Rat cellular retinol-binding protein II: use of a cloned cDNA to define its primary structure, tissue-specific expression, and developmental regulation.

The primary structure of rat cellular retinol-binding protein (CRBP) II has been determined from a cloned cDNA. Alignment of this 134-amino acid, 15,580-Da polypeptide with rat CRBP revealed that 75 of 133 comparable residues are identical. Both proteins contain four tryptophan residues, which occupy identical relative positions in the two primary structures, providing a structural explanation for their similar fluorescence spectra when complexed to retinol. Two of the three cysteines in each single-chain protein are comparably positioned. Both polypeptides contain reactive thiol groups, but the rate of disruption of CRBP II-retinol complexes by p-chloromercuribenzoate is greater than that of CRBP-retinol. The small intestine contains the highest concentrations of CRBP II mRNA in adult rats. CRBP II mRNA is first detectable in intestinal RNA during the 19th day of gestation, a time that corresponds to the appearance of an absorptive columnar epithelium. Unlike in intestine, a dramatic fall in liver CRBP II mRNA concentration occurs immediately after birth. The CRBP II gene remains quiescent in the liver during subsequent postnatal development. These data suggest that ligand-protein interactions may be somewhat different for the two rat CRBPs. They also support the concept that CRBP II plays a role in the intestinal absorption or esterification of retinol and suggest that changes in hepatic metabolism of vitamin A occur during development.     

Localization of cellular retinol-binding protein in several rat tissues.

The distribution of cellular retinol-binding protein (CRBP) in rat liver, ileum, and epididymis was examined by the peroxidase-antiperoxidase immunolocalization technique. Positive cytoplasmic staining was seen in the liver when antiserum prepared against purified CRBP was used but not when antiserum absorbed with purified CRBP was used. Ileal mucosa, a tissue that contains no detectable CRBP, showed no positive staining. The epididymis showed strong positive staining in the caput but not in the cauda. Staining was present in principal and basal cells but not in peritubular or interstitial cells. Radioimmunoassay revealed that the CRBP within the caput epididymidis was localized in the initial segment and proximal region, areas known to be involved in the synthesis and secretion of factors necessary for sperm maturation. The results demonstrate that the expression of CRBP may vary within the same cell type, as well as between different cell types within the same tissue.     

Cellular differentiation in the emerging fetal rat small intestinal epithelium: mosaic patterns of gene expression.

We have examined the pattern of differentiation of the small intestinal epithelium in fetal rats during the 17th through 21st days of gestation. Five genes expressed in late fetal, neonatal, and adult enterocytes were used as markers of differentiation. They encode three homologous small cytoplasmic hydrophobic ligand binding proteins--liver fatty acid binding protein (L-FABP), intestinal fatty acid binding protein (I-FABP), and cellular retinol binding protein II (CRBP II)--and two apolipoproteins--apoAI and apoAIV. RNA blot hybridization studies indicated that gradients in mRNA concentration from the proximal small intestine to colon appear coincident with the initiation of rapid epithelial cell proliferation and villus formation (days 17-19 of the 22-day gestation period). Immunocytochemical studies disclosed a remarkably heterogeneous pattern of cell-specific expression of the three hydrophobic ligand binding proteins that was not apparent with either apoAIV or apoAI. This "mosaic" staining pattern was observed in morphologically similar cells occupying identical topographic positions along nascent villi in 17- to 18-day fetuses. The onset and resolution of this mosaicism varies between I-FABP, L-FABP, and CRBP II in the proximal small bowel, although it completely resolves by the first postnatal day. The distal small intestine exhibits a developmental delay of 1-2 days in the appearance of this heterogeneous pattern of initial gene expression. Double-label immunofluorescent analyses using L-FABP and I-FABP antibodies indicated that on the 18th day of gestation the proximal small intestinal columnar epithelium contains several populations of enterocytes expressing neither, one, or both proteins. The potential significance of this mosaic pattern of intestinal epithelial differentiation is discussed in light of recent studies with transgenic and chimeric mice.     

