Structural characteristics of the mucosa of the small intestine in gnotobiote rats]

Histological and electron microscopic investigations of mucosal strucutre of the small intestine in gnotobiotic and conventional rats have demonstrated that mucosal index (lenear dimentions of villi/crypt dimentions ratio) is equal to 3--4 and 1.4--2.5, respectively. In gnotobiotic rats the number of mitotic figures in crypts is less, migratory-desquamative parameters of enterocytes are 5--6 days, cranio-caudal gradient of linear dimentions of villi, crypts and the number of goblet-shaped cells is not expressed. The goblet-shaped cells and cells with acidophilic granules have secretion of moderate intensity. The number of cells infiltrating epithelium of villi and crypts is greater when microbes are present in the gut lumen. A considerable difference in the structure of the connective tissue basis of the gnotobiotic and conventional rats mucosa are also connected with presence or absence of microflora in the gut lumen.     

An early stimulation of solid feed intake slightly influences the morphological gut maturation in the rabbit

The impact of dietary factors on the gut morphological maturation is poorly documented in rabbits. The weights of the digestive segments as well as the morphology of villi and crypts along the small intestine were analysed weekly from day 14 till day 49, in two rabbit groups weaned at either 21 (W21 group, n = 12 litters) or 35 days (W35 group, n = 12 litters) of age. From 21 till 35 days, the W21 group ate 57% more solid feed than the W35 group (P < 0.01), and presented slighter body weights from day 28 till day 49 (-9%, P < 0.05). Tissue weights of the empty digestive segments, as expressed relative to the body weights, were higher in the W21 than in the W35 group from day 28 till day 49 (P < 0.001), whereas absolute tissue weights appeared similar (except for the proximal colon). From day 28 to day 49, small intestinal villi grew in height and surface area (P < 0.05) whereas the crypts deepened. Villous height followed a proximo-distal decreasing gradient from the duodenum to the ileum (P < 0.05) from day 28 onward. The villous height to width ratio changed with the beginning of significant solid feed intake: from a thin shape until day 21, villi became wider from day 28 on. The effect of weaning age on mucosal morphology was insignificant, except for the jejunal crypts whose surface area and depth were higher in the W21 group. The present results showed that morphological changes in the digestive tract of young rabbits were weakly influenced by an early stimulation of solid feed intake.     

Observation of mucosal pathology after praziquantel treatment in experimental Fibricola seoulensis infection in rats

It is well known that the duodenum of mice or rats infected with Fibricola seoulensis shows atrophy of villi (shortening, blunting, widening, fusion) and hyperplasia of crypts. This study was performed to observe healing process of these pathological changes after deworming with anthelmintic treatment. Albino rats infected each with 1,000 metacercariae of F. seoulensis were treated with single dose of 10 mg/kg praziquantel on day 15 post-infection. On day 1, 3, 5, 7, 15, 21 and 28 after the treatment, they were sacrificed and their duodenums were histopathologically studied. Control (uninfected) rats showed their normal finger-like projections of duodenal villi and well arranged crypts. In comparison, untreated (infected) controls revealed severe mucosal changes characteristic of villous atrophy and crypt hyperplasia in their duodenum. The damaged duodenal mucosa was found to restore its normal morphology after praziquantel treatment; until day 3 post-treatment the mucosa was severely atrophied; on day 5 long and slender villi sometimes appeared among the fused and stout ones; after day 15 the villi were in their normalizing process. From this experiment, it was shown that the mucosal changes in the duodenum of rats caused by F. seoulensis infection were completely reversible in 21-28 days after anthelmintic treatment.     

In vivo immediate early gene expression induced in intestinal and colonic mucosa by feeding.

Since the gut responds rapidly to food intake, the levels of expression of several immediate early genes were measured in mucosa from small and large intestine of rats starved for 3 days or refed. Within 1 h of refeeding, jejunal and ileal c-fos, jun B and zif/268 mRNA and colonic zif/268 dramatically increased. The zif/268 gene in jejunum corresponded in size to the full-length cDNA but, in ileum, an RNA band of about 1.2 kb in size increased greatly after feeding. This represents a physiologic in vivo model for the study of gene regulation associated with intestinal epithelial cellular responses to feeding.     

