Stimulation of disaccharidase activities in the jejunal brush border membrane of adult rat by total parenteral nutrition. Effects of thyroid hormones

This study examined the morphological and functional adaptations occurring in the jejunum of adult rats fed totally by parenteral nutrition during a 4-day period. Comparison was made with sham-operated animals receiving orally a similar isocaloric diet. The intravenously fed rats exhibited a 20% shortening of the villus height. The specific activity of aminopeptidase showed remarkable stability in all conditions. A major stimulation (2.5-fold) was measured for lactase-specific activity. Daily administration of thyroxine completely inhibited the rise of lactase activity. Thyroidectomy caused a significant increase of lactase activity in the orally fed controls, but did not exert any synergistic effect with parenteral nutrition on intestinal enzyme activities. In our experimental conditions intravenous feeding led to a 3-fold decrease in the concentration of thyroid hormones detected in the serum. The results show that total parenteral nutrition leads to a stimulation of the specific activity of brush border lactase in the intestine of adult rat which might be related to the level of thyroid hormones.     

Conjugated bile salts regulate turnover of rat intestinal brush border membrane hydrolases

The mechanisms whereby the conjugated bile salts regulate the activities of the brush border membrane hydrolases and its physiological significance were investigated in rat small intestine, and comparisons were made with the action of pancreatic protease. Rat brush border membrane proteins were metabolically labeled with [³⁵S]methionine, and isolated brush border membrane was incubated with taurocholate or pancreatic elastase. The activity of solubilized hydrolases was assayed and the molecular forms of the hydrolases were examined by SDS-PAGE. The activity and protein bands of alkaline phosphatase and sucrase-isomaltase were solubilized by taurocholate, while alkaline phosphatase was not solubilized by elastase. Solubilized sucrase-isomaltase molecules were proteolytically degraded by elastase, whereas the intact molecule of sucrase-isomaltase was solubilized by taurocholate. Next the physiological role of bile salts in brush border membrane hydrolase turnover were investigated using metabolic labeling of brush border membrane hydrolase and immunoprecipitation in biliary diversion rats. After three days of biliary diversion, a significant increase in alkaline phosphatase activity was observed. Although synthesis of alkaline phosphatase in biliary diversion rats was similar to that observed in control rats, biliary diversion rats showed 1.5-fold slower turnover of alkaline phosphatase when compared with control rats. These results suggest that conjugated bile salts in the intestinal lumen may cause a rapid turnover of brush border membrane hydrolases, which may be increased by the enhanced enzyme degradation. The mechanisms for the enhanced degradation appeared to be solubilization of hydrolases caused by the detergent activity of bile salts. Therefore, conjugated bile salts may play an important physiological role in the regulation of expression of the protease-resistant enzymes such as alkaline phosphatase.     

The locally acting glucocorticosteroid budesonide enhances intestinal sugar uptake following intestinal resection in rats

BACKGROUND AND AIMS: Locally and systemically acting corticosteroids alter the morphology and transport function of the intestine. This study was undertaken to assess the effect of budesonide, prednisone, and dexamethasone on sugar uptake. METHODS: Adult male Sprague Dawley rats underwent transection or resection of 50% of the middle portion of the small intestine, and in vitro uptake of sugars was measured. RESULTS: The 50% enterectomy did not alter jejunal or ileal uptake of glucose or fructose. Prednisone had no effect on the uptake of glucose or fructose in resected animals. In contrast, in resected rats budesonide increased by over 120% the value of the jejunal maximal transport rate for the uptake of glucose, and increased by over 150% ileal uptake of fructose. Protein abundance and mRNA expression of the sodium dependent glucose transporter in brush border membrane (SGLT1), sodium independent fructose transporter in the brush border membrane (GLUT5), sodium independent glucose and fructose transporter in the basolateral and brush border membranes (GLUT2), and Na(+)/K(+) ATPase alpha1 and beta1 did not explain the enhancing effect of budesonide on glucose or fructose uptake. Budesonide, prednisone, and dexamethasone reduced jejunal expression of the early response gene c-jun. In resected animals, expression of the mRNA of ornithine decarboxylase (ODC) in the jejunum was reduced, and corticosteroids reduced jejunal expression of the mRNA of proglucagon. CONCLUSIONS: These data suggest that the influence of corticosteroids on sugar uptake in resected animals may be achieved by post translational processes involving signalling with c-jun, ODC, and proglucagon, or other as yet unknown signals. It remains to be determined whether budesonide may be useful to stimulate the absorption of sugars following intestinal resection in humans.     

