Medium-chain triglycerides enhance mucous secretion and cell proliferation in the rat

Background  The specific purpose of this study was to investigate the effects of medium-chain triglycerides (MCTs) on intestinal cell proliferation and mucous secretion of the small intestine in the rat. Methods  Rats were fed chow diet and given MCTs or the same weight of corn oil (5 g/kg per day) by gavage daily for 2 weeks, and then tissue samples of the small intestines were harvested. Leptin concentration in the small intestine was measured. Cell proliferation and apoptosis in the small intestine was determined by immunohistochemistry. Diamine oxidase (DAO) activity was measured by colorimetric assay. Results  In rats fed only chow diet (normal rats), the number of goblet cells per villi was 14.2 ± 0.75 in the jejunum and 15.2 ± 1.12 in the ileum. The number of goblet cells increased significantly in rats given MCTs compared with rats given corn oil or normal rats. Ki-67-positive cells were detected on the entire villi and the crypts in the small intestine. Furthermore, the proliferative index and the apoptotic index were also significantly greater in rats given MCTs than rats given corn oil or normal rats. Moreover, DAO activity and leptin concentration in the small intestine were significantly greater in rats given MCTs than rats given corn oil or normal rats. Conclusions  MCTs enhance cell proliferation of the intestinal epithelium and mucous secretion from goblet cells in the small intestine. These effects may protect the gut in patients suffering from inflammatory bowel disease or enterogenous infection.     

Rat intestinal mucosal responses to a microbial flora and different diets.

The effects of diet on the histochemical composition of intestinal mucosubstances and the morphology of the villi and crypts were investigated by comparing the data of germ free and conventionally maintained rats fed either a purified diet or a commercial diet. The influence of intestinal microflora was evaluated by comparing the germ free rats and those harbouring either a conventional rat flora or a human microbial flora. In both germ free rats and those maintained conventionally, feeding a purified diet resulted in shallower crypts in the small intestine but deeper crypts in the large intestine compared with their counterparts fed on the commercial diet. The preliminary data obtained with association of human flora showed a reduction of the villus height and crypt depth in the small intestine and, to some extent, the amount of neutral mucins in the goblet cells of both small and large intestine and an increase in the amount of sulphated mucins in the large intestine. In rats given the commercial diet the periodic acid Schiff staining for neutral mucins was more intense in the upper crypts of the small intestine than in the lower crypts, and to a lesser extent in the upper crypts of the large intestine. These results provide evidence that the dietary composition, microbial flora, as well as the interactions between the dietary constituents and microbial flora change the mucosal architecture and the mucus composition and therefore alter the functional characteristics of the intestinal tract.     

Fetal characteristics of small intestinal crypt cells.

Nine monoclonal antibodies were prepared against luminal membranes purified from rat intestinal cells at day 19 of gestation, and seven of them were found to define antigens common to adult crypt cells and fetal or embryonic intestinal epithelial cells. The FBB 2/29 antigen was first detected over the entire intestinal epithelial population at days 14-15 of gestation, a period of development characterized by formation of a stratified intestinal epithelium and differentiation of the surrounding mesenchyme. This antigen, identified as a set of high molecular mass proteins, became restricted to the crypt and lower villus cells after birth and was exclusively expressed by the crypt cells in adult intestine. It also was found to be expressed by the epithelial cells of the distal tubuli in the kidney of adult rats and by cultured human tumor colonic cells. The FBB 1/54/1, FBB 3/46, and FBB 3/78/9 antibodies stained only the epithelial cells present at the base of the villi in fetal intestine, starting at days 20-21 of gestation (about 1-2 days before birth), and stained the crypt and lower villus cells in newborn and adult intestine; these antigens may be regarded as specific markers for the developing crypt cells in fetal intestine shortly before birth. The FBB 1/20 and FBB 4/2 antigens were first detected on the fetal intestinal cells at day 18 of gestation; they were located over the entire epithelium in newborn rats and became restricted to the crypts after weaning. The FBB 2/28 antigen was expressed by the entire intestinal epithelium at all stages of development, starting from days 18-19 of gestation in the fetus. Two antibodies, FBB 3/4 and FBB 3/24, were found to be specific for lactase. These results have demonstrated the expression of cell- and tissue-specific components in rat intestine during early embryonic development and revealed a marked similarity in surface membrane antigens between fetal intestinal epithelial cells and adult crypt cells.     