Correlation between methotrexate-induced intestinal damage and decrease in polyamine content

A synthetic analog of prostaglandin E(1), OP-1206 [17S, 20-dimethyl-trans-Delta(2)-prostaglandin E(1)] protects the small intestine from the methotrexate (MTX)-induced damage. The purpose of this study is to evaluate the protective effect of OP-1206 on the methotrexate-induced small intestinal damage in rats from the biochemical point of view. MTX (15 mg/kg body weight) was orally administered to rats once daily for 5 days. OP-1206 (0.5 microg/kg body weight) was orally administered to rats twice a day for 5 days, and on the 6th day biochemical components in the jejunal mucosa of the treated rats were determined. The contents of DNA, RNA, proteins and polyamines (spermine and spermidine) in the jejunal mucosa of rats were markedly decreased by the MTX treatment. The coadministration of OP-1206 with MTX prevented such decreases caused by the MTX treatment. The MTX treatment decreased the incorporation of 3H-thymidine into DNA in the jejunal mucosa, while the coadministration of OP-1206 with MTX prevented it. These results indicated that OP-1206 could protect the intestinal mucosa against the biochemical effects of MTX through a trophic action on intestinal villi. Further, it should be noted that polyamines may possibly play an important role of modulation action on intestinal mucosa.     

Protection of mouse jejunum against lethal irradiation by Podophyllum hexandrum

Radiation induced gastrointestinal damage occurs due to the destruction of the clonogenic crypt cells and eventual depopulation and denudation of the villi. P. hexandrum, a plant, known for its antitumour activity, has been shown to protect the mice against whole body lethal (10 Gy) irradiation. Present study was undertaken to investigate the radioprotective effect of P. hexandrum on jejunal villi cells, crypt cells, their proliferative capacity and mitigation of apoptosis.

In an in vivo micro colony survival assay, pre-irradiation administration of P. hexandrum (–2 h) increased the number of surviving crypts in the jejunum by a factor of 3.0 (P < 0.05) and villi cellularity by 2.7 (P < 0.05) fold in comparison to irradiated control. Pre-irradiation administration of P. hexandrum reduced the incidence of apoptotic bodies in the crypts (P < 0.05) in a time dependent manner and depicted a mitotic arrest till the 24 h. However, after 84 h the percentage of mitosis was observed to be nearly similar to that of unirradiated control.

This study suggests that arrest of cell division may help in protecting the clonogenic cells against radiation. It would be interesting to investigate further the role of P. hexandrum in influencing various cell cycle regulators like bcl-2, TGF-β, Cyclin-E etc.     

Morphological changes in basement membrane associated with jejunal graft injury

Jejunal mucosa injury and morphometric continuity of the intestinal epithelial basement membrane (BM) in experimental groups after 1 h and 6 h of reperfusion following intestinal heterotopic allotransplation in male Wistar rats were observed. Intestinal graft tissue was compared to the jejunum of recipients. Significant damage to the jejunal mucosa was detected and followed by complete intestinal villi destruction, damage to epithelial intestinal crypts and the surrounding lamina propria mucosae. There was a local interruption of BM continuity, as well as its absence in particular areas. The percentage of continuous, clearly visible and undamaged BM detected using the periodic acid and Schiff reagent (PAS) method was 22.4% in the graft samples taken 1 h after transplantation. In the same samples, 43.1% undamaged BM was visualised by silver impregnation. Disappearance of the BM was significant in comparison with recipient samples using both staining methods (both p < 0.001) Biopsies of the intestinal mucosa graft 6 h after transplantation showed significant reduction of the damage to the jejunal mucosa (p < 0.05 and p < 0.01, respectively). The percentage of continuous, clearly visible and undamaged PAS-positive BM was 54%, and argyrophilic BM rose to approximately 83%. The difference between BM continuity detected by PAS method was significant when comparing graft and recipient samples (p < 0.001), but impregnation method did not revealed significant distinction. Based on our results, the silver impregnation method seems to be more sensitive to damage assessment of the jejunal graft.     

Diet, mucosal architecture and epithelial cell production in the small intestine of specified-pathogen-free and conventional rats.