Enteral and parenteral feeding to evaluate malabsorption in intestinal parasitism.

General evidence of malnutrition such as loss in body weight associated with intestinal parasitism has been attributed to decreased food intake, to intestinal malabsorption, and to change in host basal metabolism. To establish the relative importance of these factors in this regard, rats with trichinosis were studied. The weights of infected and uninfected animals were followed after being placed on one of three feeding regimens for 1 week--stock diet ad libitum, intraduodenal nutrition, and intravenous nutrition. Infected rats on a stock diet lost weight whereas those on the other two regimens maintained the same weight pattern as uninfected counterparts. The maintainance of body weight occurred despite alterations at the level of the intestinal brush border as indicated by a depression of intestinal disaccharidase activities (sucrase and lactase) and by reduction of monosaccharide absorption (measured as accumulation of beta-methyl glucoside) in the proximal, heavily infected region of the small intestine. There was no compensatory increase in enzyme activity nor in the absorptive capacity in the distal gut. Results support the conclusion that inadequate oral food intake rather than changes in basal metabolism or intestinal pathophysiology accounts for weight loss during the intestinal phase of infection.     

Phosphate transport adaptation in rat jejunum and plasma level of 1,25-dihydroxyvitamin D3

Intestinal absorption of inorganic phosphate (Pi) increases in response to a reduction in the dietary supply of Pi. In this work this adaptive response has been characterized in jejunal brush border membrane vesicles and studied in temporal relationship with the change in the plasma level of 1,25(OH)2D3. The results indicate that in rat jejunal brush border membrane vesicles the activity of the sodium-dependent Pi transport system is stimulated by a low Pi diet. This adaptive response was the result of an increase in the Vmax and a reduction in the Km of the cotransport system. This change in Pi transport was correlated with an increase in the circulating level of 1,25(OH)2D3 in a time-related fashion. In conclusion, these results are consistent with the notion that Pi restriction leads to an increase in Pi transport activity in the luminal membrane of the intestine. A time course study suggests that the elevation in plasma 1,25(OH)2D3 might be involved in the adaptation of the intestinal Pi transport system to Pi restriction.     

Altered intestinal and renal brush border amino-oligopeptidase structure in diabetes and metabolic acidosis: normal and biobreed (BB) rats.

Amino-oligopeptidase (AOP, aminopeptidase N), a major glycoprotein hydrolase in intestinal and kidney brush border membranes, plays a crucial role in digesting peptide nutrients and salvaging filtered peptides. The molecular structure of rat intestinal and kidney AOP was compared for normal Wistar and congenitally diabetic BB Wistar (BBd) rats. Brush border membranes were isolated, solubilized with Triton X-100, and the AOP specifically immunoprecipitated with polyvalent rabbit antiserum and analyzed on 7% sodium dodecyl sulfate (SDS)-acrylamide electrophoresis. While the specific hydrolytic activity was maintained, BBd rats displayed an altered migration of AOP on SDS gels. Intestinal AOP migrated as a smaller species (130 kd) in the BBd than in the normal Wistar (135 to 140 kd). In some BBd rats, additional intestinal AOP species were observed (a 130- to 135-kd doublet or a 125-, 130-, or 135-kd triplet). Kidney AOP migrated as a broader band (125 to 140 kd) than intestine for all rat groups, probably due to carbohydrate chain heterogeneity, and was approximately 5 kd smaller in the BBd rat than in the normal Wistar. In contrast, no mass change was found in diabetes induced by streptozotocin (STZ). The altered intestinal AOP in the BBd rat was present when first inserted into the brush border membrane (6 hours after intraperitoneal [35S]methionine labeling), and hence was not due to nonenzymatic glycosylation (NEG). Abnormal intestinal and kidney AOP structure appeared in early diabetes, irrespective of high plasma glucose levels or ketoacidosis, and was reversed following evolution of the diabetes under prolonged (21 to 120 days) insulin treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Functional characterization of luminal enterocyte membranes of the small intestine mucosa using isolated brush border membranes