E2 prostaglandins modulate cell proliferation in the small intestinal epithelium of the rat.

Groups of Sprague-Dawley rats were administered 1 mg/kg indomethacin subcutaneously, indomethacin subcutaneously plus 200 micrograms/kg oral 15-R-15 methyl-prostaglandin E2 (MePGE2) or oral MePGE2 twice daily for 10 days. The animals were treated with antibiotics to prevent mortality. Two control groups were used: control 1 was given placebo and control 2 was treated with antibiotics. All rats were killed 4 h after injection of a metaphase blocker, and the proliferative activity of the distal small intestine was examined in histological sections by means of the cumulative mitotic index (MI). A reduction in the number of villous cells was observed in the rats given antibiotics (p < 0.05 vs. control 1). The small intestinal villi of the rats treated with indomethacin had fewer cells than those of both control groups (p < 0.05) whereas the crypts contained more cells (p < 0.05) and had a higher MI than those of the controls (p < 0.05 vs. controls 1 and 2). These changes were reverted by the prostaglandin analogue. The number of cells of the small intestinal crypts and the cumulative MI in the rats who received indomethacin and the prostaglandin analogue were similar to controls, and they were significantly lower than the values observed in the animals treated with indomethacin (p < 0.05). The animals treated with the prostaglandin analogue and placebo developed a marked hyperplasia of the small intestinal villi (p < 0.05 vs. both control groups), but the atrophy of the villi induced by indomethacin was not prevented by simultaneous administration of the analogue.(ABSTRACT TRUNCATED AT 250 WORDS)     

Postnatal Epithelial Growth of the Small Intestine in the Rat Occurs by Both Crypt Fission and Crypt Hyperplasia

Studies of growth of the small intestine have largely concentrated on crypt hyperplasia rather than crypt fission. The aim of this study was to investigate quantitatively both crypt fission and crypt hyperplasia. DAxPVG/c rats were killed at 7, 11, 14, 17, 19, 21, 25, 55, and 72–73 days of life. Samples of jejunum at one third of the intestinal length were taken for morphometry (villous area, crypt area, percentage of bifid crypts, and crypt mitotic count) by microdissection. Growth factors and their receptors were assessed by oligonucleotide microarray. Crypt fission was 10.5%, 5.2%, and 1.5% at days 11, 25, and 72–73 of life, respectively. Crypt hyperplasia increased from day 21. No conventional growth factor was identified during crypt fission. We conclude that crypt fission contributes to growth of the small intestine prior to weaning and crypt hyperplasia to growth after weaning.     

Effect of fasting and refeeding on basolateral polyamine uptake and metabolism by the rat small bowel.

Fasting reduced the weight, protein, DNA, RNA and polyamine contents of the small intestine of rats, but its effects on the in vivo uptake of intraperitoneally injected 14C-spermidine through the basolateral membrane of the small intestine were small. The uptake of putrescine was nearly doubled by fasting for 48 h. Fasting for 48 h had reduced villus length but was without effects on the crypts. Refeeding for 6 h of rats fasted for 48 h led to hypertrophic growth: the length of both crypts and villi increased by about 50% without changes in cell number. The uptake of spermidine by the small intestine increased above not only that in fasted rats but also that in the controls fed ad libitum. The high putrescine uptake of rats fasted for 48 h was unchanged after refeeding for 6 h, but returned to control values after 12 h. Spermidine in the gut was well conserved, while most of the putrescine was transformed into non-polyamine metabolites. It is concluded that refeeding stimulates basolateral spermidine uptake, and this may be a general mechanism for polyamine accretion in adaptive growth of the small intestine.     