Cellular and subcellular localization of the Nramp2 iron transporter in the intestinal brush border and regulation by dietary iron.

Genetic studies in animal models of microcytic anemia and biochemical studies of transport have implicated the Nramp2 gene in iron transport. Nramp2 generates two alternatively spliced mRNAs that differ at their 3' untranslated region by the presence or absence of an iron-response element (IRE) and that encode two proteins with distinct carboxy termini. Antisera raised against Nramp2 fusion proteins containing either the carboxy or amino termini of Nramp2 and that can help distinguish between the two Nramp2 protein isoforms (IRE: isoform I; non-IRE: isoform II) were generated. These antibodies were used to identify the cellular and subcellular localization of Nramp2 in normal tissues and to study possible regulation by dietary iron deprivation. Immunoblotting experiments with membrane fractions from intact organs show that Nramp2 is expressed at low levels throughout the small intestine and to a higher extent in kidney. Dietary iron starvation results in a dramatic upregulation of the Nramp2 isoform I in the proximal portion of the duodenum only, whereas expression in the rest of the small intestine and in kidney remains largely unchanged in response to the lack of dietary iron. In proximal duodenum, immunostaining studies of tissue sections show that Nramp2 protein expression is abundant under iron deplete condition and limited to the villi and is absent in the crypts. In the villi, staining is limited to the columnar absorptive epithelium of the mucosa (enterocytes), with no expression in mucus-secreting goblet cells or in the lamina propria. Nramp2 expression is strongest in the apical two thirds of the villi and is very intense at the brush border of the apical pole of the enterocytes, whereas the basolateral membrane of these cells is negative for Nramp2. These results strongly suggest that Nramp2 is indeed responsible for transferrin-independent iron uptake in the duodenum. These findings are discussed in the context of overall mechanisms of iron acquisition by the body.     

Identification, retinoid binding, and x-ray analysis of a human retinol-binding protein

Two cellular retinol-binding proteins (CRBP I and II) with distinct tissue distributions and retinoid-binding properties have been recognized thus far in mammals. Here, we report the identification of a human retinol-binding protein resembling type I (55.6% identity) and type II (49.6% identity) CRBPs, but with a unique H residue in the retinoid-binding site and a distinctively different tissue distribution. Additionally, this binding protein (CRBP III) exhibits a remarkable sequence identity (62.2%) with the recently identified iota-crystallin/CRBP of the diurnal gecko Lygodactylus picturatus [Werten, P. J. L., R?ll, B., van Alten, D. M. F. & de Jong, W. W. (2000) Proc. Natl. Acad. Sci. USA 97, 3282-3287 (First Published March 21, 2000; 10.1073/pnas.050500597)]. CRBP III and all-trans-retinol form a complex (K(d) approximately 60 nM), the absorption spectrum of which is characterized by the peculiar fine structure typical of the spectra of holo-CRBP I and II. As revealed by a 2.3-A x-ray molecular model of apo-CRBP III, the amino acid residues that line the retinol-binding site in CRBP I and II are positioned nearly identically in the structure of CRBP III. At variance with the human CRBP I and II mRNAs, which are most abundant in ovary and intestine, respectively, the CRBP III mRNA is expressed at the highest levels in kidney and liver thus suggesting a prominent role for human CRBP III as an intracellular mediator of retinol metabolism in these tissues.     

