Sections Of Mucosa Research Articles

Effects of carprofen on the integrity and barrier function of canine colonic mucosa

To measure effects of carprofen on conductance and permeability to mannitol and histologic appearance in canine colonic mucosa. Colonic mucosa from 13 mature mixed-breed dogs. Procedures-Sections of mucosa from the transverse colon and proximal and distal portions of the descending colon were obtained immediately after dogs were euthanized. Sections were mounted in Ussing chambers. Carprofen (400 microg/mL) was added to the bathing solution for treated sections. Conductance was calculated at 15-minute intervals for 240 minutes. Flux of mannitol was calculated for three 1-hour periods. Histologic examination of sections was performed after experiments concluded. Conductance was graphed against time for each chamber, and area under each curve was calculated. Conductance X time, flux of mannitol, and frequency distribution of histologic findings were analyzed for an effect of region and carprofen. Carprofen significantly increased mean conductance X time, compared with values for control (untreated) sections for all regions of colon. Carprofen significantly increased mean flux of mannitol from period 1 to period 2 and from period 2 to period 3 for all regions of colon. Carprofen caused a significant proportion of sections to have severe sloughing of cells and erosions involving >or= 10% of the epithelium, compared with control sections. Carprofen increased in vitro conductance and permeability to mannitol in canine colonic mucosa. Carprofen resulted in sloughing of cells and erosion of the colonic mucosa. These findings suggested that carprofen can compromise the integrity and barrier function of the colonic mucosa of dogs.

Duodenum mucosa denudation after gamma rays irradiation of rat small intestine.

In order to understand the mechanism of denudation of intestine mucosa after irradiation stress with doses used in cancer radiotherapy, rats were whole body irradiated using 60-Co-gamma rays at three doses and two dose rates. Fragments of duodenum mucosa were take off 24 or 48 h after irradiation, and processed for microscopy observations. For each dose and dose rate, one thousand 2 micrometer-thick serial epon sections were pictured and digital images were imported into softwares, ImageJ and Reconstruct for 3d reconstruction. Morphological changes were observed within irradiated mucosa when compared to control that worst with dose and dose rate. Numerous lesions were noted as villous atrophy, crypts and villi branched, lamina propria vacuoles, large and numerous irregular villus pleats. Lost of large fragments of epithelial layer were observed. Ultrathin sections of irradiated mucosa revealed typical apoptotic lesions within columnar cells as chromatin condensation of irregular shape nuclei, nuclear fragmentation and swollen mitochondria. Apoptotic cells were detected at crypt bottom but not at villi tip as are usually reported. Apoptotic index increased with dose and post irradiation time. Detachment of two neighbour epithelial layers was detected at crypt stem cells site. Animation of mucosa after 3dr has show entire villi detachment from crypts that propagate longitudinally through mucosa. Apoptotic process induced by radiation and epithelial layer detachment occurred at same site (between villi bottom and crypt corresponding to stem cells position). Thus we established that mucosa denudation occur by entire villi detachment via lost of intercellular contacts of apoptotic stem cells. This study elucidates the enigma of the fast denudation of intestine mucosa following irradiation.

This Month in Gastroenterology

This Month in Gastroenterology

Characterization of immunoisolated human gastric parietal cells tubulovesicles: identification of regulators of apical recycling

Gastric parietal cells possess an amplified apical membrane recycling system dedicated to regulated apical recycling of H-K-ATPase. While amplified in parietal cells, apical recycling is critical to polarized secretory processes in most epithelial cells. To clarify putative regulators of apical recycling, we prepared immunoisolated parietal cell H-K-ATPase-containing recycling membranes from human stomachs and analyzed protein contents by tryptic digestion and mass spectrometry. We identified and validated by Western blots many of the proteins previously identified on immunoisolated rabbit tubulovesicles, including Rab11, Rab25, syntaxin 3, secretory carrier membrane proteins (SCAMPs), and vesicle-associated membrane protein (VAMP)2. In addition, we detected several previously unrecognized proteins, including Rab10, VAMP8, syntaxin 7, and syntaxin 12/13. We also identified the K(+) channel component KCNQ1. Immunostaining of human gastric mucosal sections confirmed the presence of each of these proteins in parietal cells and their colocalization with H-K-ATPase on tubulovesicles. To investigate the role of the identified soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins in apical recycling, we transfected them as DsRed2 fusions into an enhanced green fluorescent protein (EGFP)-Rab11a-expressing Madin-Darby canine kidney (MDCK) cell line. Syntaxin 12/13 and VAMP8 caused a collapse of the EGFP-Rab11a compartment, whereas a less dramatic effect was observed in cells transfected with syntaxin 3, syntaxin 7, or VAMP2. The five DsRed2-SNARE chimeras were also transfected into a MDCK cell line overexpressing Rab11-FIP2(129-512). All five of the chimeras were drawn into the collapsed apical recycling system. This study, which represents the first proteomic analysis of an immunoisolated vesicle population from native human tissue, demonstrates the diversity of putative regulators of the apical recycling system.

