences in the left-handed population in literature, may be associated with different immune machinery in left-handed people than in right-handed ones.

  1. Intestinal tuberculosis sometimes mimics Crohn's disease.

    Directory of Open Access Journals (Sweden)

    Esfandiar Shojaei

    2013-11-01

    Full Text Available Intestinal tuberculosis is an uncommon presentation of tuberculosis (TB and has clinicopathological similarities with Crohn's disease. In regions where TB is endemic clinicians must aware of this condition and fully evaluate their patients when Crohn's disease is diagnosed. We recommend all pathologic specimens be evaluate effectively for TB.Smear,culture and PCR for Mycobacterium.tuberculosis from samples aside the pathological reviews help for better diagnosis. Here we present a case of intestinal tuberculosis which initially diagnosed as Crohn's disease but after starting immunosuppressive agents he presented with disseminated tuberculosis.

  2. Effects of post-conditioning with sevoflurane on the expressions of intestinal AQP8 and I-FABP in pigs with hemorrhagic shock

    Directory of Open Access Journals (Sweden)

    Yan-hong CHEN

    2015-11-01

    Full Text Available Objective To observe the effects of sevoflurane post-conditioning on the expression of Aquaporin 8 (AQP8 and intestinal fatty acid binding protein (I-FABP, in order to investigate the protective role of sevoflurane post-conditioning on intestinal injury and its underlying mechanism. Methods Eighteen bama miniature pigs were randomly divided into three groups (6 each using a random number table: control group (S group, hemorrhagic shock group (HS group, and sevoflurane post-coditioning group (Post/ Sev group. Experimental animals were fasted for 8 hours before surgery, and propofol 3mg/kg was given viathe ear vein. Endotracheal intubation was done when the animal fell asleep. Bloodletting from the femoral artery after anesthesia was done to reproduce hemorrhagic shock. In Post/Sev group, 2% sevoflurane was given by inhalation for 30min (post-conditioning after successful reproduction of the model. Blood samples were collected prior to anesthesia (T0 and 30min (T1, 1h (T2, 1.5h (T3, 2h (T4, 3h (T5, 4h (T6 after hemorrhagic shock. The quantity of blood I-FABP and intestinal AQP8 levels were determined with ELISA. Water content in the intestinal tissue was determined by wet and dry weight method. Histopathological changes in the intestinal tissue were observed with HE staining. Results Compared with the control group, the serum I-FABP content, the expressions of intestinal AQP8, and water content in the intestinal tissue were significantly increased in HS group and Post/Sev (P<0.05 group. Compared with HS group, the above indices in Post/Sev group were significantly lower (P<0.05. These results were confirmed by pathological examination. Conclusion Postconditioning with sevoflurane could improve, to some extent, pig's intestinal barrier function in hemorrhagic shock, and this effect is likely related with lowering of intestinal AQP8 and I-FABP expression and mucosal edema. DOI: 10.11855/j.issn.0577-7402.2015.11.11

  3. Partial Enteral Nutrition Preserves Elements of Gut Barrier Function, Including Innate Immunity, Intestinal Alkaline Phosphatase (IAP) Level, and Intestinal Microbiota in Mice.

    Science.gov (United States)

    Wan, Xiao; Bi, Jingcheng; Gao, Xuejin; Tian, Feng; Wang, Xinying; Li, Ning; Li, Jieshou

    2015-08-03

    Lack of enteral nutrition (EN) during parenteral nutrition (PN) leads to higher incidence of infection because of gut barrier dysfunction. However, the effects of partial EN on intestina linnate immunity, intestinal alkaline phosphatase (IAP) and microbiota remain unclear. The mice were randomized into six groups to receive either standard chow or isocaloric and isonitrogenous nutritional support with variable partial EN to PN ratios. Five days later, the mice were sacrificed and tissue samples were collected. Bacterial translocation, the levels of lysozyme, mucin 2 (MUC2), and IAP were analyzed. The composition of intestinal microbiota was analyzed by 16S rRNA pyrosequencing. Compared with chow, total parenteral nutrition (TPN) resulted in a dysfunctional mucosal barrier, as evidenced by increased bacterial translocation (p < 0.05), loss of lysozyme, MUC2, and IAP, and changes in the gut microbiota (p < 0.001). Administration of 20% EN supplemented with PN significantly increased the concentrations of lysozyme, MUC2, IAP, and the mRNA levels of lysozyme and MUC2 (p < 0.001). The percentages of Bacteroidetes and Tenericutes were significantly lower in the 20% EN group than in the TPN group (p < 0.001). These changes were accompanied by maintained barrier function in bacterial culture (p < 0.05). Supplementation of PN with 20% EN preserves gut barrier function, by way of maintaining innate immunity, IAP and intestinal microbiota.