Lymphoid cell subsets in normal human small intestine

Nineteen specimens of normal tissue were obtained from the duodenum, upper jejunum and terminal ileum. Specimens were fixed with periodate-lysine-paraformaldehyde and then frozen sections were made. Lymphoid cell subsets were identified by indirect immunoperoxidase staining using mouse anti-human monoclonal antibodies. Results: There was a remarkable variation in the distribution of positive cells among individuals and even within single sections. Lymphoid cell subsets of the three parts of the small intestine were almost identical except for the difference in IgA subclasses of IgA-containing cells. IgA1 predominated over IgA2 in the jejunum while IgA2 predominated over IgA1 in the ileum. The subsets in the lamina propria (LP) and in the epithelium were clearly distinct; Leu 3+ (helper/inducer T) predominated in LP while Leu 2+ (suppressor/cytotoxic T) predominated in the epithelium and no Ig-containing cells were found in the epithelium. In LP the mean ratio of Leu 1+ (pan T):IgA (IgA1 + IgA2) containing cells: IgM: IgG: IgD: IgE: HNK-1+ was 48.9: 36.6: 9.0: 1.8: 1.0: 0.3: 2.4. The staining patterns of HLA-DR+ and Leu 10+ were similar; dense in the top in LP of the villi. In some individuals the apical portion of epithelial cells was stained by anti-HLA-DR. A few positive cells were observed in only two out of 19 cases by anti IL-2R.     

Immunochemical detection of metallothionein in specific epithelial cells of rat organs.

The distribution of a heavy metal binding protein, metallothionein, was studied immunocytochemically by using antimetallothionein antibody and the immunoperoxidase staining technique on histological sections of liver, kidney, intestine, lung, and testis from cadmium-treated rats. These tissues either accumulate heavy metals (e.g., liver, kidney, and testis) or are exposed to metal by ingestion or inhalation (intestine and lung). Staining for metallothionein was observed intracellularly in epithelial parenchymal cells of the liver and kidney; all hepatocytes and most renal tubular cells stained for the protein. Accumulation of metallothionein was not seen in connective tissue cells surrounding either blood vessels or renal tubules. Extracellular localization of metallothionein was also observed in the liver sinusoids and within the lumina of the renal tubules, suggesting a metal transport or excretory function for this protein. Surface columnar epithelial cells of the intestinal villi indicated the presence of metallothionein but connective tissue cells of the lamina propria were negative for the protein. The granular secretory Paneth cells of the small intestine also stained strongly for metallothionein as did respiratory epithelial cells of the lung. In the testis, metallothionein was detected in the Sertoli cells and interstitial cells but not within the spermatogonia. Sertoli cells are closely associated with the developing spermatogonia and appear to serve a nutritive role in spermatogenesis. Because of the secretory, absorptive, or nutritive function of the metallothionein-localizing cells in the organs studied, we suggest that metallothionein may be involved in metal storage or transport in addition to its commonly proposed detoxification role.     

A fluorescence confocal endomicroscope for in vivo microscopy of the upper- and the lower-GI tract

BACKGROUND: This report describes the development and the clinical evaluation of a novel confocal endomicroscope for obtaining fluorescence images of cellular morphology of the mucosae of the upper- and the lower-GI tract in vivo. The work assessed the feasibility of performing in vivo microscopy at endoscopic examination and evaluated fluorescence imaging protocols. METHODS: Images were collected in real time by using two prototype endoscope configurations, featuring slightly different miniaturized fiber-optic confocal microscopes, fitted integrally into the tips of conventional endoscopes. Confocal scanning was performed at 488 nm illumination for excitation of exogenously applied fluorophores (topical acriflavine and intravenous fluorescein). The images were compared with conventional histology of biopsy specimens and the findings of white-light endoscopy. RESULTS: Confocal endomicroscopy enabled imaging of cellular and subcellular structures (i.e., nuclei) of the GI tract. The crypts of the colonic mucosa, the villi of the terminal ileum and duodenum, the gastric pits of the stomach, and the squamous epithelium of the distal esophagus could be clearly visualized. Acriflavine strongly contrasted the cell nuclei of the surface epithelium, including the absorptive epithelial cells and the mucous secreting goblet cells. Fluorescein stained the extracellular matrix of the surface epithelium and also the subepithelial layers of the lamina propria. Images at increasing depth beneath the epithelium showed the mucosal capillary network. The findings correlated with the histology of biopsy specimens. CONCLUSIONS: The development of a fluorescence confocal endomicroscope makes it practical to examine the upper- and the lower-GI mucosa in cellular detail during otherwise routine endoscopic examination. The results represent a major technical advance in the development of this new optical imaging modality for the in vivo examination of GI tissue.     