Intestinal Tight Junction in Allograft After Small Bowel Transplantation

Introduction In the intestinal physical barrier, tight junctions between intestinal epithelial cells play a central role. There is increasing evidence that rejection after small bowel transplantation promotes intestinal barrier injury to allografts. Our aim was to study the morphological changes of tight junctions in allografts during rejection. Methods Small bowel transplantation was performed using the F344 to Lewis rat model. Animals were divided into three groups: isogeneic controls, acute rejection group, and chronic rejection group. Allograft rejection was characterized by hemotoxylin and eosin staining of mucosal tissue sections. Tight junctions in grafts were investigated by transmission electron microscopy. Results Acutely rejected allografts showed severe mucosal injury and completely loosened tight junctions, while chronically rejected allografts revealed less mucosal injury and remained with partial integrity of their tight junctions. Conclusion Our study demonstrated for the first time the morphological alterations of tight junctions in allograft mucosa during acute and chronic rejections, suggesting disruption of tight junctions was relative to the intestinal inflammatory processes.

Detection of PrPCWD in Postmortem Rectal Lymphoid Tissues in Rocky Mountain Elk (Cervus Elaphus Nelsoni) Infected with Chronic Wasting Disease

Preclinical diagnostic tests for transmissible spongiform encephalopathies have been described for mule deer (Odocoileus hemionus), using biopsy tissues of palatine tonsil, and for sheep, using lymphoid tissues from palatine tonsil, third eyelid, and rectal mucosa. The utility of examining the rectal mucosal lymphoid tissues to detect chronic wasting disease (CWD) was investigated in Rocky Mountain elk (Cervus elaphus nelsoni), a species for which there is not a live-animal diagnostic test. Postmortem rectal mucosal sections were examined from 308 elk from two privately owned herds that were depopulated. The results of the postmortem rectal mucosal sections were compared to immunohistochemical staining of the brainstem, retropharyngeal lymph nodes, and palatine tonsil. Seven elk were found positive using the brainstem (dorsal motor nucleus of the vagus nerve), retropharyngeal lymph nodes, and palatine tonsil. Six of these elk were also found positive using postmortem rectal mucosal sections. The remaining 301 elk in which CWD-associated abnormal isoform of the prion protein (PrP(CWD)) was not detected in the brainstem and cranial lymphoid tissues were also found to be free of PrP(CWD) when postmortem rectal mucosal sections were examined. The use of rectal mucosal lymphoid tissues may be suitable for a live-animal diagnostic test as part of an integrated management strategy to limit CWD in elk.

Keratinocyte-Secreted Laminin 5 Can Function as a Transient Receptor for Human Papillomaviruses by Binding Virions and Transferring Them to Adjacent Cells

Human papillomaviruses (HPVs) replicate only in the terminally differentiating epithelium of the skin and mucosa. While infection of basal keratinocytes is considered a requirement for permissive infection, it remains unclear whether virions can specifically target basal cells for adsorption and uptake following epithelial wounding. We present evidence that HPV binds specifically to laminin 5 (LN5), a component of the extracellular matrix (ECM) secreted by migrating and basal keratinocytes. HPV type 11 capsids colocalized with LN5 in the ECM secreted by vaginal keratinocytes. Binding of both virions and virus-like particles to purified LN5 and to the LN5-rich ECM secreted by cultured keratinocytes was effectively blocked by pretreatment with anti-LN5 antibodies. HPV capsid binding to human cervical mucosa sections included the basement membrane which contains LN5. Cultured keratinocytes expressing alpha6 integrin, a transmembrane protein known to bind LN5, were readily infected by virions preadsorbed to LN5-containing substrates, whereas mutant keratinocytes lacking alpha6 integrin were relatively resistant to infection via this route. These findings suggest a model of natural HPV infection in which proliferating keratinocytes expressing alpha6 integrin at the site of epithelial wounding might be targeted by virions adsorbed transiently to LN5 secreted by migrating keratinocytes.

