Expression Of Sucrase Research Articles

ChREBP deficiency prevents high sucrose diet-induced obesity through reducing sucrase expression.

Obesity appears to be a major contributing factor for many health problems. Effective treatments for reducing weight gain, other than caloric restriction and exercise, are limited. The consumption of sugars is a major factor in the development of obesity in part by stimulating the transcription factor, carbohydrate response element binding protein (ChREBP), a process that is driven by de novo lipogenesis. Therefore, we hypothesized that inhibiting the action of ChREBP would be a promising strategy for alleviating these diseases. Using ChREBP deficient mice, the effect of a high intake of sucrose on body weight and blood glucose levels were investigated. Unlike wild type mice, ChREBP deficient mice did not gain much weight and their blood glucose and cholesterol levels remained relatively constant. In tracing it's cause, we found that the levels of expression of sucrase, an enzyme that digests sucrose, and both Glut2 and Glut5, a transporter of glucose and fructose, were not induced by feeding a high sucrose diet in the small intestine of ChREBP deficient mice. Our findings suggest that the inhibition of ChREBP could suppress weight gain even on a high sucrose diet.

S-layer protein 2 of vaginal Lactobacillus crispatus 2029 enhances growth, differentiation, VEGF production and barrier functions in intestinal epithelial cell line Caco-2

We have previously demonstrated the ability of the human vaginal strain Lactobacillus crispatus 2029 (LC2029) for strong adhesion to cervicovaginal epithelial cells, expression of the surface layer protein 2 (Slp2), and antagonistic activity against urogenital pathogens. Slp2 forms regular two-dimensional structure around the LC2029 cells,which is secreted into the medium and inhibits intestinal pathogen-induced activation of caspase-9 and caspase-3 in the human intestinal Caco-2 cells. Here, we elucidated the effects of soluble Slp2 on adhesion of proteobacteria pathogens inducing necrotizing enterocolitis (NEC), such as Escherichia coli ATCC E 2348/69, E. coli ATCC 31705, Salmonella Enteritidis ATCC 13076, Campylobacter jejuni ATCC 29428, and Pseudomonas aeruginosa ATCC 27853 to Caco-2 cells, as well as on growth promotion, differentiation, vascular endothelial growth factor (VEGF) production, and intestinal barrier function of Caco-2 cell monolayers. Slp2 acts as anti-adhesion agent for NEC-inducing proteobacteria, promotes growth of immature Caco-2 cells and their differentiation, and enhances expression and functional activity of sucrase, lactase, and alkaline phosphatase. Slp2 stimulates VEGF production, decreases paracellular permeability, and increases transepithelial electrical resistance, strengthening barrier function of Caco-2 cell monolayers. These data support the important role of Slp2 in the early postnatal development of the human small intestine enterocytes.

Chronic Effects of an Olive Leaf Extract on Sucrose Hydrolysis and Transport in the Caco‐2/TC7 Model of the Small Intestine