Ubiquitous expression of HLA-DR antigens on human small intestinal epithelium

To clarify the frequency of the expression of HLA-DR antigens on the human small intestinal epithelium and the variation of such expression, 43 specimens (duodenum 16, jejunum 8, ileum 19; histologically normal 33, pathologic lesions 10) were studied for HLA-DR expression by indirect immunoperoxidase staining using monoclonal antibodies against HLA-DR antigens. All specimens reacted with anti-HLA-DR antibodies with minor variations. Most specimens showed decreasing intensity from the top of the villi toward the base of the villi and no staining in the crypt. In about half of the specimens, clear peripheral and cytoplasmic staining of the epithelial cells was observed. In some cases, the staining of the apical portion of the epithelium was patchy. The expression was unrelated to the lymph follicle. This ubiquitous expression of HLA-DR antigens on the small intestinal epithelium presents a striking contrast to an absence of the antigens on the normal large intestinal epithelium. A preliminary study on the pathologic lesions demonstrated the following changes in the expression of HLA-DR antigens; appearance of HLA-DR antigens on the crypt epithelium in Crohn's disease, loss of HLA-DR antigens on the epithelium covering the area of cell infiltration in macroglobulinemia and covering adenoma in familial polyposis coli.     

Localization of cellular retinol-binding protein and retinol-binding protein in cells comprising the blood-brain barrier of rat and human.

Brain is not generally recognized as an organ that requires vitamin A, perhaps because no obvious histologic lesions have been observed in severely vitamin A-deficient animals. However, brain tissue does contain cellular vitamin A-binding proteins and a nuclear receptor protein for retinoic acid. In the present study, immunohistochemical techniques were used to determine the cell-specific location of cellular retinol-binding protein in human and rat brain tissue. Cellular retinol-binding protein was localized specifically within the endothelial cells of the brain microvasculature and within the cuboidal epithelial cells of the choroid plexus, two primary sites of the mammalian blood-brain barrier. In addition, autoradiographic procedures demonstrated binding sites for serum retinol-binding protein in the choroidal epithelium. These observations suggest that a significant movement of retinol across the blood-brain barrier may occur.     

Plasma and cellular retinoid-binding proteins and transthyretin (prealbumin) are all localized in the islets of Langerhans in the rat.

The immunohistochemical localization of plasma retinol-binding protein (RBP), cellular retinol-binding protein (CRBP), and transthyretin (TTR) was studied in rat pancreas. The studies employed antibodies purified by immunosorbent affinity chromatography, permitting the specific staining and localization of each antigen by the unlabeled peroxidase-antiperoxidase method. Specific immunostaining for each of these three proteins was found localized to the islets of Langerhans. Both RBP and CRBP were localized in cells that were peripherally distributed within the islets, with an anatomic distribution that resembled that of the glucagon-containing A cells. Immunoreactive TTR was localized in cells that were more centrally distributed in the islets, with an anatomic distribution that resembled that of the insulin-containing B cells. These findings were confirmed by radioimmunoassay of a homogenate of isolated rat islets. By using sensitive and specific radioimmunoassays for each antigen, unusually high levels of CRBP, RBP, TTR, and cellular retinoic acid-binding protein (CRABP) were found in rat islets. The physiological significance of the localization of RBP, CRBP, CRABP, and TTR in the islets is not known. The findings suggest that retinoids and their binding proteins may play important metabolic roles within islet cells, and hence that they may be involved in some way in the biological, endocrine, function of the islets.     