Diffusion of naltrexone across reconstituted human oral epithelium and histomorphological features

In transbuccal absorption a major limitation could be the low permeability of the mucosa which implies low drug bioavailability. The ability of naltrexone hydrochloride (NLX) to penetrate a resembling histologically human buccal mucosa was assessed and the occurrence of any histomorphological changes observed. We used reconstituted human oral (RHO) non-keratinised epithelium as mucosal section and a Transwell diffusion cells system as bicompartmental model. Buccal permeation was expressed in terms of drug flux ( J s) and permeability coefficients ( K p). Data were collected using both artificial and natural human saliva. The main finding was that RHO does not restrain NLX permeation. Drug transport across the epithelium was observed also in presence of various concentrations of penetration enhancers, without any significant differences. On the contrary, the flux throughout the mucosa was extensively affected by iontophoresis. Histologically, no sign of flogosis was observed in any specimen under experiment without iontophoresis, whereas cytoarchitectural changes, up to nuclear pycnosis or cellular swelling, were determined as a consequence of the application of electric fields.

Richard M. Peek, Jr, MD

Dr Peek received his BS degree cum laude from Davidson College in 1984. Later he attended the University of North Carolina at Chapel Hill where he received his MD degree in 1988. After Internal Medicine training and Chief Residency at the University of Alabama School of Medicine, he completed a fellowship in Gastroenterology at Vanderbilt University School of Medicine in 1995. In 1996, he became an Assistant Professor at Vanderbilt, where he is now the Mina Cobb Wallace Professor of Medicine and Cancer Biology and Director of the Division of Gastroenterology, Hepatology, and Nutrition. Dr. Peek is also co-Director of the Vanderbilt Digestive Diseases Research Center at Vanderbilt. Dr. Peek has received several awards and honors and is a member of numerous professional societies. He is the recipient of the Grant Liddle Scholarship for Excellence in Clinical Research (1994), the Glaxo Institute of Digestive Health Basic Research Award (1995), the Miles and Shirley Fiterman Foundation Basic Research Award (2001), the Young Investigator Award from the American Gastroenterological Association (2002), and in 2003 he was elected to the American Society for Clinical Investigation. A charter member of the Gastrointestinal Mucosal Pathobiology Study Section at NIDDK, Dr. Peek served as co-chair of the Biology Section for the National Cancer Institute-sponsored Stomach/Esophageal Cancers Progress Review Group in 2002, and was an invited participant in the NCI-sponsored Inflammation and Cancer Think Tank Meeting June 2–4, 2004, which was organized to identify research advances and opportunities in the study and clinical applications of inflammation on cancer initiation, progression, and metastasis. He also serves as a Section Editor for Gastroenterology and is on the Editorial board of the American Journal of Physiology: Gastrointestinal and Liver Physiology. Dr. Peek’s research efforts are grounded in understanding the molecular mechanisms through which H pylori enhances the risk for gastric cancer using both in vitro and in vivo models.

The immunohistology of CD40 in human oral epithelium in health and disease

CD40 has a role in the regulation of immune responses, cell proliferation and migration, and apoptosis. Little is known of its distribution in oral mucosal pathology. Oral keratinocyte lines were tested for CD40 protein by Western blotting. Immunohistochemistry was used to stain paraffin sections of oral mucosa in health and in inflammatory, reactive, dysplastic and malignant disease. Western blotting confirmed the presence of CD40 in oral keratinocytes. CD40 was generally expressed by keratinocytes in the basal layer, with variable parabasal expression. Langerhans cells also stained positively. Expression was lost in nine of 33 (27%) epithelial dysplasias, seven of which were severe. Eighty-one percent of well, 69% of moderately and 50% of poorly differentiated oral squamous cell carcinomas (OSCC) expressed CD40. Overall, 45 of 65 (69%) OSCC were positive. The pattern of expression was unrelated to tumour differentiation. CD40 expression by basal and parabasal oral keratinocytes is physiological. Expression is lost in approximately one-third of oral epithelial dysplasias and OSCC. The significance of such loss remains unknown, but may be related to immunological or other abnormalities of keratinocyte homeostasis.