Atrophy of the small intestinal mucosa is functionally characterized by a reduction in non-electrolyte transport in vivo. In order to elucidate the cellular defect being responsible for this malabsorption, we have studied the Na+-dependent D-glucose accumulation as well as the activities of aminopeptidase M and maltase in brush border membrane vesicles prepared from jejunal self-emptying blind loops and corresponding intestinal segments of sham-operated control rats. Membrane vesicles from atrophic mucosa did not show any differences in D-glucose uptake or in enzyme activities when compared with those derived from normal intestine. Thus it is unlikely that the impaired non-electrolyte absorption in the atrophic mucosa in vivo is due to a defect in cellular transport processes. It is more probable that the functional impairment is the result of the diminished absorptive surface in this pathophysiological condition.     

The influence of age on intestinal dipeptidyl peptidase IV (DPP IV/CD26), disaccharidases, and alkaline phosphatase enzyme activity in C57BL/6 mice

The objective of this study was to determine and describe the age-related changes in intestinal brush border membrane enzyme activities that occur in C57Bl/6 mice. Specifically, jejunal, duodenal, and ileal dipeptidyl peptidase IV/CD26, disaccharidase (lactase, sucrase, and maltase), and alkaline phosphatase activities were determined. A significant correlation between analyzed intestinal brush border membrane enzyme activities and animal age was found. Our study revealed that intestinal dipeptidyl peptidase IV/CD26, lactase, sucrase, maltase, and alkaline phosphatase activities decline significantly with age (p < .05). Nevertheless, the horizontal (duodenum to ileum) enzyme activity patterns are not affected by age.     

The function of Gp170, the multidrug-resistance gene product, in the brush border of rat intestinal mucosa.

Gp170 is a transmembrane glycoprotein that is overexpressed in multidrug-resistant tumor cells and is present in the apical plasma membrane domain of small intestinal mucosal cells. The function of Gp170 was studied in the small intestine of the rat. Jejunal and ileal brush border membrane vesicles, but not basolateral membrane vesicles, manifested adenosine triphosphate (ATP)-dependent transport of daunomycin, a substrate for Gp170, and contained a approximately 170-kilodalton protein that reacts with anti-Gp170 monoclonal antibody. Whereas ATP supported daunomycin transport, nonhydrolyzable ATP analogues were ineffective. ATP-dependent daunomycin transport by brush border vesicles was unidirectional (inside to outside) and temperature dependent and was blocked by Gp170 inhibitors but not by taurocholate or bromsulphalein glutathione. Studies using everted small intestine revealed transport of rhodamine 123, a Gp170 substrate, from the serosal surface through the mucosa and inhibition by Gp170 inhibitors. These results suggest that Gp170 in rat small intestinal brush border membrane vesicles is an ATP-dependent efflux pump responsible for the transport of Gp170 substrates into the small intestinal lumen. Gp170 may protect against exogenously derived potentially damaging hydrophobic cations and contribute to the rarity of small intestinal cancer in humans and many animals.     

Characterization of calcium uptake by brush border membrane vesicles of human small intestine

Calcium uptake was characterized in human duodenal, jejunal, and ileal brush border membrane vesicles. Calcium uptake into human intestinal brush border membrane vesicles represented uptake into intravesicular space as evidenced by studies of osmolality, temperature dependence, calcium ionophore A23187-induced efflux and influx, and lanthanum displacement. Calcium uptake into membrane vesicles was sodium-independent. Negative membrane potential induced by valinomycin and anion substitution studies indicated an electroneutral process. Initial rate of uptake of calcium was linear up to 30 s (Y = 0.11 + 0.02x, r = 0.99). Kinetic parameters were determined from uptake measurements at 7 s, well within the linear phase of uptake. Calcium uptake represented mediated and nonmediated components. These components showed changes along the intestinal tract. Km values of the mediated component increased aborally, being lowest in the duodenum and highest in the terminal ileum. Vmax was highest in the duodenum, followed by, in descending order, the ileum, terminal ileum, and jejunum. The nonmediated component was greatest in the duodenum and decreased aborally. The duodenum appears to have a high-affinity, high-capacity system for the transport of calcium in humans. These studies are the first to characterize calcium transport by brush border membranes of the human small intestine.     

