Biomedical subjects
M D Sitrin
Publications and source records attributed to M D Sitrin.
Correction of abnormal small intestinal cytosolic protein kinase C activity in streptozotocin-induced diabetes by insulin therapy.
Diabetes was induced in rats by administration of a single intraperitoneal injection of streptozotocin (50 mg/kg body wt). After 7 days, one group of diabetic animals was treated with insulin for an additional 5 days. Control, diabetic and diabetic + insulin rats were then killed, their distal small intestines were removed and the epithelial cells were examined and compared with respect to polyphosphoinositide turnover, total protein kinase C activity and cellular distribution, and 1,2-diacylglycerol mass and production. The results of these experiments demonstrated that, compared with their control counterparts, the intestines from diabetic rats had a decreased turnover of polyphosphoinositides, but an increase in 1,2-diacylglycerol mass which was a result, at least in part, of an increase in the synthesis of this lipid de novo. Total protein kinase C activity was decreased in the diabetic rats due to a decrease in cytosolic activity, with no significant change in particulate activity. Moreover, insulin administration for 5 days to diabetic animals did not affect their lowered intestinal polyphosphoinositide turnover, but did further accentuate their increased 1,2-diacylglycerol mass and synthesis de novo; this treatment also corrected total protein kinase C activity by increasing the cytosolic activity of this enzyme. These results indicate that signalling mechanisms involving polyphosphoinositides, 1,2-diacylglycerol and protein kinase C are abnormal in the intestines of diabetic rats and that some of these biochemical parameters can be modulated by insulin administration in vivo.
1,2-Dimethylhydrazine-induced alterations in protein kinase C activity in the rat preneoplastic colon.
Recently, a number of studies in experimental animals and humans have suggested that alterations in the activity of protein kinase C (PKC) may be involved in the malignant transformation process. To determine whether such alterations in this kinase were present before the development of 1,2-dimethylhydrazine (DMH)-induced colon cancers, rats were given s.c. injections of this procarcinogen (20 mg/kg body weight/week) or diluent for 10 or 15 weeks. Animals were sacrificed after these time periods and colonic epithelium was harvested from each group. The activity and distribution of PKC in the cytosolic and membrane fractions of these preparations as well as 1,2-diacylglycerol mass and phosphoinositide turnover were then examined and compared in the presence and absence of 10 nM 1,25-dihydroxycholecalciferol, an agent which has previously been found to influence these biochemical parameters in the normal rat colonic epithelium. The results of these studies demonstrate that: (a) the percentage of PKC activity in the membrane fraction was significantly greater in DMH-treated animals compared to their control counterparts at 10 and 15 weeks; (b) the total PKC activity was similar at 10 weeks, but markedly reduced in the colonic mucosa of the DMH-treated group at 15 weeks; (c) 1,2-diacylglycerol mass and phosphoinositide turnover were increased in the colonic mucosa of rats administered this carcinogen at both time points; and (d) in control, but not in DMH-treated animals, in vitro addition of 1,25-dihydroxycholecalciferol increased PKC activity, 1,2-diacylglycerol mass and phosphoinositide turnover at each of the times studied. Based on these findings, it would appear that alterations in PKC activity may play a role in the malignant transformation process of the colon in animals administered DMH.
Effect of chronic selenite supplementation on selenium excretion and organ accumulation in rats.
We examined the effect of chronic selenite supplementation on whole body and selected organ selenium (Se) accumulation, urine excretion of total Se and trimethylselenonium ion, and Se balance in adult male rats. Animals were housed in metabolic cages and given either deionized water or water containing 4 micrograms of Se/mL as selenite for 30 d. Absorption of selenite was nearly complete, with only approximately 10% of ingested Se appearing in feces. There was a rapid rise in urinary Se that reached a plateau within a few days and accounted for 54 +/- 2% of the intake. Excretion of trimethylselenonium ion (TMSe) in urine increased rapidly, representing 35-40% of urinary Se in the supplemented animals compared with only 2% for the control group. In one experiment, rats were killed at 30 d and total carcass Se was measured using isotope dilution analysis. Supplemented rats had only a modest increase in whole body Se (94 +/- 4 micrograms Se vs. 66 +/- 3 in controls). Calculation of Se balance in the supplemented rats showed that approximately 35% of ingested Se could not be accounted for by urine plus fecal losses combined with the portion retained in the carcass. The results from this study demonstrate that under the condition of supplementation at 4 micrograms of Se/mL of drinking water, pathways other than urinary and fecal excretion may account for a substantial portion of Se loss.
Intestinal malabsorption syndromes.
This chapter is a review of selected causes of intestinal malabsorption. The pathogenesis and clinical features of malabsorption due to acquired immunodeficiency syndrome (AIDS), celiac sprue, and small bowel bacterial overgrowth are discussed, with emphasis on recent developments. The etiology and diagnostic challenges of malabsorptive disorders in the elderly are also reviewed.
1,25(OH)2 vitamin D3 stimulates membrane phosphoinositide turnover, activates protein kinase C, and increases cytosolic calcium in rat colonic epithelium.