Upper jejunum and terminal ileum were examined in specified-pathogen-free (SPF), conventional and conventional after SPF rearing (ex-SPF) rats. The effect of 2 differential diets on the last 2 groups was examined. Ex-SPF rats had taller villi and deeper crypts than SPF rats, but similar crypt to villus ratios and cell production rates. Ex-SPF rats had similar crypt depth and jejunal villus height to conventional rats on the same diet, but taller ileal villi and a lower cell production rate. Even after 6-8 weeks, in a conventional environment, ex-SPF rat intestine was still not identical with conventional rat intestine. Diet had a significant effect on mucosal architecture, and a smaller effect on cell production rate. It is concluded that diet, microbiological status of colony of origin, and environment after weaning, can all affect mucosal architecture and epithelial cell production, and should be properly controlled in experimental studies.     

Liporotein particles in the jejunal mucosa of fetal rats.

The jejunal mucosa of fetal rats contains particles of the same size, electron density and distribution as particles identified by others in adult jejunum as very low density lipoprotein particles. These particles are first seen in 19-day fetuses at which time the jejunal mucosa is still relatively undifferentiated and formation of villi is just beginning. They become progressively more abundant during the last 3 days of gestation as differentiation proceeds and are seen in virtually all absorptive cells in the apical third of jejunal villi of fetuses at term.     

L-Glutamate Supplementation Improves Small Intestinal Architecture and Enhances the Expressions of Jejunal Mucosa Amino Acid Receptors and Transporters in Weaning Piglets

L-Glutamate is a major oxidative fuel for the small intestine. However, few studies have demonstrated the effect of L-glutamate on the intestinal architecture and signaling of amino acids in the small intestine. The aim of this study was to investigate the effects of dietary L-glutamate supplementation on the intestinal architecture and expressions of jejunal mucosa amino acid receptors and transporters in weaning piglets. A total of 120 weaning piglets aged 35±1 days with an average body weight at 8.91±0.45 kg were randomly allocated to two treatments with six replicates of ten piglets each, fed with diets containing 1.21% alanine, or 2% L-glutamate. L-Glutamate supplementation increased the activity of glutamate oxaloacetate transaminase (GOT) in the jejunal mucosa. Also, the mRNA expression level of jejunal mucosa glutamine synthetase (GS) was increased by L-glutamate supplementation. The height of villi in duodenal and jejunal segments, and the relative mRNA expression of occludin and zonula occludens protein-1 (ZO-1) in jejunal mucosa were increased by dietary L-glutamate supplementation. L-Glutamate supplementation increased plasma concentrations of glutamate, arginine, histidine, isoleucine, leucine, methionine, phenylalanine and threonine. L-Glutamate supplementation also increased the relative mRNA expression of the jejunal mucosa Ca²⁺-sensing receptor (CaR), metabotropic glutamate receptor 1 (mGluR1) and metabotropic glutamate receptor 4 (mGluR4), and neutral amino acid transporter B⁰-like (SLC1A5) in the jejunal mucosa. These findings suggest that dietary addition of 2% L-glutamate improves the intestinal integrity and influences the expression of amino acid receptors and transporters in the jejunum of weaning, which is beneficial for the improvement of jejunal nutrients for digestion and absorption.     

Glutamine and ketone-body metabolism in the gut of streptozotocin-diabetic rats.

1. In short- and long-term diabetic rats there is a marked increase in size of both the small intestine and colon, which was accompanied by marked decreases (P less than 0.001) and increases (P less than 0.001) in the arterial concentrations of glutamine and ketone bodies respectively. 2. Portal-drained viscera blood flow increased by approx. 14-37% when expressed as ml/100 g body wt., but was approximately unchanged when expressed as ml/g of small intestine of diabetic rats. 3. Arteriovenous-difference measurements for ketone bodies across the gut were markedly increased in diabetic rats, and the gut extracted ketone bodies at approx. 7 and 60 nmol/min per g of small intestine in control and 42-day-diabetic rats respectively. 4. Glutamine was extracted by the gut of control rats at a rate of 49 nmol/min per g of small intestine, which was diminished by 45, 76 and 86% in 7-, 21- and 42-day-diabetic rats respectively. 5. Colonocytes isolated from 7- or 42-day-diabetic rats showed increased and decreased rates of ketone-body and glutamine metabolism respectively, whereas enterocytes of the same animals showed no apparent differences in the rates of acetoacetate utilization as compared with control animals. 6. Prolonged diabetes had no effects on the maximal activities of either glutaminase or ketone-body-utilizing enzymes of colonic tissue preparations. 7. It is concluded that, although the epithelial cells of the small intestine and the colon during streptozotocin-induced diabetes exhibit decreased rates of metabolism of glutamine, such decreases were partially compensated for by enhanced ketone-body utilization by the gut mucosa of diabetic rats.     