  4. Diagnosis of intestinal and extra intestinal amoebiasis

    International Nuclear Information System (INIS)

    Lopez, Myriam Consuelo; Quiroz, Damian Arnoldo; Pinilla, Analida Elizabeth

    2007-01-01

    The objective is to carry out a review of the national and international literature as of the XXth century in order to update the advances for the diagnosis of complex odd Entamoeba histolytic / Entamoeba dispar and that of intestinal and extra intestinal amoebiasis that may be of use to the scientific community. As well as to unify the diagnostic criteria of this parasitosis known as a public health problem, and as a consequence of that, optimize the quality of population care. Data source: there was a systematic search for the scientific literature Publisher in Spanish and English since 1960 until today, this selection started on the first semester of 2006 until 2007, in the development of the line on intestinal and extra-intestinal amoebiasis of the Medical School of the National University of Colombia. A retrospective search process was carried out, systematically reviewing the most relevant articles as well as the products of this research line. In deciding how to make this article, there was a continuous search in different data bases such as Medline, SciELO and other bases in the library of the National University of Colombia, as well as other classical books related to the subject. For that purpose the terms amoebiasis, odd Entamoeba histolytic, Entamoeba, diagnosis, epidemiology, dysentery, amoebic liver abscess, were used. Studies selection: titles and abstracts were reviewed to select the original publications and the most representative ones related to this article's subject. Data extraction: the articles were classified according to the subject, the chronology and the authors according to the scientific contribution to solve the problem. Synthesis of the data: in the fi rst instance, a chronological critical analysis was carried out to order and synthesize the progress made in the diagnosis until confirmation of the experts' agreements in the field of amoebiasis was obtained throughout the world. Conclusion: this article summarizes what has taken place

  5. Intestinal parasitiasis: Positive cases and low haematocrit among ...

    African Journals Online (AJOL)

    %) of stool samples examined. No characteristic symptoms of intestinal parasitic disease were observed in any of the infected women. However haematocrit results show low PCV (20-29%) in more of the infected than the uninfected women.

  6. Molecular detection of intestinal parasites for clinical diagnosis and epidemiology

    NARCIS (Netherlands)

    Hove, Robert Jan ten

    2009-01-01

    The detection of intestinal parasitic infections for routine diagnosis and for epidemiological research still depends mainly on microscopical examination of stool samples for the identification of helminth eggs and protozoan trophozoites and cysts. Because microscopy has several limitations,

  7. Intestinal anisakidosis (anisakiosis).

    Science.gov (United States)

    Takei, Hidehiro; Powell, Suzanne Z

    2007-10-01

    A case of intestinal anisakidosis in a 42-year-old man in Japan is presented. His chief complaint was an acute onset of severe abdominal pain. Approximately 12 hours before the onset of this symptom, he had eaten sliced raw mackerel ("sashimi"). Upper endoscopy was unremarkable. At exploratory laparotomy, an edematous, diffusely thickened segment of jejunum was observed, which was resected. The postoperative course was uneventful. The segment of small intestine showed a granular indurated area on the mucosal surface, and microscopically, a helminthic larva penetrating the intestinal wall, which was surrounded by a cuff of numerous neutrophils and eosinophils, as well as diffuse acute serositis. A cross section of the larva revealed the internal structures, pathognomonic of Anisakis simplex. Although anisakidosis is rare in the United States, with the increasing popularity of Japanese cuisine, the incidence is expected to increase, and pathologists should be familiar with this disease.