Effects of urogastrone-epidermal growth factor on intestinal brush border enzymes and mitotic activity.

The wet weight of the stomach, small intestine, caecum, and colon were significantly reduced (p less than 0.001) in intravenously fed rats compared with orally fed controls. Human epidermal growth factor (urogastrone) reversed this atrophy. Detailed analysis of the small intestine showed a similar effect on intestinal crypt cell population, mitoses per crypt, and protein content. Brush border gamma glutamyltransferase and alpha glucosidase activities were reduced by up to 50% throughout the small intestine of the animals fed intravenously. The specific activities (mU/mg protein) were unchanged, as a concomitant decrease in the tissue weight and protein content also occurred. Intestinal brush border enzyme activities in the rats treated with urogastrone-epidermal growth factor were restored to those seen in the orally fed rats except for alpha glucosidase activity in the proximal gut. In addition, the specific activity of gamma glutamyltransferase was highly significantly increased (p less than 0.01) in all regions of the small intestine. Thus, although urogastrone administration prevents the decrease in brush border enzyme activity seen after the removal of luminal nutrition, the response seems to differ depending on the intestinal location, with the specific activities of some enzymes being higher than those seen in orally fed rats. Urogastrone-epidermal growth factor can thus significantly increase the functional ability of the intestine in addition to its trophic effects.     

Diarrhoea and malabsorption in acquired immune deficiency syndrome: a study of four cases with special emphasis on opportunistic protozoan infestations.

Chronic diarrhoea is frequent in acquired immune deficiency syndrome (AIDS) but has been poorly investigated so far. We report four patients with AIDS in whom diarrhoea and malabsorption were outstanding features, and who underwent extensive digestive investigations. Diarrhoea was a presenting symptom in all subjects and was of secretory type in three of them. D-xylose and vitamin B12 were malabsorbed in all cases; steatorrhea was found in the two patients who could ingest significant amounts of fat. Faecal alpha 1-antitrypsin clearance was increased in all subjects. Search for digestive pathogens showed unusual protozoans in all patients: in case 1, optical and electron microscopy revealed the presence in the cytoplasm of villous enterocytes of Microsporidia protozoans still unreported in AIDS. Stool and jejunal fluid examination showed Isospora belli in case 2 and Cryptosporidium in cases 3 and 4. On histological and ultrastructural study the former was localised in the cytoplasm of a few enterocytes and the latter was scattered throughout the villus and crypt brush border. Otherwise small intestinal histology only showed minor non-specific changes and the enterocytes were ultrastructurally normal. In patient 3 the slow marker intestinal perfusion technique showed a profuse fluid secretion in the duodenum and proximal jejunum. All patients needed prolonged total parenteral nutrition. Cryptosporidium and Microsporidia could not be eradicated despite multiple drug trials. Isospora belli was transiently cured by pyrimethamine-sulphadiazine. Only patient 2 is presently at home, and patients 1, 3, and 4 died after two, six, and nine months of total parenteral nutrition, respectively.     

Defective jejunal brush border membrane sodium/proton exchange in association with lethal familial protracted diarrhoea.

The spectrum of clinical disease associated with specific defects in jejunal brush border membrane sodium/proton exchange is poorly defined and only two patients have been described so far. Jejunal brush border membrane transport studies were performed in a boy who presented with lethal familial protracted diarrhoea in the first few days of life. Using jejunal brush border membrane vesicles prepared from conventional jejunal biopsy specimens, initial sodium uptake under H+ gradient conditions was found to be only 6% of the mean control value. In contrast, sodium stimulated glucose uptake was normal. Our data confirm the importance of a congenital defect in this exchanger as a cause of severe sodium-losing diarrhoea and extend the spectrum of disorders characterised by a specific defect in brush border membrane Na+/H+ exchange to include some forms of lethal familial protracted diarrhoea.     