Fascin is a potential biomarker for early-stage oesophageal squamous cell carcinoma

Background: Fascin, an actin-binding protein, is usually expressed at a low level in normal epithelium, but is markedly up regulated in several types of carcinomas. Reports on fascin expression in...

Investigation of Equine Influenza Cases Exhibiting Neurological Disease: Coincidence or Association?

Equine influenza is usually a transient and self-limiting disease. However, during an outbreak of equine influenza in the UK in 2003 there were reports of unusually severe clinical signs among unvaccinated animals. Two influenza-infected horses developed neurological signs, and one was subjected to euthanasia. Post-mortem examination of the brain revealed viral-type non-suppurative encephalitis, and influenza virus antigen was demonstrated by immunolabelling of sections of nasal mucosa. A syndrome known as influenza-associated encephalopathy has been described in man. Although not proved, the data suggest that similar disease mechanisms may operate in horses, and that equine influenza virus infection can result in encephalitis in the natural host, perhaps due to an aberrant host immune response.

Expression of retinoid receptors in multiple cell lineages in the gastric mucosae of mice and humans

In mice and humans, the gastric epithelial progenitors undergo proliferation and bipolar migration from the isthmus associated with their differentiation into mucus-, acid- and pepsinogen-secreting cell lineages. Little is known about factors that control the dynamics of these isthmal progenitor cells. Retinoids have long been known as chemopreventive agents against gastric mucosal damage and carcinogenesis. The aim of the present study was to examine the cellular localization of the various retinoid receptors proteins (RAR and RXR) in the gastric epithelium of mice and humans. Gastric antral biopsies of normal individuals and the oxyntic and antral regions of the mouse stomach were processed for immunohistochemistry using anti-RAR and anti-RXR antibodies. To label the progenitor cell zone, some sections were also probed with antibodies specific for proliferating cell nuclear antigen. The immunoprobed oxyntic mucosal sections of the mice showed that RXRbeta protein was present in the epithelial isthmal cells, neck cells, zymogenic cells and some pit and parietal cells. In addition, RARbeta was found in isthmal and neck cells, and RARgamma was mainly found in neck cells. In the mouse antrum, only RXRbeta was detected in the isthmal cells and their pit and gland cell descendents. In humans, immunoprobed antral sections showed that RARbeta, RARgamma, RXRalpha and RXRgamma proteins are expressed in the isthmal, pit and gland cells. Retinoid receptors are expressed in multiple cell lineages of the mouse and human gastric epithelium and may, therefore, account for the possible effects of retinoids on gastric epithelial cell proliferation and differentiation.

Correlation of spectral imaging and visual grading for the quantification of thymidylate synthase protein expression in rectal cancer

Quantification of protein expression in tissue sections stained by immunohistochemistry has traditionally involved visual grading techniques. However, if these results are to be used to predict tumor behavior and permit targeted therapy, there is a need for more accurate, objective, and reproducible methods. This study investigated the utility of spectral imaging as a method of quantifying thymidylate synthase protein expression in immunohistochemically stained sections of primary rectal cancer and normal rectal mucosa by comparing it with the current gold standard of manual visual grading. There was good correlation between estimates of thymidylate synthase stain intensity and area derived by spectral imaging and visual grading in both tumor and normal mucosal sections, suggesting that spectral imaging is a valid way of quantifying biologic sections stained by immunohistochemistry.