Growing evidence from both animal models and human intervention studies indicates that polyphenols can potentially attenuate postprandial glycemic responses and improve insulin sensitivity. A possible mechanism of action is through the inhibition of α‐glucosidases, critical for carbohydrate digestion in the gut, and by attenuation of glucose and fructose transport across small intestinal enterocytes. Sugar intake has increased over the last few decades, in part due to “hidden sugars” in foods, and a recent study demonstrated that excess sugar availability is a significant determinant of diabetes risk [1]. The research here focuses on the gut brush border enzyme sucrase which hydrolyses sucrose into glucose and fructose prior to absorption. We hypothesized that decreased expression or activity of sucrase would reduce the glycemic response to sucrose, potentially leading to positive effects on the glycemic response. The objective of the research is to evaluate if an extract from olive leaf, rich in the polyphenol oleuropein, chronically affects sucrose hydrolysis and transport.Fully differentiated Caco‐2/TC7 cells were used as a model for the small intestine and were exposed chronically to 1.5 mg/mL olive leaf extract (1.1 mM oleuropein) for the final 72 h of differentiation. This concentration is theoretically achievable in the gut lumen after consumption of the supplement “Bonolive”. An in vitro assay was developed using the cells as a source of sucrase to determine effects on activity and kinetic parameters. After treatments, sucrase mRNA was determined in whole cell lysates using droplet digital PCR, apical sucrase protein was quantitated after cell surface biotinylation using automated quantitative capillary Protein Simple Western blotting. Transport experiments were performed using cells cultured on porous supports, and the apical and basolateral concentrations of glucose and fructose were quantified using high performance anion exchange chromatography with pulsed amperometric detection.Olive leaf extract treatment reduced the Vmax of sucrase by 34%, decreased the specific activity by 32% and reduced total sucrase mRNA by 75% (all p<0.001). There was no change in total sucrase protein, but the proportion at the cell surface was lowered by 41% (p<0.001). Chronic treatment also affected sucrose hydrolysis and transport of the products; both apical glucose and fructose were attenuated by 37% and 21% respectively, in agreement with the data on sucrase activity. No change in glucose transport was observed when glucose was used as the substrate.In conclusion, chronic treatment by oleuropein‐rich olive leaf extract reduced the hydrolysis of sucrose. These results warrant future investigation into the use of olive leaf extract in humans for glycemic control after sugar consumption.Support or Funding InformationEuropean Research Council Advanced Grant “POLYTRUE?” (grant agreement 322467) and European Union Seventh Framework Programme (FP7/2007‐2013) “BACCHUS” (grant agreement 312090). Bonolive was kindly supplied by BioActor BV, The Netherlands. The Caco2/TC7 cell line was a kind gift by Prof. Monique Rousset, Centre de Recherche des Cordeliers, Paris, France.

Sodium/glucose cotransporter-1, sweet receptor, and disaccharidase expression in the intestine of the domestic dog and cat: two species of different dietary habit

The domestic cat (Felis catus), a carnivore, naturally eats a very low carbohydrate diet. In contrast, the dog (Canis familiaris), a carno-omnivore, has a varied diet. This study was performed to determine the expression of the intestinal brush border membrane sodium/glucose cotransporter, SGLT1, sweet receptor, T1R2/T1R3, and disaccharidases in these species adapted to contrasting diets. The expression (this includes function) of SGLT1, sucrase, maltase and lactase were determined using purified brush border membrane vesicles and by quantitative immunohistochemistry of fixed tissues. The pattern of expression of subunits of the sweet receptor T1R2 and T1R3 was assessed using fluorescent immunohistochemistry. In proximal, middle, and distal small intestine, SGLT1 function in dogs was 1.9- to 2.3-fold higher than in cats (P = 0.037, P = 0.0011, P = 0.027, respectively), and SGLT1 protein abundance followed an identical pattern. Both cats and dogs express T1R3 in a subset of intestinal epithelial cells, and dogs, but not cats, express T1R2. In proximal and middle regions, there were 3.1- and 1.6-fold higher lactase (P = 0.006 and P = 0.019), 4.4- and 2.9-fold higher sucrase (both P < 0.0001), and 4.6- and 3.1-fold higher maltase activity (P = 0.0026 and P = 0.0005), respectively, in the intestine of dogs compared with cats. Dogs have a potential higher capacity to digest and absorb carbohydrates than cats. Cats may suffer from carbohydrate malabsorption following ingestion of high-carbohydrate meals. However, dogs have a digestive ability to cope with diets containing significant levels of carbohydrate.

Morphological and biochemical alterations in the jejunum following iodoacetamide-induced colitis in rats

This study aims to describe the morphological alterations in the small and large intestines as well as the expression of some enterocyte enzymes and carriers in a rat model of iodoacetamide-induced colitis. Biopsies from the large and small intestines were taken at 1, 2, 4, 8, and 16 days postinduction and studied by light microscopy. The expressions of lactase, sucrase, aminopeptidase, and Glut-5 in the jejunum were studied by immunohistochemistry. Gene expressions of enterocyte lactase and sucrase were determined by RT-PCR using specific oligonucleotides. Microscopic examination of the large intestines revealed manifestations concordant with inflammation. Such alterations peaked at 2 days, were maintained to a lesser extent for 4 days, regressed by 8 days, and healed by 16 days. In the jejunum, the expression of lactase, sucrase, and aminopeptidase decreased 2 days after colitis induction, and recovered 2 days later. Similarly, Glut-5 expression decreased transiently with partial recovery by day 8. Compared with sham, gene expression of jejunal brush border enzymes sucrase and lactase showed a 4-fold increase in lactase and a 9-fold increase in sucrase after 4 days. We conclude that colitis can induce significant functional abnormalities in distant noninflamed small bowel regions.