The Influence of Age on Intestinal Dipeptidyl Peptidase IV (DPP IV/CD26), Disaccharidases,and Alkaline Phosphatase Enzyme Activity in C57BL/6 Mice

The objective of this study was to determine and describe the age-related changes in intestinal brush border membrane enzyme activities that occur in C57Bl/6 mice. Specifically, jejunal, duodenal, and ileal dipeptidyl peptidase IV/CD26, disaccharidase (lactase, sucrase, and maltase), and alkaline phosphatase activities were determined. A significant correlation between analyzed intestinal brush border membrane enzyme activities and animal age was found. Our study revealed that intestinal dipeptidyl peptidase IV/CD26, lactase, sucrase, maltase, and alkaline phosphatase activities decline significantly with age (p < .05). Nevertheless, the horizontal (duodenum to ileum) enzyme activity patterns are not affected by age.     

Effects of non-steroidal anti-inflammatory drugs on the methotrexate transport in rat small intestine

BACKGROUND: Methotrexate (MTX) is used in the treatment of rheumatic disease, sometimes along with non-steroidal anti-inflammatory drugs (NSAIDs), and is actively co-transported with H+ in the small intestine, mediated by a reduced folate carrier (RFC). The co-administration of NSAIDs with MTX might cause a decrease in MTX absorption through the small intestine, since some NSAIDs are uncouplers of oxidative phosphorylation. The present study investigates whether flufenamic acid, diclofenac and indomethacin, NSAIDs, decrease the ATP content of small intestinal epithelial cells and affect MTX transport (the secondary active transport) in the small intestine. METHODS: MTX transport was examined in the presence and absence of the NSAIDs, using the everted intestine technique and brush border membrane vesicles (BBMVs) from the rat small intestine. The change in physical properties of the membrane was studied in BBMVs using the fluorescence techniques. RESULTS: MTX absorption in the small intestine with H+ gradient (mucosal side, pH 6.0; serosal side, pH 7.4) decreased in the presence of the NSAIDs, but absorption without H+ gradient (both sides, pH 7.4) was unaffected. The intestinal mucosal ATP content decreased in the presence of the NSAIDs. The uptake of MTX in BBMVs was unaffected by the NSAIDs. The activity of intestinal Na+-K+-ATPase was enhanced in the presence of the NSAIDs. The fluorescence measurements showed that membrane fluidity, membrane potential and membrane hydrophobicity of BBMVs were unaffected by the NSAIDs. CONCLUSIONS: NSAIDs decreased the H+/MTX absorption in the small intestine, but not the passive transport. The uncoupling effect of the NSAIDs decreased the ATP content in the small intestine, resulting in inhibition of the secondary active transport.     

Cholecystokinin and secretin prevent the intestinal mucosal hypoplasia of total parenteral nutrition in the dog.

Because the pancreas undergoes involutional changes during total parenteral nutrition (TPN) and because pancreatico-biliary secretions are trophic to the intestine, we studied jejunal and ileal structure and function and exocrine pancreatic function before and after 6 weeks of TPN in two groups of beagle dogs, one of which had TPN alone, the other having TPN plus daily stimulation of pancreatico-biliary secretions with intravenous infusions of cholecystokinin (CCK) and secretin. The injections of 1 U each per kg of body weight per day of CCK and secretin completely prevented the proximal and distal small bowel mucosal hypoplasia which developed in the TPN alone group. They also resulted in significant increases in in vivo galactose absorption (64 mM) per unit length of jejunum and ileum. However, there was no significant change in mucosal alpha-glucosidase and catalase activity or in in vitro mucosal uptake of 1 mM [14C]leucine when expressed per unit weight of intestinal mucosa. The capacity of the pancreas to respond to CCK and secretin was unaffected by excluding food from the intestine with 6 weeks of TPN in terms of pH, volume, and peak secretion rates of bicarbonate and protein, but maximum amylase output (units per 15 min per kg of body weight) fell significantly (P less than 0.05) from a mean of 1022 +/- 155 to 874 +/- 426 in TPN alone group and to 472 +/- 79 in the TPN dogs given CCK and secretin. These results show that daily CCK and secretin is trophic to the intestine of dogs nourished by TPN but do not indicate whether this trophic effect is attributable to CCK alone, secretin alone, the combination of the two hormones, or to the resultant stimulation of pancreatico-biliary secretions.     