同种异体大鼠骨髓间充质干细胞移植对放射性空肠损伤的修复作用

目的观察同种异体骨髓间充质干细胞（MSCs）移植对放射性空肠的修复作用。方法全骨髓贴壁体外培养大鼠MSCs并进行DAPI标记。对大鼠行5 Gy X线全腹部照射,每隔72 h照射1次,共5次,制备放射性空肠损伤动物模型。取40只大鼠,随机分成正常对照组、模型及MSCs治疗组及DAP标记组。激光共聚焦显微镜下观察DAPI标记的MSCs在空肠组织中富集情况。病理学观察MSCs对放射性空肠损伤的修复作用。结果正常大鼠空肠黏膜结构清楚,隐窝深遂,腺体丰富,模型组7 d黏膜上皮细胞坏死脱落,隐窝几乎完全破坏,治疗组7 d黏膜坏死组织较少,黏膜增厚,30 d治疗组核分裂相增加,较模型组增加明显。结论成功建立放射性空肠损伤动物模型,MSCs可以促进空肠再生与修复。     

Cell migration and cortisone induction of sucrase activity in jejunum and ileum

The increase of sucrase activity in homogenates of jejunum and ileum of suckling rats after cortisone administration has been investigated. Serial tissue sections of villi and crypts were also assayed for sucrase activity and these results were compared with the migration of cells labelled with [(3)H]thymidine along the villus. By using a low dose of cortisone (0.5mg/day per 100g body wt.) it was found that the sensitivity of the small intestine producing system to cortisone stimulation increased during the suckling period. On the other hand, 5mg of cortisone/day per 100g body wt. produced practically the same increase of sucrase during the entire suckling period. Sucrase activity in homogenates of the entire small-intestinal wall was first detected 24h after the first injection of cortisone (5mg/day per 100g body weight) to 9-day-old animals and maximum activity both in the jejunum and ileum was reached by 120h. Jejunal activity was greater than ileal activity, but the rate of the increase was similar. The half-time of the increase was 23-27h, whereas enterocytes migrate from the base to the tip of the villi in approximately 72h. Comparison of sucrase activity in serial tissue sections of villi and crypts at various times after cortisone treatment showed that the leading edge of sucrase activity proceeds toward the tip of the villi at the same rate as the advancing edge of newly formed cells. Sucrase activity increased in the newly induced cells as they migrated to the tip of the villi. It was concluded that the increase of sucrase activity in suckling rats after cortisone stimulation is due to at least three factors: (1) increase of activity in newly differentiating cells, (2) increased percentage of villus cells with sucrase activity and (3) continued production or activation of sucrase activity as the cells migrate along the villi.     

Trichostrongylus colubriformis: effects on villi and crypts along the whole small intestine in infected rabbits

In control and Trichostrongylus colubriformis-infected rabbits histological samples were taken from the small intestine at 25-cm intervals from the pylorus. Length of villi, surface of glands of Lieberkühn, and number of epithelial cells per unit length were measured with a digitized table. Two zones were so delimited along the parasitized mucosa: in the proximal part of the gut, both shortened villi and dilated crypts were found; in the distal region of the intestine, which was also the area with the lowest density of parasites, dilatation of crypts was associated with an increase in the size of villi. In both regions, the size of enterocytes was unchanged. Correlation coefficients calculated between worm burdens and relative variation in length of villi were significant and negative. Conversely, the same coefficients were significant and positive between worm burdens and surface of crypts. The possibility of local and general actions of Trichostrongylus colubriformis on both villi and crypts is evoked to explain the changes in the two zones. The functional importance of the hypertrophy in the distal region is still to be investigated during a T. colubriformis infection.     

Regulation of perfused capillary density in canine intestinal mucosa during endotoxemia.