Activity of hepatocyte nuclear factor 1α and hepatocyte nuclear factor 1β isoforms is differently affected by the inhibition of protein phosphatases 1/2A

Phosphorylation/dephosphorylation processes are known to control the activity of several transcription factors. The nutrition-dependent expression of sucrase–isomaltase and Na+/glucose co-transporter 1, two proteins implicated in the intestinal absorption of glucose, has been shown to be closely related to modifications of hepatocyte nuclear factor 1 (HNF1) activity. This study was conducted to determine whether phosphorylation/dephosphorylation processes could control HNF1 activity. We show that expression of the gene encoding sucrase–isomaltase is inhibited in the enterocytic Caco-2 clone TC7 by okadaic acid at a concentration that is known to inhibit protein phosphatases 1/2A and that does not affect cell viability. At the same concentration, phosphorylation of the HNF1α and HNF1β isoforms is greatly enhanced and their DNA-binding capacity is decreased. The phosphorylation state of HNF1β isoforms directly affects their DNA-binding capacity. In contrast, the decreased DNA-binding activity of the HNF1α isoforms, which was observed after the inhibition of protein phosphatases 1/2A, is due to a net decrease in their total cellular and nuclear amounts. Such an effect results from a decrease in both the HNF1α mRNA levels and the half-life of the protein. This is the first evidence for the implication of protein phosphatases 1/2A in the control of the activity of HNF1 isoforms. Moreover, these results emphasize a physiological role for the balance between phosphatases and kinases in the nutrition-dependent regulation of HNF1-controlled genes.

Expression of sucrase and intestinal-type alkaline phosphatase in colorectal carcinoms in rats treated with methylazoxymethanol acetate

In this study the small-intestine phenotype in rat colonic tumors was investigated in terms of sucrase and intestinal-type alkaline phosphatase (I-ALP) activity. F344 rats were given intraperitoneal injections of methylazoxymethanol acetate at a dose level of 25 mg/kg body weight once a week for 8 weeks and were killed 40 weeks after the first injection. Sucrase and I-ALP activities in proximal and distal colon adenocarcinomas were significantly higher than those in the normal colon epithelium. In the jejunum, by contrast, normal tissue had significantly higher levels than tumors. Immunohistochemical staining of I-ALP was also strong in striated cell borders of colon adenocarcinoma cells. These data suggest that, whereas absorptive cells of the small intestine lose their own traits with tumor development, colonocytes acquire phenotypic features of the small intestine. Intestinal enzymes associated with the striated-cell border, such as sucrase and I-ALP, may be useful markers for malignant phenotypic expression in colonocytes.

Increased expression of sucrase and intestinal-type alkaline phosphatase in human gastric carcinomas with progression.

The activities of sucrase, total alkaline phosphatase (total ALP) and intestinal‐type alkaline phosphatase (I‐ALP) were assayed in gastric carcinomas and in their surrounding mucosae from 57 patients with advanced cancers, and the localization of sucrase in 203 carcinomas, including 86 early cancers, was examined immunohistochemically using polyclonal anti‐sucrase antibody. All three enzymes were active in the 57 carcinomas as well as in their surrounding mucosae, but the levels were fairly low as compared to those in normal jejunum mucosa. A considerable part of the total ALP activity in tumor specimens was assumed to be due to I‐ALP itself. Increased sucrase and I‐ALP were found with greater depth of invasion by undifferentiated‐type carcinomas. The pattern of immunohistochemical localization of sucrase in the 203 carcinomas also clearly indicated increased expression with greater depth of invasion even in differentiated‐type carcinomas.