The hormonal form of vitamin D, 1,25(OH)2 vitamin D3 [1,25(OH)2D3], regulates colonic calcium absorption and colonocyte proliferation and differentiation. In this study, we have examined the effect of 1,25(OH)2D3 on membrane phosphoinositide turnover, protein kinase C activation, and regulation of intracellular calcium concentration [( Ca+2]i) in isolated rat colonic epithelium. In a concentration-dependent manner, 1,25(OH)2D3 stimulated breakdown of membrane phosphoinositides within 15 s, generating diacylglycerol and inositol 1,4,5-triphosphate (IP3). 1,25(OH)2D3 rapidly activated colonic protein kinase C, with maximal translocation of activity from the cytosol to the membrane occurring within 1 min of exposure to the secosteroid. Studies performed in isolated colonocytes with the fluorescent dye fura-2 demonstrated that 10(-8) M 1,25(OH)2D3 caused a rapid rise in [Ca+2]i which then transiently decreased before rising to a new plateau value. When these experiments were performed in a calcium-free buffer, an increase in [Ca+2]i was observed, but both the transient and secondary rise were diminished in magnitude, suggesting that 1,25(OH)2D3 may stimulate both release of intracellular calcium stores and calcium influx. 1,25(OH)2D3 stimulated [3H]thymidine uptake in rat colonocytes, 4 h after an in vivo injection. These studies indicate that 1,25(OH)2D3 exerts a rapid influence on membrane phosphoinositide metabolism which may mediate certain of the secosteroid's effects on colonocyte calcium transport and proliferation.
Serum unconjugated bile acids in patients with small bowel bacterial overgrowth.
Concentrations of total and unconjugated bile acids in serum were measured fasting and 2 h postprandially in 9 patients with a positive [14C]glycocholate breath test consistent with small bowel bacterial overgrowth and in 13 controls. Gas-liquid chromatography-mass spectrometry (GLC-MS) and enzymatic-fluorometric assays were both used. In contrast to previous work, total serum bile acids were only occasionally elevated in patients with bacterial overgrowth. Total 2 h postprandial unconjugated bile acids, however, were elevated in 7/9 patients when measured by GLC-MS and in 6/9 when measured by the enzymatic-fluorometric method. The best separation between patients and controls was achieved by GLC-MS determinations of 2 h postprandial unconjugated cholic acid or primary bile acids, which were abnormal in 8/9 patients. This study indicates that measurement of serum bile acids may be a useful approach to the diagnosis of bacterial overgrowth, but would require accessible methods for separating and measuring cholic acid or unconjugated primary bile acids in post-prandial sera.
Dietary triacylglycerol modulates sodium-dependent D-glucose transport, fluidity and fatty acid composition of rat small intestinal brush-border membrane.
Rats were maintained on nutritionally complete diets enriched in unsaturated (menhaden fish oil) or saturated (butter fat) triacylglycerols. After 4 weeks, the animals were killed, proximal small intestinal brush-border membranes were prepared, and examined and compared with respect to their lipid composition, molecular species of phosphatidylcholine, lipid fluidity and sodium-dependent D-glucose transport. Membranes prepared from the two dietary groups were found to possess similar ratios of cholesterol/phospholipid (mol/mol), sphingomyelin/phosphatidylcholine (mol/mol), and protein/lipid (w/w). In contrast to these findings, however, striking differences were noted in the total fatty acid compositions of these membranes. Plasma membranes prepared from animals fed the fish oil diet possessed higher percentages of saturated fatty acids as well as (n - 3) unsaturated fatty acids and lower percentages of monounsaturated and (n - 6) unsaturated fatty acids than those prepared from animals fed the butter fat diet. Analysis of the molecular species of phosphatidylcholine by HPLC, moreover, revealed that membranes from rats fed fish oil had higher levels of 16:0-20:5, 16:0-22:6 and 18:0-20:5 and lower levels of 18:0-18:2 and 16:0-18:1 than their butter fat counterparts. As assessed by steady-state fluorescence polarization, differential polarized phase fluorometric and excimer/monomer fluorescence intensity techniques using various fluorophores, the lipid fluidity of membranes from rats fed fish oil was also found to be significantly lower compared to membranes from rats fed butter fat. Finally, comparison of the kinetic parameters of Na+-dependent D-glucose transport revealed that fish oil-membrane vesicles had a higher maximum velocity (Vmax) than butter fat membrane vesicles but a similar Km for glucose.
Effect of phosphorus on endogenous calcium losses during total parenteral nutrition.
Intravenous phosphorus can reduce urinary calcium losses in patients receiving total parenteral nutrition (TPN). We investigated the effect of intravenous P on urinary and fecal Ca loss in intravenously fed normal and thyroparathyroidectomized (TPTX) rats to assess the role of parathyroid hormone (PTH) and endogenous fecal Ca losses. Doubling the intravenous P load during TPN decreased urinary Ca losses by 54% (0.299 vs 0.137 mmol/d) in intact rats and by 43% (0.514 vs 0.294 mmol/d) in TPTX rats. Increased P load in normal rats had no effect on urinary cyclic AMP excretion, serum Ca, serum P, serum 1,25-dihydroxycholecalciferol, or endogenous fecal Ca losses. These observations suggest that the hypocalciuric effect of P during TPN is independent of PTH and is not caused by a repartitioning of obligatory Ca losses from the renal to the intestinal route.