Polymorphism at the 3′-UTR of the thymidylate synthase gene: A potential predictor for outcomes in Caucasian patients with esophageal adenocarcinoma treated with preoperative chemoradiation

To test the hypothesis that TS3'UTR polymorphisms predict outcomes in 146 Caucasian patients with esophageal adenocarcinoma treated with preoperative 5-fluorouracil-based chemoradiation. DNA was extracted from hematoxylin-and-eosin stained histologic slides of normal esophageal or gastric mucosa sections from paraffin blocks of esophagectomy specimens. Genotypes of the TS3'UTR polymorphism were determined by polymerase chain reaction for a 6-bp insertion. The genotype groups (0bp/0bp, 6bp/0bp, and 6bp/6bp) were compared for clinical features and overall survival, recurrence-free-survival, locoregional control (LRC), and distant metastasis control. Multivariable Cox regression analyses were performed to find independent predictors for the stated outcomes. There was a trend of association between 6bp/6bp genotype and a decreased risk of local regional recurrence (hazards ratio = 0.211, 95% confidence interval = 0.041-1.095, p = 0.06) compared with other genotypes. There was a trend that patients with 6bp/6bp genotype had a higher 3-year probability of LRC compared with patients with the other two genotypes combined (p = 0.07); however, the difference was not statistically significant. The null hypotheses were not rejected in this study, probably owing to small sample size or the single gene examined. Prospective studies with adequate statistical power analyzing a family of genes involved in the 5-fluorouracil metabolism are needed to assess genetic determinants of treatment-related outcomes in esophageal adenocarcinoma.

662 POSTER Modification of the circadian rest/activity rhythm by 1st line oxaliplatin (I-OHP), 5-fluorouracil (5FU) and leucovorin (LV) in patients (pts) with metastatic colorectal cancer (MCC). an international study (EORTC 05963)

Purpose: To test the hypothesis that TS3′UTR polymorphisms predict outcomes in 146 Caucasian patients with esophageal adenocarcinoma treated with preoperative 5-fluorouracil-based chemoradiation.Methods and Materials: DNA was extracted from hematoxylin-and-eosin stained histologic slides of normal esophageal or gastric mucosa sections from paraffin blocks of esophagectomy specimens. Genotypes of the TS3′UTR polymorphism were determined by polymerase chain reaction for a 6-bp insertion. The genotype groups (0bp/0bp, 6bp/0bp, and 6bp/6bp) were compared for clinical features and overall survival, recurrence-free-survival, locoregional control (LRC), and distant metastasis control. Multivariable Cox regression analyses were performed to find independent predictors for the stated outcomes.Results: There was a trend of association between 6bp/6bp genotype and a decreased risk of local regional recurrence (hazards ratio = 0.211, 95% confidence interval = 0.041–1.095, p = 0.06) compared with other genotypes. There was a trend that patients with 6bp/6bp genotype had a higher 3-year probability of LRC compared with patients with the other two genotypes combined (p = 0.07); however, the difference was not statistically significant.Conclusions: The null hypotheses were not rejected in this study, probably owing to small sample size or the single gene examined. Prospective studies with adequate statistical power analyzing a family of genes involved in the 5-fluorouracil metabolism are needed to assess genetic determinants of treatment-related outcomes in esophageal adenocarcinoma.

Steroid receptor expression in vaginal epithelium of healthy fertile women and influences of hormonal contraceptive usage

Background The objective of this study was to evaluate whether long-term usage of hormonal contraceptives modifies the steroid receptor expression in the human vaginal epithelium of healthy young women. Methods In a cross-sectional study, three groups of hormonal contraceptive users [combined oral contraceptives (COCs), levonorgestrel implants (LNG) and depot medroxyprogesterone acetate injections (DMPAs)] were compared to controls. Fifteen subjects (20–34 years) were enrolled to each group. Vaginal biopsies were collected at two occasions from each subject, and serum concentrations of E 2 and progesterone were measured. Monoclonal antibodies directed against progesterone receptors (PRs) and estrogen receptors (ERs) were used in immunohistochemistry on formalin-fixed tissue sections of vaginal mucosa. A program for immunohistomorphometric quantification was devised to estimate frequency of epithelial steroid receptor-expressing cells. Results Progesterone receptor expression was markedly down-regulated and significantly reduced in DMPA users compared to controls, COC and LNG users. In DMPA users, the ER expression was significantly elevated in the first compared to the second sample, and significantly elevated compared to LNG users. Estradiol concentration in serum was significantly reduced in hormonal contraceptive users compared to controls. Conclusions Steroid receptor expression in human vaginal epithelium is altered by long-term use of DMPA compared to controls.