  8. Intestinal failure: a review

    Science.gov (United States)

    Allan, Philip; Lal, Simon

    2018-01-01

    Intestinal failure (IF) is the inability of the gut to absorb necessary water, macronutrients (carbohydrate, protein, and fat), micronutrients, and electrolytes sufficient to sustain life and requiring intravenous supplementation or replacement. Acute IF (types 1 and 2) is the initial phase of the illness and may last for weeks to a few months, and chronic IF (type 3) from months to years. The challenge of caring for patients with IF is not merely the management of the underlying condition leading to IF or the correct provision of appropriate nutrition or both but also the prevention of complications, whether thromboembolic phenomenon (for example, venous occlusion), central venous catheter-related bloodstream infection, IF-associated liver disease, or metabolic bone disease. This review looks at recent questions regarding chronic IF (type 3), its diagnosis and management, the role of the multidisciplinary team, and novel therapies, including hormonal treatment for short bowel syndrome but also surgical options for intestinal lengthening and intestinal transplant. PMID:29399329

  9. Small intestine diverticuli

    International Nuclear Information System (INIS)

    Pomakov, P.; Risov, A.

    1991-01-01

    The routine method of contrast matter passage applied to 850 patients with different gastrointestinal diseases proved inefficient to detect any small-intestinal diverticuli. The following modiffications of the method have been tested in order to improve the diagnostic possibilities of the X-ray: study at short intervals, assisted passage, enteroclysm, pharmacodynamic impact, retrograde filling of the ileum by irrigoscopy. Twelve diverticuli of the small-intestinal loops were identified: 5 Meckel's diverticuli, 2 solitary of which one of the therminal ileum, 2 double diverticuli and 1 multiple diverticulosis of the jejunum. The results show that the short interval X-ray examination of the small intestines is the method of choice for identifying local changes in them. The solitary diverticuli are not casuistic scarcity, its occurrence is about 0.5% at purposeful X-ray investigation. The assisted passage method is proposed as a method of choice for detection of the Meckel's diverticulum. 5 figs., 3 tabs. 18 refs

  10. Chronic intestinal pseudoobstruction syndrome

    International Nuclear Information System (INIS)

    Yeon, Kyung Mo; Seo, Jeong Kee; Lee, Yong Seok

    1992-01-01

    Chronic intestinal pseudoobstruction syndrome is a rare clinical condition in which impaired intestinal peristalsis causes recurrent symptoms of bowel obstruction in the absence of a mechanical occlusion. This syndrome may involve variable segments of small or large bowel, and may be associated with urinary bladder retention. This study included 6 children(3 boys and 3 girls) of chronic intestinal obstruction. Four were symptomatic at birth and two were of the ages of one month and one year. All had abdominal distension and deflection difficulty. Five had urinary bladder distension. Despite parenteral nutrition and surgical intervention(ileostomy or colostomy), bowel obstruction persisted and four patients expired from sepses within one year. All had gaseous distension of small and large bowel on abdominal films. In small bowel series, consistent findings were variable degree of dilatation, decreased peristalsis(prolonged transit time) and microcolon or microrectum. This disease entity must be differentiated from congenital megacolon, ileal atresia and megacystis syndrome

  11. Small Intestinal Infections.

    Science.gov (United States)

    Munot, Khushboo; Kotler, Donald P

    2016-06-01

    Small intestinal infections are extremely common worldwide. They may be bacterial, viral, or parasitic in etiology. Most are foodborne or waterborne, with specific etiologies differing by region and with diverse pathophysiologies. Very young, very old, and immune-deficient individuals are the most vulnerable to morbidity or mortality from small intestinal infections. There have been significant advances in diagnostic sophistication with the development and early application of molecular diagnostic assays, though these tests have not become mainstream. The lack of rapid diagnoses combined with the self-limited nature of small intestinal infections has hampered the development of specific and effective treatments other than oral rehydration. Antibiotics are not indicated in the absence of an etiologic diagnosis, and not at all in the case of some infections.