Changes in the structure and regeneration mode of the rat small intestinal mucosa following benzalkonium chloride treatment

Tritiated thymidine was administered IP to rats that had been exposed to benzalkonium chloride in the duodenum, jejunum, and ileum, resulting in neuronal ablation. Epithelial cell proliferation and migration were studied 21 and 7 days after treatment. Significant hyperplasia and hypertrophy of the villi and crypts was seen from day 7 on. This was half as pronounced as that of the muscle layer, whose maximal percent increase was not seen until day 21. In the crypt, the proliferation had increased significantly (65% 3H index corrected) and its zone had expanded proportionally to the total crypt depth. After an average of 36 hours in the ileum (48 hours in normal rats), labeled cells reached the tip of the lengthened villi, reflecting significantly accelerated migration. Concerning the distributional pattern of the labeled cells in the crypt, a nonsignificant shift to the lower two thirds of the crypt could be distinguished. From this the author concludes that treatment with benzalkonium chloride influences the proliferation and migration of the epithelial cells in the treated area. These alterations may result from loss of the myenteric plexus, but other factors cannot be excluded.     

Initial kinetic changes of prostaglandin E2-induced hyperplasia of the rat small intestinal epithelium occur in the villous compartments

This study was performed to further identify the sequence of cell kinetics that occurs in the development of gastric and intestinal epithelial hyperplasia after orally administered prostaglandins of the E series. A high-dose, short-treatment schedule was used to examine the initial effects on kinetic parameters in the rat small intestinal epithelium. Groups of rats were killed after a single dose of oral prostaglandin E2 at 1 h after in vivo labeling with [methyl-3H]thymidine and during continued treatment at 6, 12, 24, 48, 72, and 96 h. As evidenced by autoradiography, the earliest change produced by prostaglandin E2 was an increased cellularity of the villous compartment (p less than 0.05 after 24 h). There was no change of labeling index of the villous compartment or of the leading edge of labeled cells within 24 h. At 48 h, the increased cellularity was accompanied by a significantly elevated labeling index of the villi. Throughout the study period no significant differences were observed between groups in the number of cells or labeling indices in the jejunal crypts, or in cellular input from the crypts to the villi. Epithelial turnover time in the placebo and treatment groups was 69 and 71 h, respectively. To exclude the possibility that prostaglandin E2 initially affects cell birth rate and mean cell cycle time, a metaphase blocker was given after 4 days of treatment in a second study. Animals were killed after 0, 0.5, 1.0, 1.5, and 2.5 h. The rate of entry into mitoses was 8.1% cells/h in controls compared with 8.2% cells/h in treated rats. The distribution of mitoses within crypts was identical in the two groups and the mean cell cycle time was 13.6 and 13.2 h, respectively. Also in this study there were trophic changes of the villi. It is concluded that the hyperplasia produced by oral prostaglandin E2 starts in the villi of the small intestine and is initiated by reduced cell exfoliation from the villous tips. Previously recorded retention of cellular elements in villi and crypts, increased cellularity of the proliferative compartments, and reduced mitotic index are secondary events.     

Mechanism of Excretion of Iron by the Small Intestine: an Experimental Study in Rats with Normal and Abnormal Mucosae

The findings in the present study have demonstrated the presence of two distinct mechanisms of iron excretion by the small intestine in rats. Loss of cell bound iron is the principal pathway of excretion and takes place as epithelial cells are shed off from the tips of villi. The available evidence suggests that the epithelial cells of the mucosa sequestrate iron from a storage pool which might be identical to the labile iron store in man. Transferrin bound iron makes a small contribution to the total loss. An abnormally large amount of iron was excreted in the small intestine of rats infested with Nippostrongylus brasiliensis. At the same time an unduly large amount was also sequestrated by the intestines of these rats. These facts were attributed to increased population of maturing cells in the crypts and the rapid rate of renewal of the villous epithelium.
The morphological appearances of the intestinal villi of the infested rats have a resemblance to those in human subjects with coeliac disease. It is, therefore, suggested that these observations may have some bearing on the status of iron balance in patients suffering from coeliac disease.     