Phosphate Transport Adaptation in Rat Jejunum and Plasma Level of 1,25-Dihydroxyvitamin D3

Intestinal absorption of inorganic phosphate (Pi) increases in response to a reduction in the dietary supply of Pi. In this work this adaptive response has been characterized in jejunal brush border membrane vesicles and studied in temporal relationship with the change in the plasma level of 1,25(OH)2D₃. The results indicate that in rat jejunal brush border membrane vesicles the activity of the sodium-dependent Pi transport system is stimulated by a low Pi diet. This adaptive response was the result of an increase in the V_max and a reduction in the K_m of the cotransport system. This change in Pi transport was correlated with an increase in the circulating level of 1,25(OH)2D₃ in a time-related fashion. In conclusion, these results are consistent with the notion that Pi restriction leads to an increase in Pi transport activity in the luminal membrane of the intestine. A time course study suggests that the elevation in plasma 1,25(OH)2D₃ might be involved in the adaptation of the intestinal Pi transport system to Pi restriction.     

Reduced intestinal epithelial cell brush border membrane calcium transport in spontaneously hypertensive rats.

OBJECTIVE: The present study aimed to determine whether there were alterations in intestinal calcium homeostasis in the spontaneously hypertensive rats (SHR) and to identify at which interface of the intestinal epithelial cell (brush border or basolateral) this occurs. DESIGN: Controversy exists as to whether intestinal calcium transport is altered in association with hypertension. Studies using perfused duodenal segments of the SHR have shed little light on the problem; other studies have only measured calcium transport in brush border membrane vesicles. This study allows specific focus on calcium transport mechanisms at both the brush border and basolateral membrane using simultaneously prepared membrane vesicles. METHODS: Calcium transport was studied by measuring radiolabelled calcium (45Ca) uptake in isolated brush border and basolateral membrane vesicles, prepared from the small intestines of SHR and Wistar-Kyoto (WKY) rats. Calcium uptake was measured when vesicles were incubated in solutions containing different concentrations of ATP and calcium. Orientation and membrane marker assays were used to confirm the phenotypes of the two membrane vesicle preparations. RESULTS: ATP-dependent calcium efflux was only observed in the basolateral membrane, which contains the Ca2+ -ATPase pump. SHR brush border membrane vesicles displayed no significant increase in calcium incorporation, whereas WKY brush border vesicles showed a 500% increase in uptake (ANOVA, P<0.05, n = 7). CONCLUSIONS: This study indicates that deficiencies exist in SHR intestinal calcium transport at the brush border membrane of intestinal epithelial cells. While further studies are required to ascertain the exact mechanisms involved, postulated deficiencies in the actions of calcium regulating hormones at this membrane suggest the need for concurrent intake of a calcitrophic agent to assist calcium uptake at the brush border membrane.     

Beneficial effects of insulin-like growth factor I on epithelial structure and function in parenterally fed rat jejunum

BACKGROUND & AIMS: The functional significance of intestinal hyperplasia stimulated by insulin-like growth factor (IGF)-I is unclear and has not been studied in a model of mucosal atrophy induced by total parenteral nutrition (TPN). The aim of this study was to determine how IGF-I affects intestinal structure and epithelial function in the absence of luminal nutrition caused by TPN. METHODS: Rats were maintained with TPN with or without IGF-I (800 micrograms/day), and jejunal histology and epithelial ion transport were measured after 5 days. In a third TPN group without IGF-I, a short-term dose of IGF-I was added during in vitro flux chamber experiments. RESULTS: Rats given TPN with IGF-I had greater jejunal mucosal weight, greater protein and DNA content, and increased villus height and crypt depth compared with rats given TPN only. TPN increased ionic permeability and ion transport responses to secretory and absorptive agents. IGF-I in vivo reversed most of these changes; IGF-I in vitro enhanced sodium-dependent glucose absorption but had no other effects. CONCLUSIONS: Coinfusion of recombinant human IGF-I with TPN solution stimulates intestinal hyperplasia and attenuates transport changes induced by TPN. The latter effect seems to be primarily associated with the growth state of the epithelium. (Gastroenterology 1996 Dec;111(6):1501-8)