Regulation of sucrase and lactase in developing rats: Role of nuclear factors that bind to two gene regulatory elements

BACKGROUND & AIMS: Sucrase-isomaltase and lactase-phlorizin hydrolase expressions change remarkably during postnatal development in rats. The aim of this study was to explore the role of transacting nuclear proteins, proteins that bind to the SIF1 cis-regulatory element of the sucrase-isomaltase gene and to the CE-LPH1 cis-regulatory element of the lactase-phlorizin hydrolase gene, in this regulation.METHODS: Enzyme activity, Northern analysis, and electrophoretic mobility shift assays were used to study the relationship of these nuclear proteins to sucrase-isomaltase and lactase-phlorizin hydrolase gene expression in rats during development.RESULTS: A rapidly migrating low-molecular- weight SIF1-binding protein was found in suckling animals without sucrase-isomaltase messenger RNA (mRNA), and a higher-molecular-weight- binding protein was found in older animals with expression of sucrase- isomaltase mRNA. Supershift experiments and Western analysis showed that neither protein is Cdx-2, the only previously described SIF1- binding protein. CE-LPH1-binding protein was found only in adult animals (with low lactase activity), and there was no relationship between enzymatic activity and levels of lactase-phlorizin hydrolase mRNA.CONCLUSIONS: SIF1-binding proteins may regulate sucrase- isomaltase expression during postnatal development, but CE-LPH1-binding proteins do not seem to regulate lactase-phlorizin hydrolase expression during this period.(Gastroenterology 1997 Mar;112(3):803-12)

Epidermal Growth Factor Selectively Enhances Functional Enterocyte Adaptation after Massive Small Bowel Resection

After massive small bowel resection, the intestine adapts to compensate. In addition to proliferation, enterocytes also undergo selective functional adaptation. In this study we examined the effect of intraperitoneal administration of epidermal growth factor (EGF) on the expression of the brush border dissacharidase sucrase, the sodium glucose cotransporter (SGLT1), and the sodium–potassium ATPase pump (NaK ATPase) by enterocytes in the remnant intestine after massive small bowel resection. Adult Lewis rats underwent either ileal transection or 70% proximal intestinal resection. These animals were subdivided into groups that received either saline or EGF intraperitoneally for 1 week. Ilea from each group were harvested 4 weeks postoperatively. Enterocytes were separated from these segments by calcium chelation. The total protein from the isolated cells was subjected to Western blot analysis. Administration of EGF to animals that underwent transection did not significantly alter the expression of sucrase, SGLT1, or NaK ATPase. After intestinal resection, the expressions of sucrase and SGLT1 were significantly increased. The combination of EGF administration and intestinal resection resulted in a further increase in SGLT1 expression. The intraperitoneal administration of EGF selectively enhanced the expression of SGLT1 by enterocytes after massive small bowel resection. Administration of EGF to sham-operated animals did not have similar effects. These results suggest that EGF augments the adaptive response and may therefore have a therapeutic role in the management of patients with short bowel syndrome.

Sucrase-isomaltase gene expression in suckling rat intestine: hormonal, dietary, and growth factor control.

The effect of hydrocortisone, sucrose, and epidermal growth factor (EGF), alone or in combination, was investigated on the precocious expression of sucrase activity and sucrase-isomaltase (SI) mRNA in normal and adrenalectomized (ADX) suckling rats. In normal rats, each regulatory factor administered separately from 12 to 13 days of age induced a parallel expression of sucrase activity and SI mRNA. A potentiation of both sucrase activity and SI mRNA was observed after administration of hydrocortisone plus sucrose. Only SI mRNA was potentiated after EGF-sucrose administration or EGF-hydrocortisone administration. However, the potentiation of the activity was obtained after a prolonged treatment period of 4 days. In adrenalectomized rats, sucrose and EGF were still able to induce precocious induction of SI mRNA and sucrase activity, but sucrase activity was lowered by 75% by sucrose and by 30% by EGF. The combined treatments gave results similar to those obtained in normal rats. The results obtained in ADX rats indicate that the effects of sucrose and EGF on the precocious expression of sucrase and the potentiation of SI mRNA are independent of endogenous glucocorticoids. The rate of [3H]thymidine incorporation into mucosal DNA of normal sucklings was similar after single or combined treatments. From this study, we conclude that the precocious induction of intestinal sucrase by various regulatory factors, alone or in combination, is primarily regulated at the level of mRNA and is independent of increases in cellular proliferation and circulating glucocorticoids.