Distribution of vitamin D-dependent calcium-binding protein messenger ribonucleic acid in rat placenta and duodenum

The vitamin D-dependent calcium-binding protein (CaBP), cholecalcin or calbindin, is one of the best documented molecular expressions of 1,25-dihydroxyvitamin D, the hormonal metabolite of vitamin D. In this report, DNA/RNA hybridization assays have been used to examine cholecalcin (CaBP) mRNA production in the placenta and duodenum of 21-day pregnant rats. A cloned CaBP cDNA which codes for the rat intestinal 9000 mol wt cholecalcin (9KCaBP) was radiolabeled and used in hybridization assays to explore 1) the size and relative quantities of CaBP mRNA extractable from placenta and duodenum by molecular hybridization, and 2) the localization and quantification, by in situ hybridization histochemistry, of CaBP mRNA in specific cells in rat placenta and duodenum. Northern hybridization studies show that the [32P]cDNA sequence hybridizes to a single 500- to 600-nucleotide species in the placenta as in the duodenum and, therefore, demonstrate identical 9KCaBP mRNA processing in both tissues. Dot blot hybridization studies show that the concentration of 9KCaBP mRNA was greatest in the duodenum, while that of the inner (fetal) placenta was about 50% the duodenal level. Considerably less CaBP mRNA was found in the outer (maternal) placenta. The observed differences in 9KCaBP mRNA levels correlate well with the in vivo variations in 9KCaBP concentrations. In situ hybridization histochemistry using [3H]cDNA reveals that 9KCaBP mRNA visualized by silver grains was concentrated in the inner placenta over the cytoplasm of syncytial cells in the trophoblastic epithelium of the labyrinth and much less frequently in the cells of the outer placenta. In the duodenum, 9KCaBP mRNA was found only in the absorptive epithelial cells from the crypt region to the upper part of the villi. The silver grains were distributed throughout the cytoplasm of the columnar cells; they were densest in the perinuclear region and rarest in the nuclear region. The concentration was greater in the cells at the villous tips than in those of the crypts. This difference in 9KCaBP mRNA levels correlates well with the distribution of the protein itself along the villi. CaBP mRNA quantities detected by hybridization histochemistry showed greater labeling in the syncytial cells of the trophoblastic epithelium of the labyrinth than in the absorptive epithelial cells of the upper part of the villi (200% less), indicating accumulation of CaBP mRNA at a greater rate in the trophoblastic epithelium than in the absorptive epithelial cells. These results indicate that in the rat, 9KCaBP is synthesized in both the absorptive cells of the duodenum and the cells of the trophoblastic epithelium of the placenta.     

Determination of cellular retinol-binding protein in human hepatocellular and colorectal carcinomas by radioimmunoassay

A study was conducted to determine the tissue levels of cellular retinol-binding protein (CRBP), serum retinol-binding protein (RBP) and retinoids in hepatocellular carcinoma (HCC) and in colorectal adenocarcinoma. CRBP, which has a molecular weight of 14 900 daltons and an isoelectric point of 4.9, was purified from human liver. An antihuman CRBP antibody was raised in the turkey and further purified by immunosorbent affinity chromatography on CRBP-coupled Sepharose column. A radioimmunoassay for CRBP using this antibody was established. RBP was measured by an enzyme immunoassay and retinoids were measured by high-performance liquid chromatography analysis. Retinol levels in liver tumours were significantly decreased compared with those in respective non-cancerous adjacent tissues. Retinyl ester levels in liver tumours were also significantly decreased compared with those in the adjacent tissues. CRBP levels in liver tumours were significantly decreased compared with those in the adjacent tissues, whereas no significant difference was observed in the CRBP level between the colon tumours and adjacent colon tissues. RBP levels in liver tumours were also significantly decreased compared with those in the adjacent tissues. The decreased CRBP levels in liver tumours may, at least in part, account for the local deficiency of retinol in HCC, whereas colon tumours may grow independently, regardless of retinol status.     