When O2 delivery (blood flow X arterial O2 content) is reduced, many tissues respond by increasing perfused capillary density. This facilitates the increase in O2 extraction required to maintain tissue O2 consumption in the face of limited O2 supply. In a previous study of isolated canine small intestine (J. Appl. Physiol. 64: 2410-2419, 1988), endotoxin administration was associated with an impaired ability to increase O2 extraction in response to progressive reductions in O2 delivery. The aim of the present study was to determine whether reductions in perfused capillary density occur after endotoxin administration. Fourteen male dogs were anesthetized with chloralose (150 mg/kg iv) and urethan (750 mg/kg iv), and a segment of small intestine was exteriorized through a midline laparotomy. The segment was isolated vascularly, autoperfused, and maintained at body temperature. Escherichia coli endotoxin (5 mg/kg) or sham challenge was administered, and the animals were allowed to stabilize. Blood flow and arterial and gut venous blood O2 contents were measured after 3 h. Perfused vessels were then labeled by injecting colloidal carbon (less than 0.8 microns) through the arterial cannula and clamping the artery and vein as the bolus passed through the tissue. In some of the experiments a second gut segment was successfully obtained within 1 h of the first, yielding a total of 14 gut segments in nine endotoxin animals and nine segments in five control animals. Morphological analysis of capillary surface density in mucosal villi and crypts showed a significantly higher perfused capillary density in control tissue blocks (77.8 +/- 9.2%) than in blocks from endotoxin-treated animals (68.8 +/- 8.0%, P less than 0.04).(ABSTRACT TRUNCATED AT 250 WORDS)     

Persistence of gut mucosal innate immune defenses by enteric α-defensin expression in the simian immunodeficiency virus model of AIDS

Gastrointestinal mucosa is an early target of HIV and a site of viral replication and severe CD4(+) T cell depletion. However, effects of HIV infection on gut mucosal innate immune defense have not been fully investigated. Intestinal Paneth cell-derived α-defensins constitute an integral part of the gut mucosal innate defense against microbial pathogens. Using the SIV-infected rhesus macaque model of AIDS, we examined the level of expression of rhesus enteric α-defensins (REDs) in the jejunal mucosa of rhesus macaques during all stages of SIV infection using real-time PCR, in situ hybridization, and immunohistochemistry. An increased expression of RED mRNAs was found in PC at the base of the crypts in jejunum at all stages of SIV infection as compared with uninfected controls. This increase correlated with active viral replication in gut-associated lymphoid tissue. Loss of RED protein accumulation in PC was seen in animals with simian AIDS. This was associated with the loss of secretory granules in PC, suggesting an increase in degranulation during advanced SIV disease. The α-defensin-mediated innate mucosal immunity was maintained in PC throughout the course of SIV infection despite the mucosal CD4(+) T cell depletion. The loss of RED protein accumulation and secretion was associated with an increased incidence of opportunistic enteric infections and disease progression. Our findings suggest that local innate immune defense exerted by PC-derived defensins contributes to the protection of gut mucosa from opportunistic infections during the course of SIV infection.     

Regulation of Epithelial Differentiation in Rat Intestine by Intraluminal Delivery of an Adenoviral Vector or Silencing RNA Coding for Schlafen 3

Although we stimulate enterocytic proliferation to ameliorate short gut syndrome or mucosal atrophy, less effort has been directed at enterocytic differentiation. Schlafen 3 (Slfn3) is a poorly understood protein induced during IEC-6 enterocytic differentiation. We hypothesized that exogenous manipulation of Slfn3 would regulate enterocytic differentiation in vivo. Adenoviral vector coding for Slfn3 cDNA (Ad-GFP-Slfn3) or silencing RNA for Slfn3 (siSlfn3) was introduced intraluminally into rat intestine. We assessed Slfn3, villin, sucrase-isomaltase (SI), Dpp4, and Glut2 by qRT-PCR, Western blot, and immunohistochemistry. We also studied Slfn3 and these differentiation markers in atrophic defunctionalized jejunal mucosa and the crypt-villus axis of normal jejunum. Ad-GFP-Slfn3 but not Ad-GFP increased Slfn3, villin and Dpp4 expression in human Caco-2 intestinal epithelial cells. Injecting Ad-GFP-Slfn3 into rat jejunum in vivo increased mucosal Slfn3 mRNA three days later vs. intraluminal Ad-GFP. This Slfn3 overexpression was associated with increases in all four differentiation markers. Injecting siSlfn3 into rat jejunum in vivo substantially reduced Slfn3 and all four intestinal mucosal differentiation markers three days later, as well as Dpp4 specific activity. Endogenous Slfn3 was reduced in atrophic mucosa from a blind-end Roux-en-Y anastomosis in parallel with differentiation marker expression together with AKT and p38 signaling. Slfn3 was more highly expressed in the villi than the crypts, paralleling Glut2, SI and Dpp4. Slfn3 is a key intracellular regulator of rat enterocytic differentiation. Understanding how Slfn3 works may identify targets to promote enterocytic differentiation and maintain mucosal function in vivo, facilitating enteral nutrition and improving survival in patients with mucosal atrophy or short gut syndrome.     