Localization of carbonic anhydrase activity in the developing rat colon

Carbonic anhydrase activity was localized histochemically by light and electron microscopy in the proximal and distal colon of developing rats. Fixed tissue was taken for normal morphology and carbonic anhydrase localization from fetal (20-22 days gestation), suckling (1-19 days postnatal), weanling (20-25 days postnatal), and adult rats. The proximal colon had distinct villi at birth which were diminished between days 5 and 11 postnatally. The distal colon lacked villi at birth but had rudimentary crypts (ridges and furrows) which were replaced during the suckling period by a flat mucosa interspersed with true crypts. Carbonic anhydrase first appeared in both proximal and distal colonic epithelial cells on the day of birth (22 days gestation). Goblet cells were nonreactive at each developmental period. In neonatal rats, epithelial cells in the upper half of the villi of the proximal colon and on the surface and upper crypts of the distal colon were positive for carbonic anhydrase throughout the cytoplasm. Cells at the villar base (proximal colon) or in the deep crypt (distal colon) had reaction product in the intercellular spaces but not the cytoplasm. By 11 days postnatal, cytoplasmic reaction product was present in proximal colonic cells in the upper three-fourths of the crypt and was concentrated in a heavy band in the apical cytoplasm. In the distal colon, cytoplasmic positive cells did not extend as deeply into the crypts and the apical banding pattern was weak. Intercellular spaces in the deeper crypt epithelium were positive in both proximal colon and distal colon, suggesting a membrane-bound carbonic anhydrase. It was concluded that carbonic anhydrase appeared suddenly at birth and was continuously present in mid- to upper-crypt (or upper villus in early neonatal proximal colon) non-goblet cells into adulthood. This suggests a functional role for carbonic anhydrase in chloride-bicarbonate exchange across the neonatal and adult colonic mucosa.     

Effect of dietary fat on the distribution of mucosal mass and cell proliferation along the small intestine.

This study investigated how substitution of long chain triglycerides for glucose in a mixed diet affects the overall small intestinal mucosal mass and the distribution of mucosal mass and cell proliferation along the small intestine. Four groups of eight female Wistar rats (180-200 g) were isocalorically fed mixed diets containing the essential fatty acid rich oil Efamol substituted for glucose at concentrations of 1.2%, 10%, 25%, and 50% total calories for 20 to 23 days. The small intestine was divided into three equal length segments and whole gut weights, mucosal weights, protein and DNA determined. Cell proliferation was estimated from the two hour accumulation of vincristine arrested metaphases in microdissected crypts at points 0%, 17%, 33%, 50%, 66%, and 100% small intestinal length. There were no differences between groups in parameters of overall small intestinal or distal segment mucosal mass. With increasing levels of fat, however, there was a significant trend for the mucosal mass of the proximal segment to fall and that of the middle segment to rise. The pattern of two hour metaphase accumulation reflected these changes. These regional changes in mucosal mass and cell proliferation may reflect differences in the sites of absorption of fat and glucose.     

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.     

Proglucagon-Derived Peptides in Intestinal Epithelial Proliferation

Proglucagon-derived peptides have been implicated in the control of intestinal mucosal cell division. To investigate the actions of these peptides on intestinal cell proliferation, different doses of enteroglucagon, oxyntomodulin, glucagon-like peptide-1 (GLP-1) and glucagon-like peptide-2 (GLP-2) were tested in male Wistar rats maintained on total parenteral nutrition. Crypt cell proliferation was assessed by the analysis of arrested metaphases in microdissected crypts. Enteroglucagon and oxyntomodulin had no effect on intestinal weight or cell proliferation. GLP-1 had a slight effect on stomach and small intestinal weights and on epithelial cell proliferation in the small and large intestines. GLP-2 infusion dose-dependently increased the weights of the stomach, small intestine, colon, and cecum and increased crypt cell proliferation in the small and large intestines of parenterally fed rats. In orally fed animals, GLP-2 increased intestinal weight but had little effect on proliferation. Therefore, of the proglucagon-derived peptides, GLP-2 appears to be a major mediator of intestinal epithelial proliferation.     