  12. Intestinal Stem Cell Markers in the Intestinal Metaplasia of Stomach and Barrett's Esophagus.

    Directory of Open Access Journals (Sweden)

    Bo Gun Jang

    Full Text Available Gastric intestinal metaplasia (IM is a highly prevalent preneoplastic lesion; however, the molecular mechanisms regulating its development remain unclear. We have previously shown that a population of cells expressing the intestinal stem cell (ISC marker LGR5 increases remarkably in IM. In this study, we further investigated the molecular characteristics of these LGR5+ cells in IM by examining the expression profile of several ISC markers. Notably, we found that ISC markers-including OLFM4 and EPHB2-are positively associated with the CDX2 expression in non-tumorous gastric tissues. This finding was confirmed in stomach lesions with or without metaplasia, which demonstrated that OLFM4 and EPHB2 expression gradually increased with metaplastic progression. Moreover, RNA in situ hybridization revealed that LGR5+ cells coexpress several ISC markers and remained confined to the base of metaplastic glands, reminiscent to that of normal intestinal crypts, whereas those in normal antral glands expressed none of these markers. Furthermore, a large number of ISC marker-expressing cells were diffusely distributed in gastric adenomas, suggesting that these markers may facilitate gastric tumorigenesis. In addition, Barrett's esophagus (BE-which is histologically similar to intestinal metaplasia-exhibited a similar distribution of ISC markers, indicating the presence of a stem cell population with intestinal differentiation potential. In conclusion, we identified that LGR5+ cells in gastric IM and BE coexpress ISC markers, and exhibit the same expression profile as those found in normal intestinal crypts. Taken together, these results implicate an intestinal-like stem cell population in the pathogenesis of IM, and provide an important basis for understanding the development and maintenance of this disease.

  13. Effects of probiotic on the intestinal morphology with special reference to the growth of broilers

    International Nuclear Information System (INIS)

    Lutfullah, G.; Ahmad, I.

    2011-01-01

    The probiotic (Protexin) increases the growth rate in broilers. It must interfere with the intestinal cell morphology and absorption. The intestinal epithelium is one of the most rapidly renewed tissues in the body and is renewed by a process of continuous cell division. This study was carried out with an aim to establish a link between the use of probiotic doses, growth rate, and intestinal cell proliferation by measuring the length and weight of the intestine and intestinal crypt cell proliferation (CCP) of broiler chicks. The results revealed significant increase in intestinal CCP but no effect was observed on the intestinal weight and length. The increase in CCP has also no significant influence towards growth factor. The increased weight gain in this study is associated with more feed consumption which is observed with Protexin dose 1.0 g / 10 kg of feed. Furthermore, feed consumption reduced beyond this dose may lead to reduced weight gain. (author)

  14. The intestinal calcistat

    Directory of Open Access Journals (Sweden)

    M K Garg

    2013-01-01

    Full Text Available The main physiological function of vitamin D is maintenance of calcium homeostasis by its effect on calcium absorption, and bone health in association with parathyroid gland. Vitamin D deficiency (VDD is defined as serum 25-hydroxy vitamin D (25OHD levels <20 ng/ml. Do all subjects with VDD have clinical disease according to this definition? We hypothesize that there exist an intestinal calcistat, which controls the calcium absorption independent of PTH levels. It consists of calcium sensing receptor (CaSR on intestinal brush border, which senses calcium in intestinal cells and vitamin D system in intestinal cells. CaSR dampens the generation of active vitamin D metabolite in intestinal cells and decrease active transcellular calcium transport. It also facilitates passive paracellular diffusion of calcium in intestine. This local adaptation adjusts the fractional calcium absorption according the body requirement. Failure of local adaptation due to decreased calcium intake, decreased supply of 25OHD, mutation in CaSR or vitamin D system decreases systemic calcium levels and systemic adaptations comes into the play. Systemic adaptations consist of rise in PTH and increase in active vitamin D metabolites. These adaptations lead to bone resorption and maintenance of calcium homeostasis. Not all subjects with varying levels of VDD manifest with secondary hyperparathyroidism and decreased in bone mineral density. We suggest that rise in PTH is first indicator of VDD along with decrease in BMD depending on duration of VDD. Hence, subjects with any degree of VDD with normal PTH and BMD should not be labeled as vitamin D deficient. These subjects can be called subclinical VDD, and further studies are required to assess beneficial effect of vitamin D supplementation in this subset of population.