Unique insights into the intestinal absorption,transit, and subsequent biodistribution of polymer-derived microspheres

Polymeric microspheres (MSs) have received attention for their potential to improve the delivery of drugs with poor oral bioavailability. Although MSs can be absorbed into the absorptive epithelium of the small intestine, little is known about the physiologic mechanisms that are responsible for their cellular trafficking. In these experiments, nonbiodegradable polystyrene MSs (diameter range: 500 nm to 5 µm) were delivered locally to the jejunum or ileum or by oral administration to young male rats. Following administration, MSs were taken up rapidly (≤5 min) by the small intestine and were detected by transmission electron microscopy and confocal laser scanning microscopy. Gel permeation chromatography confirmed that polymer was present in all tissue samples, including the brain. These results confirm that MSs (diameter range: 500 nm to 5 µm) were absorbed by the small intestine and distributed throughout the rat. After delivering MSs to the jejunum or ileum, high concentrations of polystyrene were detected in the liver, kidneys, and lungs. The pharmacologic inhibitors chlorpromazine, phorbol 12-myristate 13-acetate, and cytochalasin D caused a reduction in the total number of MSs absorbed in the jejunum and ileum, demonstrating that nonphagocytic processes (including endocytosis) direct the uptake of MSs in the small intestine. These results challenge the convention that phagocytic cells such as the microfold cells solely facilitate MS absorption in the small intestine.     

Age related increase of brush border enzyme activities along the small intestine.

Intestinal morphology and brush border hydrolase activities were determined along the small intestine of young adult (three months, n = 10), mature (12 months, n = 10), and senescent (29 months, n = 15) rats. The intestinal segments of the senescent rats contained higher mucosal mass and protein content (p less than 0.05) compared with the young and mature animals. A significant reduction of villus height and crypt depth (p less than 0.05) was found in the proximal intestine during aging. A 35% increase in villus height (p less than 0.05) without changes in crypt depth, was observed in the distal ileum in senescent rats. The activities of sucrase and isomaltase were significantly increased during aging in the duodenum and jejunum (p less than 0.05). Lactase and aminopeptidase activities which showed only minor changes between young and mature animals were significantly enhanced in senescent animals (p less than 0.05) with aminopeptidase exhibiting a three-fold increase in activity in the proximal ileum. The results when combined with those of previous studies suggest that in the aged animal, the increased level of intestinal hydrolase activities may be the consequence of prolonged cellular maturation along the villi in the proximal intestine, and of adaptation to increased concentrations of intraluminal substrates in the distal intestine.     

The development of the fetal rat intestine and its reaction to maternal diabetes. I. Morphometrical analysis in normal conditions

In rodents, the fetal intestine develops rapidly during the last 5 days of gestation. The present investigation describes the events which occur in the duodenum, jejunum and ileum of fetal Wistar rat from day 16.5 to 21.5. The first villi and microvilli as well as endocrine cells already appear at 17.5 days in the duodenal mucosae. Goblet cells are detected at 18.5 days. The structure of the intestinal mucosa at 21.5 days is similar to that of adults. The evolution was quantified by morphometric analysis. The external and inner circumference, the length of the villi profile and the increased absorption area due to the villi profile were measured. We demonstrated that the total enlargement of the luminal surface area due to the villi and the microvilli in the duodenum of the fetus at 21.5 days is similar to that in the adult duodenum. This morphometric analysis could be used to detect possible disturbances in the development of the fetal intestine.     