Triiodothyronine Stimulates the Expression of Sucrase–Isomaltase in Caco-2 Cells Cultured in Serum-Free Medium

In a previous study we have shown that triiodothyronine (T3) added to a serum-free medium supplemented with insulin, transferrin, and selenous acid (ITS) can stimulate Caco-2 cell differentiation. In this study we have focused on the effects of T3on sucrase activity. The results obtained demonstrate that T3(50 nM) does not change Caco-2 cell proliferation but enhances sucrase activity from 50 to 80%. Similar increases were observed whether or not insulin was present in the culture medium, showing that there was no synergistic effect between T3and insulin on sucrase activity. Moreover, T3acts specifically during the differentiation period since addition of T3to the defined TS medium before confluency is reached does not stimulate sucrase activity. Sucrase kinetic parameters were evaluated for the first time in Caco-2 cells under various culture conditions. The presence of a single enzyme was verified, with aKmof about 7 mMand aVmaxaround 20 nmol of substrate hydrolyzed min−1mg−1of protein. Our results showed that T3did not change the enzyme's affinity for sucrose but doubled theVmax. Moreover, immunoblotting using anti-sucrase–isomaltase (SI) antibodies revealed an approximately twofold increase in the relative amount of SI immunoreactive protein in T3-stimulated cells compared to untreated cells. Results obtained by both Northern hybridization and RT-PCR amplification showed a significant increase in SI mRNA contents. These results suggest that T3acts primarily on sucrase expression at the mRNA level.

Small intestinal injury in a neonatal rat model of giardiasis is strain dependent

Background & Aims : The factors that determine the severity of giardiasis are poorly understood. Host factors are important, but parasite virulence may also play a role. The aim of this study was to compare the apparent virulence of three genotypically different Giardia isolates (P01, VNB3, and WB). Methods : Infection rates, parasite loads, structural damage, disaccharidase activity, and water and electrolyte absorption were observed at 10 days after inoculation in a neonatal rat model of infection. Results : DNA fingerprinting showed differences between isolates studied. The infective rate varied between 67% and 100%. There were no differences in intestinal parasite load. Infection with strains P01 and WB, but not with VNB3, was associated with a reduction in villus height. There was precocious expression of sucrase at 10 days after inoculation in all infected groups. Water absorption of a plasma electrolyte solution was decreased in VNB3-infected animals when compared with P01- and WB-infected animals and controls. Water absorption and lactose hydrolysis were impaired during perfusion with a lactose-containing solution in all infected groups. Conclusions : Three genotypically different Giardia isolates that infect neonatal rats with the same trophozoite load differ in their ability to cause functional mucosal damage. Infection with Giardia lamblia induced precocious expression of sucrase activity and impaired mucosal absorption

Maternal state affects intestinal changes of rat pups at weaning

Two treatments of hooded rat (Rattus norvegicus) mothers after the birth of their pups affected the stage of development at which their pups started to take solid food and showed an increase in the expression of sucrase, an enzyme needed for digesting solid food. The pups of mothers that became pregnant in a post-partum oestrus showed the changes significantly earlier than those in the control group (Fig. 1A, B). The pups of mothers that had been mildly food-restricted after birth showed the changes significantly later than those in the control group. In brief, the pups of the pregnant mothers weaned first, the control pups next and those of food-restricted mothers weaned last. Differences in the timing of weaning between the three groups occurred in the absence of differences in pup body weight at the time of weaning. Within each experimental group, on the day in which sucrase expression showed the first detectable increase, sucrase activity was strongly predicted by the weight of the pups shortly after birth (Fig. 3). Pregnant mothers put on weight rapidly before implantation and their pups prepared for early weaning even though they did not differ in body weight from control pups (Fig. 4A, B). Food-restricted mothers were significantly lighter than control mothers and, with lighter pups before weaning started, settled for a longer period of suckling.