  15. Human intestinal mucus proteins isolated by transanal irrigation and proctosigmoidoscopy

    OpenAIRE

    Paola Andrea Gómez Buitrago; Carlos Augusto González Correa; Mario Santacoloma Osorio; Gonzalo Taborda Ocampo; Marco Aurelio Zezzi Arruda

    2014-01-01

    Human intestinal mucus essentially consists of a network of Mucin2 glycoproteins embedded in many lower molecular weight proteins. This paper contributes to the proteomic study of human intestinal mucus by comparing two sample collection methods (transanal irrigation and brush cytology during proctosigmoidoscopy) and analysis techniques (electrophoresis and digestion in solution). The entire sample collection and treatment process is explained, including protein extraction, digestion and desa...

  16. Foreign body-associated intestinal pyogranuloma resulting in intestinal obstruction in four dogs.

    Science.gov (United States)

    Papazoglou, L G; Tontis, D; Loukopoulos, P; Patsikas, M N; Hermanns, W; Kouti, V; Timotheou, T; Liapis, I; Tziris, N; Rallis, T S

    2010-04-17

    Intestinal obstruction resulting from an intramural foreign body-associated pyogranuloma was diagnosed in four dogs. Vomiting and weight loss were the main clinical signs. On physical examination, a mass in the abdomen was detected in three dogs. Abdominal radiography demonstrated the presence of soft tissue opacity in three of the dogs and gas-filled dilated intestinal loops in all four dogs. Abdominal ultrasonography showed hyperkinetic fluid-filled dilated intestinal loops and a hypoechoic small intestinal mass in all the dogs. Exploratory coeliotomy confirmed the presence of a jejunal mass, which was removed by resection and anastomosis in all the dogs. In one of the dogs a linear foreign body was also found cranial to the mass and was removed through a separate enterotomy incision. The lesions were diagnosed as foreign body-associated intestinal pyogranulomas on histological examination. Three dogs recovered without complications, but the fourth showed signs of septic peritonitis four days after surgery and was euthanased at the owner's request. The other three dogs remained disease-free 12 to 42 months after surgery.

  17. Detection of free and covalently bound microcystins in different tissues (liver, intestines, gills, and muscles) of rainbow trout (Oncorhynchus mykiss) by liquid chromatography–tandem mass spectrometry: Method characterization

    International Nuclear Information System (INIS)

    Cadel-Six, Sabrina; Moyenga, David; Magny, Stéphanie; Trotereau, Sophie; Edery, Marc; Krys, Sophie

    2014-01-01

    So far only a few publications have explored the development of extraction methods of cyanotoxin extracted from complex matrices. With regard to cyanobacterial microcystins (MCs), the data on the contamination of the flesh of aquatic organisms is hard to compare and very limited due to the lack of validated methods. In recent years, evidence that both free and bound fractions of toxin are found in these tissues has highlighted the need to develop effective methods of quantification. Several techniques do exist, but only the Lemieux oxidation has so far been used to investigate complex tissue matrices. In this study, protocols based on the Lemieux approach were adapted for the quantitative chemical analysis of free MC-LR and MMPB derived from bound toxin in the tissues of juvenile trout gavaged with MC-LR. Afterwards, the NF V03 110 guideline was used to characterize the protocols elaborated and evaluate their effectiveness. -- Highlights: • We adapted the quantitative chemical analysis of free and total MC for the tissues of trout. • We characterize with the NF V03 110 guideline the protocols for free MC-LR and MMPB. • We quantify the free MC-LR and MMPB in the tissues of trout gavaged with MC-LR. -- We develop and characterize with the guideline NF V03 110 the protocols for the extraction and quantification of the free and total MC for different matrices

  18. Culture of human intestinal epithelial cell using the dissociating enzyme thermolysin and endothelin-3

    OpenAIRE

    Liu, Z.; Zhang, P.; Zhou, Y.; Qin, H.; Shen, T.

    2010-01-01

    Epithelium, a highly dynamic system, plays a key role in the homeostasis of the intestine. However, thus far a human intestinal epithelial cell line has not been established in many countries. Fetal tissue was selected to generate viable cell cultures for its sterile condition, effective generation, and differentiated character. The purpose of the present study was to culture human intestinal epithelial cells by a relatively simple method. Thermolysin was added to improve the yield of epithel...

  19. Presence of intestinal<