RNA原位杂交法动态观察弓形虫速殖子对BALB/c小鼠小肠黏膜的黏附及侵入

目的 RNA原位杂交法观察弓形虫速殖子黏附和侵入小肠黏膜的部位及时间。 方法 30只BALB/c小鼠随机分为两组,实验组（24只）每鼠灌胃感染2×104个弓形虫速殖子（悬于0.2 ml PBS）,对照组（6只）给予等量PBS。分别于感染后15 min、30 min、1 h、2 h、4 h和8 h各处死实验组小鼠4只和对照组小鼠1只,取十二指肠、空肠和回肠标本制备石蜡切片,并作RNA原位杂交。光学显微镜观察原位杂交切片,随机选取50个视野,计数所有观察视野内的虫体总数并计算平均数。 结果 侵入的弓形虫速殖子可位于小肠上皮细胞（吸收细胞、杯状细胞和内分泌细胞）的纹状缘、吸收细胞胞浆内或相邻吸收细胞间及固有层内。感染后15 min,黏附于空肠的速殖子数量（4.93±3.949）显著高于回肠（3.78±3.102） （P<0.05）;侵入空肠的速殖子数量（4.92±4.164）显著高于十二指肠（4.10±3.532）和回肠（3.68±3.301） （P＜0.05）。随着感染后时间的延长,黏附于各肠段的速殖子数量逐渐减少,而侵入的数量逐渐増多。与感染后15 min相比,感染后8 h,黏附于十二指肠（2.32±3.039）、空肠（3.15±3.241）和回肠黏膜（2.59±3.028）的速殖子数量均显著减少（P<0.05）,而侵入十二指肠（8.92±8.955）、空肠（9.11±6.667）和回肠黏膜（9.15±10.192）的速殖子数量均显著增加（P<0.05）。 结论 弓形虫速殖子对其所黏附的小肠上皮细胞无严格的选择性,但其侵入小肠的部位具有选择性,空肠为速殖子侵入的易感部位。     

Coeliac disease is associated with impaired expression of acyl-CoA-synthetase 5

Background and aims Coeliac disease and other disorders of the small intestine are associated with disturbances in mucosal architecture. The most severe injury to tissue architecture is villus atrophy. In coeliac disease, molecules reflecting the state of the villus architecture are not well characterized at present.Materials and methods Expression of acyl-CoA-synthetase 5 (ACS5) was studied in unaffected human small/large intestinal tissue and in coeliac disease using several methods including molecular techniques, as well as an in situ approach using a novel established monoclonal antibody directed against human ACS5.Results Strong expression, synthesis, and enzymatic activity of ACS5 were found in normal small intestinal mucosa compared with unaffected colon mucosa. In normal small intestine, ACS5 preferentially located to the epithelium covering villi. In coeliac disease, expression of ACS5 was regularly associated with differentiation of villi. Thus, ACS5 was found in the villus epithelium of the small intestine with coeliac disease of Marsh grades I, II, IIIa, or IIIb respectively. In Marsh grade IIIc coeliac disease lesions, strong expression of ACS5 was detectable neither in the surface epithelium nor in the epithelium lining hyperplastic crypts.Conclusion These data suggest that ACS5 is a very suitable marker molecule for the detection of villus atrophy in the small intestine.     

Immunocytochemical detection of leptin-like immunoreactivity in the chicken gastroenteric tract

Leptin is a hormone produced and secreted mainly by adipocytes, but also by other tissues such as placenta, brain, mammary, and pituitary glands. The gastric epithelium has also been reported as a source of leptin in mammals. In this study we examined the presence of leptin in the chicken gastroenteric tract by immunohistochemistry and Western blotting. Strong and widespread leptin-like immunoreactivity was observed in the mucosal epithelium of proventriculus plicae and in the epithelium of the complex compound glands. Numerous leptin-immunoreactive cells were found along lining epithelium of duodenum villi, while few leptin-immunoreactive cells were observed in the basal zone of duodenum glands. Many leptin-immunoreactive cells were found in the caeca at the basal zone of glands, while very few leptin-immunoreactive cells were found in the deep glandular structures of the large intestine. In the homogenates of chicken gastroenteric tract the protein detected using a human leptin-specific antibody had estimated molecular weight of approximately 15-16 kDa. To our knowledge, this is the first report demonstrating leptin-like protein distribution in the whole gastroenteric tract of bird. This finding constitute important data for the further understanding of the mechanism that regulate feeding behaviour in birds, farm animal for which the control of food intake and fatness are a high economic interest.