Effect of Ethanol on Human Colon Carcinoma CaCo‐2 and HT‐29 Cell Lines during the Maturation Process

The aim of the study was to ascertain whether the exposure to ethanol of human colon carcinoma CaCo-2 and HT-29 cell lines affects the differentiation process. As an index of enterocytic differentiation, the expression of sucrase, alkaline phosphatase, alpha 2,6-sialyltransferase toward the N-acetyllactosaminic sequence, and beta 1,4-N-acetylgalactosaminyltransferase (beta 1,4GalNAc-transferase) was examined. The latter enzyme is responsible for the biosynthesis of Sda carbohydrate histo-blood antigen, which mainly occurs in human colonic cells; its expression in CaCo-2 cells depends strictly on the enterocytic differentiation. The addition of ethanol in the culture medium resulted in a significant increment of sucrase and alpha 2,6-sialyltransferase activities in both cell lines, as well as the beta 1,4GalNAc-transferase activity in CaCo-2 cells and alkaline phosphatase activity in HT-29 cells. The increment was dose-dependent in the range between 50 and 200 mM ethanol and evident after 2 days of exposure in both cell systems. These results support the notion that, as occurs for cell lines of different origin, the ethanol in vitro positively affects the differentiation of intestinal cells, namely along the enterocytic lineage. The putative mechanism by which ethanol interferes with the maturation process of colonic cells is discussed.

REGULATION OF INTESTINAL SUCRASE EXPRESSION IN SUCKLING MOUSE INTESTINE

Intestinal disaccharidase expression in the developing post-natal animal is under a complex system of regulatory mechanisms involving pre-programmed genetic timing as influenced by circulating humoral factors, paracrine factors, and luminal nutrients. The aim of this study was to examine the relative contribution of circulating glucocorticoids and pre-programmed genetic expression of sucrase and lactase in a suckling mouse between post-natal days 13 and 15. We examined primary jejunal and ileal organ culture explants in serum-free medium at 0-2 hours and 22-24 hours of organ culture, respectively. Animals were studied at days 13, 14, and 15, with an additional group of adrenalectomy animals who underwent adrenalectomy on day 13 with organ culture at day 15. The results of sucrase activity are shown below.In summary, day 14 tissues in organ culture for 24 hours reached the same level of expression in organ culture of scrum-free media as in the intact animal on day 15. In addition, adrenalectomy on day 13 did not abolish the time-dependent expression of sucrase. However, the magnitude of expression in the basal state was diminished. We speculate that normal quantitative expression of intestinal disaccharidase in the 15-day suckling mouse is dependent upon a combination of intrinsic tuning in conjunction with circulating glucocorticoids.

Perinatal expression of brush-border hydrolases in rat colon: hormonal and tissue regulations.

The evolution pattern of brush-border digestive hydrolases and their hormonal regulation were studied in the proximal colon of newborn rats. The potentiality of the colon to express a small intestinal enzymatic pattern was also examined in associations made up of colonic endoderm and small intestinal mesenchyme, developed as either intracelomic grafts in 3-day-old chick embryos or as intrarenal grafts in adult rats. A transient increase of lactase- and aminopeptidase-specific activities occurred in the colon from the 19th day of gestation to 14 days after birth, but sucrase activity could never be detected. Immunocytochemical studies with antibodies specific for rat lactase, aminopeptidase, and sucrase confirmed these results. However, the levels of hydrolase activities were lower in the colon than in the jejunum at the same age. Thyroxine or hydrocortisone treatment during the first 4 days postpartum decreased lactase activity by 70 and 30%, respectively, but did not affect aminopeptidase activity. A slight but significant induction of sucrase activity was obtained with both hormones. In contrast, in the jejunum, only thyroxine decreased lactase activity with a lesser effect (30%), but both hormones increased aminopeptidase activity and induced the marked well-known appearance of sucrase activity. The fetal small intestinal mesenchyme was not able to induce the colonic endoderm to achieve a small intestinal-like differentiation. But the exposure of the developed hybrid intestines to glucocorticoids in organ culture allowed expression of sucrase in one-third of the cases. These results demonstrate the presence of brush-border hydrolases in the proximal colon of newborn rats, normally expressed in the small intestine, but never in the adult colon.(ABSTRACT TRUNCATED AT 250 WORDS)

Induction of intestinal differentiation by systemic and not by luminal corticosterone in adrenalectomized rat pups.