Diagnosis and therapy of ultrashort Hischsprung's disease: Meier-Ruge WA, Bruder E, Holschneider, et al. Eur J Pediatr Surg 2004 (December);14:392-397

Although ultrashort Hirschsprung's disease (UHD) was enzyme-histochemically characterised about 35 years ago, its existence is still often ignored. The aim of this study was to summarize diagnostic criteria, incidence, gender ratio, morphology and therapy over 15 years. Reliable diagnosis of suspected UHD requires contrast enema to exclude HD. In UHD, no reflux of contrast medium is observed during pressing or crying. Final proof of UHD is enzyme-histochemical biopsy examination of distal rectal mucosa. The biopsies must be taken from the dentate line and 1, 2, 4 and 6 cm above the dentate line and then subjected to an enzyme-histochemical acetylcholinesterase (AChE) reaction of native sections of rectal mucosa. UHD develops with first symptoms of therapy resistant chronic constipation during the second half of the first year of life. In contrast, constipation occurs in the first weeks of life or after weaning in HD. Also in contrast to HD, no nerve fibers with increased AChE activity are observed in the lamina propria mucosae in UHD. Nets of nerve fibers with increased AChE activity can be found only in the muscularis mucosae and the musculus corrugator cutis ani (MCCA). Therapy of choice has proven to be partial myectomy of the distal internal sphincter if dilatation of the internal sphincter is ineffective. UHD is either limited to the anal ring, or extends only 3-4 cm into the distal rectum. UHD had an incidence of 13.4% of all aganglionoses in the series presented. Gender ratio boys to girls was 2:1.

Effect of bladder ischaemia/reperfusion on superoxide dismutase activity and contraction

To correlate the effect of bilateral in-vivo bladder ischaemia/reperfusion on superoxide dismutase activity (SOD) and then to correlate this with contractile responses to various forms of stimulation. Twenty mature male New Zealand White rabbits were divided into five equal groups: group 1 (controls); group 2, 2 h of in-vivo bilateral bladder ischaemia; and groups 3-5, 2 h of in-vivo ischaemia followed by 1, 7 or 14 days of reperfusion (recovery). At the end of the treatment period, bladder strips were incubated and placed in isolated baths for contractile studies. The contractile responses to field stimulation, carbachol (10 micromol/L), ATP and KCl were determined. The balance of the bladder body was separated into muscle and mucosa sections and analysed for SOD activity. There were few effects on contraction either directly after ischaemia or after 1 day of reperfusion. However, all contractile responses were significantly reduced at 7 and 14 days after ischaemia. SOD activity of the detrusor muscle was reduced significantly immediately after ischaemia and at 7 and 14 days of reperfusion. SOD activity of the mucosa was significantly greater than that of the muscle, and was significantly reduced by both ischaemia and all times of reperfusion. These studies show clearly that both ischaemia and reperfusion result in significantly lower activity of SOD, and in contractile dysfunctions, and that reperfusion results in greater decreases in both SOD activity and contractile responses than ischaemia alone.

Assessment of c-Jun, c-Fos and cyclin D1 in premalignant and malignant oral lesions

Some oral cancers are known to develop from dysplastic oral epithelium. In the present study, the expression of c-Jun, c-Fos, and cyclin D1 proteins in oral epithelial lesions with different degrees of dysplasia, and in oral squamous cell carcinomas (OSCCs) was evaluated. Eighteen cases of mild dysplasia, 23 cases of moderate to severe dysplasia and 24 OSCCs were studied immunohistochemically. Additionally, 15 sections of oral mucosa without any evidence of dysplasia were included in the study. c-Jun expression increased according to the degree of oral dysplasia, with the greatest expression found in OSCC. c-Fos expression was intense in normal mucosa, reduced in mild dysplasia and high in moderate to severe dysplasia and in OSCCs. Cyclin D1 was expressed in only a few cases of moderate to severe dysplasia and in most of the OSCCs. Statistical analysis showed a correlation between the three proteins and the degree of epithelial alteration. The present results indicate a possible role of c-Jun and c-Fos in malignant transformation of oral mucosa.