Alterations in jejunal transport and (Na+-K+)-ATPase in an experimental model of hypoxia in rats

Hypoxia was induced by exposing rats to an atmosphere of 93% N2, 7% O2 for 4-48 hr. The animals became hypoxic as indicated by a decreased blood PaO2 (mean +/- SEM: 48 +/- 10 mm Hg). Hypoxia was accompanied by metabolic acidosis (pH 7.22 +/- 0.02) and decreased serum bicarbonate levels (9.0 +/- 4.0 meq/liter). Hypoxic rats also showed evidence of tissue hypoxia; liver tryptophan oxygenase levels were increased to 21 +/- 2 nmole/min/mg protein. In the hypoxic animals there was decreased jejunal mucosal (Na+-K+)-ATPase activity and an inhibition of active intestinal transport of sodium, glucose, 3-O-methylglucose, galactose, tyrosine, phenylalanine, and glycine as determined by in vivo perfusion studies. Jejunal fructose transport, which has a large passive component, was unaffected by hypoxia. The electrolyte, carbohydrate, and amino acid transport alterations produced by hypoxia were seen in the absence of an effect on jejunal cell number, DNA synthesis, or cell turnover. There was also no evidence of histological or ultrastructural damage. Furthermore, studies with a luminal macromolecular tracer, horseradish peroxidase, indicated that the jejunal lumen-to-blood barrier to macromolecules was also unaltered in these hypoxic animals. In vitro local oxygenation of the jejunum, by bubbling of 95% O2:5% CO2, markedly improved sodium and glucose (but not 3-O-methylglucose) absorption in hypoxic rats and control rats. The (Na+-K+)-ATPase activity of the jejunal mucosa of hypoxic rats was significantly enhanced by the local bubbling of 95% O2:5% CO2. Overall, our data indicate that during relatively mild conditions of hypoxia there is an inhibition of jejunal (Na+-K+)-ATPase activity and related transport processes that is prevented by in situ oxygenation.     

Differentiation and maturation of cultured fetal rat jejunum.

To determine whether differentiation and maturation of mammalian intestinal mucosa require influences available only in vivo or whether they can occur in vitro, fetal rat jejunum was cultured in chemically defined medium using organ culture methodology. Segments of jejunum from 18-day fetal rats were cultured in modified Liebowitz L-15 medium in room air at 37°C. Segments harvested after 24, 48, and 72 hr of culture were examined by light and electron microscopy. Uncultured jejunum from 18-day fetuses had either no or very few rudimentary villi and was lined largely by undifferentiated stratified epithelium. Goblet cells were not seen. In contrast, villi were present in the majority of 24-hr cultures, and simple columnar rather than stratified epithelium predominated. After 48 and 72 hr, villi were present in over 90% of cultured jejunal segments and stratified epithelium had disappeared. Goblet cells were seen in jejunal segments cultured 48 hr or longer in 47 of 74 fetuses. Electron microscopy further documented progressive differentiation of the epithelium during culture. Microvilli increased in number and height, a terminal web developed in the apical cytoplasm and the number of apical vesicles, mitochondria and formed elements of endoplasmic reticulum increased in absorptive cells. Jejunal lactase and alkaline phosphatase activity increased nine- and sevenfold, respectively, during 72 hr of culture, while the activity of the mitochondrial enzyme, ornithine carbamoyl transferase, increased fourfold. These observations indicate that jejunum from 18-day fetal rats can be cultured in vitro for at least 72 hr in chemically defined medium and that, during culture, maturation of the jejunal mucosa takes place with the appearance of villi, conversion of stratified to columnar epithelium and differentiation of individual epithelial cells.