Potential effects of corticosterone (cort) from maternal milk on intestinal differentiation were studied using adrenalectomized (adx) rat pups fed a formula containing differing amounts of cort. Formula cort concentrations in the range found in milk (0.1-0.5 micrograms/ml) increased the survival of adx rats, but did not induce intestinal differentiation. Formulae containing 1.0-50.0 micrograms/ml cort caused a dose-dependent elevation of serum cort concentrations and jejunal maltase activity and precocious jejunal sucrase induction. Adx rats receiving the formula containing 10 micrograms/ml cort showed serum cort concentrations similar to those in day 15 control rats and jejunal sucrase and maltase activities equivalent to those in day 18-20 control rats, suggesting that the rise in serum cort that occurs during postnatal development suffices to modulate intestinal differentiation. To distinguish between direct local and systemic effects of luminal cort on intestinal differentiation, expression of sucrase was determined in jejunal isografts and in host jejunum from adx rats fed luminal cort. Jejunal isografts and host jejunum expressed similar sucrase activities and showed similar immunofluorescent staining. Moreover, administration of cort by continual ip infusion was more effective than continual intragastric infusion in inducing intestinal sucrase and maltase activities. These data imply that luminal cort is absorbed and transported into the systemic circulation before inducing intestinal epithelial cell differentiation through the systemic route.

50 CORRELATION BETWEEN THE ONTOGENY OF SMALL INTESTINAL MUCOSAL LYMPHOCYTES AND SUCRASE IN THE PREWEANED RAT

Immunoreactive sucrase in villous enterocytes of rat jejunum appears by 21 days of age. We attempted to correlate the ontogeny and type of intraepithelial and lamina propria lymphocytes during the same period. Suckled Sprague-Dawley rat pups (0 · 24 days) were sacrificed at regular intervals. The distal jejunum was removed and 4· to 5·micron frozen sections were cut, fixed in absolute methanol, and stained with mouse monoclonal antibodies BBC 1/35 (sucrase), MRC OX-4 (B-cell), MRC OX-8 (T-suppressor), W3/25 (T-helper), MRC OX-19 (pan T-cell) and W3/13HLK (pan T-cell). The sections were counterstained with goat anti-mouse IgG-FITC conjugated antiserum. The sections were then stained with Evans blue for morphologic definition and evaluated for type, relative numbers and location of lymphocytes. T-suppressor cells were present in the intraepithelial and lamina propria areas up to age 12 days and were absent between 12 and 17 days. From 17 to 24 days, T-suppressor cell levels increased and approached adult levels. B-cells were present at all ages in the lamina propria of villi and crypts. T-helper cells were found irregularly; pan T-cell levels increased from 21 days onward. Our observations confirm and extend those of Lyscom and Brueton (Clin Exp Immunol 54:158-162, 1983) in which the number of T-suppressor intraepithelial cells increased after 21 days of life. We report that this increase coincides with the expression of sucrase by villous enterocytes.

Effects of intestinal adaptation on insulin binding to villus cell membranes.

Insulin affects the expression of brush border enzymes by villus cells in vitro and in vivo. Physiological (lactation) and surgical (jejunoileal bypass) models of hyper- and hypoplasia were established so that insulin receptor characteristics could be related to villus histology, expression of sucrase and alkaline phosphatase, and plasma insulin concentrations. In lactating rats, villus height increased up to 55% (p less than 0.005), and fasting plasma insulin increased 71% (p = 0.005), compared with controls. Insulin binding to villus cell membranes, and sucrase and alkaline phosphatase activities were, however, unchanged. In ileum of bypass operated rats, villus height increased 134% (p less than 0.005) while insulin binding fell 68% (p = 0.025). Scatchard analysis revealed that this was largely because of reduction in binding by high affinity receptors. Sucrase and alkaline phosphatase specific activities fell 57% (p = 0.03) and 49% (p = 0.02) respectively, suggesting that ileal villus cells were hypomature. The slightly hypoplastic tissue of selfemptying loops showed normal insulin binding compared to jejunum of sham operated controls. Bypass and sham operated rats had similar fasting plasma insulin concentrations. Reduced insulin binding in markedly hyperplastic gut of bypass operated rats might reflect hypomaturity of villus cells. The reduction in insulin binding, however, might significantly modulate the effect of insulin on small intestinal mucosa and account for the fall in enzyme activity which occurs despite villus hyperplasia.