The effects of glutamine on intestinal epithelial cell proliferation in parenterally fed rats

BACKGROUND: Several papers have indicated that glutamine is a preferred fuel for the enterocyte and that it can increase intestinal epithelial cell proliferation. AIMS: To investigate the effects of glutamine on intestinal epithelial cell proliferation in the parenterally fed rat. METHODS: Five groups of six rats were fed parenterally; a group of orally fed rats was also studied. Crypt cell proliferation was studied after six days using native mitoses in microdissected crypts and bromodeoxyuridine labelling. RESULTS: No effect of treatment was seen on intestinal weight; however, the weights of the small intestine, caecum, and colon were all significantly heavier in the orally fed group than in the total parenteral nutrition groups (p<0.001). There was no effect of any of the glutamine treatments on mitotic activity in the small intestine. In the colon there was a small increase in native mitoses with glutamine (p=0.03). There was also an indication of increased proliferative activity in the first fifth of the small intestine and colon with glutamine. Little effect of glutamine on bromodeoxyuridine labelling in either site was observed, but there was a small but significant reduction in growth fraction of the colon of the glutamine treated group. The labelling distribution curve from sections and the mitotic distribution curve obtained from crypt squashes showed a good correlation. CONCLUSION: Glutamine has a small, but significant effect on mitotic activity but only in the colon. Modest effects on the distribution of labelled cells were also seen.     

Changes in motility after jejunal and ileal resection: electromyographic study in rats

The aim of this work was to compare the effects of massive jejunal and ileal resections on intestinal motility using an electromyographic technique. Male Wistar rats were used: in the first group a massive jejunal resection was performed, conserving a 7-cm segment after the ligament of Treitz; the rats of the second group underwent an ileal resection, preserving 7 cm of the terminal ileum. Motility was studied at the 10th and 30th postoperative days by means of electrodes implanted throughout the remaining bowel and was expressed by the pattern of recurrence of the migrating myoelectric complex (MMC). In a fasting state, in both transected and resected animals at the 10th postoperative day, the gradient in the duration of MMC along the intestine still existed. However, on the 30th postoperative day, in animals with jejunal resection only, there was an adaptive process: the duration of MMC in the remaining jejunum was significantly increased to the duration in the ileum. After the end of the postprandial inhibition of the appearance of the MMC, on the 10th postoperative day there was a significant decrease in the duration of MMC in the ileum in both types of resection, compared to the controls. However, on the 30th postoperative day, the duration of MMC returned to its control value. In conclusion, jejunal resection seems to induce more important adaptive processes in intestinal motility than does ileal resection. The different results are discussed.     

The Expression of Msi-1 and Its Significance in Small Intestinal Mucosa Severely Damaged by High-Dose 5-FU

OBJECTIVE: The purpose was to investigate the expression of musashi-1 (msi-1) and its significances in small intestinal mucosa that was severely damaged by high-dose 5-FU. METHODS: A total of 40 adult C57BL/6J mice were divided into two groups: the control group (n = 8, group A) and experimental group (n = 32). The mice in the control group were treated with PBS by intraperitoneal injection, and the other mice were treated with high-dose 5-FU (150 mg/kg body weight for 5 consecutive days) by intraperitoneal injection. At the 1st (group B), 3rd (group C) and 5th (group D) day after treatment with high-dose 5-FU, the dying mice were killed, HE staining and immunohistochemical techniques were used to detect the expression of the putative marker of intestinal epithelial stem cells, msi-1, in samples of the middle intestine from these mice, and the percentage of the msi-1-positive cells from the intestinal mucosal cells of the mice in group B was detected by FACS. RESULTS: After treatment with high-dose 5-FU, the intestinal mucosa suffered severe damage: the villi and crypts disappeared, the number of msi-1-positive cells increased greatly, the intestinal epithelial cells could be divided into two fractions by FACS, and the percentage of msi-1-positive cells was up to 67.75% in the fraction in which the value of FSC was higher. CONCLUSIONS: After treatment with high-dose 5-FU, the percentage of intestinal stem cells had increased significantly, which was useful for the further isolation and enrichment of intestinal epithelial stem cells.     

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.