Acetate and hypercalciuria during total parenteral nutrition.
Hypercalciuria and negative calcium balance are complications of total parenteral nutrition (TPN). Because metabolism of the TPN formula generates an acid load that can induce hypercalciuria, we evaluated the effect of supplementing the formula with acetate. In a randomized crossover study six patients on continuous and six on cyclic TPN received no added acetate or 160 mmol acetate/d replacing 160 mmol chloride/d for 3 d each. Blood and urine measurements were obtained on day 3 of each formula. Acetate, which is metabolized to bicarbonate, increased blood pH and decreased renal acid excretion. Urinary Ca decreased in every patient from 422 +/- 63 to 240 +/- 46 mg/d (10.5 +/- 1.6 to 6.0 +/- 1.4 mmol/d) and from 468 +/- 68 to 285 +/- 54 mg/d (11.7 +/- 1.7 to 7.1 +/- 1.3 mmol/d) during continuous and cyclic TPN, respectively. Filtered Ca load decreased slightly whereas renal tubular Ca reabsorption increased significantly with acetate. Serum parathyroid hormone, 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D, and urinary cyclic AMP were not different.
Intestinal absorption of cholecalciferol and 25-hydroxycholecalciferol in chronic cholestatic liver disease.
We compared the absorption of cholecalciferol and 25-hydroxycholecalciferol in normal subjects and in patients with mild and severe cholestatic liver disease. 3H-cholecalciferol and 3H-25-hydroxycholecalciferol were given orally and serial blood samples were drawn for measurement of the serum level of radiolabeled vitamin. Absorption of 25-hydroxycholecalciferol peaked earlier and was greater than absorption of cholecalciferol at all times in all three groups. Patients with mild cholestasis (normal bilirubin and fecal fat excretion) absorbed both forms of the vitamin normally. Those with severe cholestasis (jaundice and steatorrhea) had minimal absorption of cholecalciferol but relatively preserved absorption of 25-hydroxycholecalciferol. Absorption of cholecalciferol and 25-hydroxycholecalciferol was inversely related to fecal fat excretion. The superior absorption of 25-hydroxycholecalciferol may partly explain its greater efficacy in oral treatment of vitamin D deficiency in patients with severe cholestasis.
Vitamin E deficiency and neurologic disease in adults with cystic fibrosis.
We report the cases of two adult patients with cystic fibrosis affecting the pancreas and liver, who also had severe vitamin E deficiency and neurologic disease. The most prominent clinical features were abnormal eye movements, diminished reflexes, decreased vibratory and position sense, ataxia, and muscle weakness. Treatment with intramuscular injections of vitamin E partially corrected the neurologic deficits. Vitamin E absorption tests documented severe malabsorption, which was later alleviated by the addition of dessicated ox bile to the regimen of alpha-tocopheryl acetate. These studies suggest that a decreased intraluminal concentration of bile salts is an important factor in the development of severe vitamin E deficiency and in the poor response to oral replacement therapy that is seen in some patients with cystic fibrosis.
Altered fractional excretion of uric acid during total parenteral nutrition.
The presence of crystal proven podagra coincident with a 52% decrease in plasma urate after a 3-day course of total parenteral nutrition (TPN) prompted a study of urate excretion in 9 patients with Crohn's disease. By Day 9 in those receiving TPN, plasma urate decreased 58% (p less than 0.001), while fractional urate excretion increased 94% (p less than 0.005). Twenty-four hour urate excretion and serum creatinine were not significantly altered. These findings persisted for the duration of TPN. In 2 patients with ileocolitis, the addition or deletion of either lipid emulsion or multivitamin infusions during TPN had no effect on urate values. Rather, the amino acid load or a specific constituent appears to be the causal factor. These data suggest that hypouricemia due to extensive net urate excretion is common during TPN therapy. Finally, patients with established gout may be at risk for acute gouty attacks during TPN therapy.
Metabolic acidosis. Development in two patients receiving a potassium-sparing diuretic and total parenteral nutrition.
Two patients developed a metabolic acidosis associated with a normal undetermined anion concentration while receiving a potassium-sparing diuretic and total parenteral nutrition. In both cases the metabolic acidosis resolved within one week after discontinuing administration of the diuretic. The use of potassium-sparing diuretics in a patient receiving total parenteral nutrition requires caution and continued monitoring for this potential drug-nutrient interaction.
Calciuretic effect of cyclic versus continuous total parenteral nutrition.
Metabolic bone disease has been reported in patients receiving long-term cyclic administration of total parenteral nutrition (TPN). The exact etiology of this disturbance in mineral homeostasis has not been identified, however many of these patients are markedly hypercalciuric and in negative calcium balance. We have studied the effects of cyclical versus continuous infusion of nutrients on urinary calcium losses in a group of patients beginning a program of long-term home TPN. Cyclic TPN, when administered over either 18 or 12 hours, significantly increased daily urinary calcium excretion compared to continuous 24-h TPN infusion by 19 and 28%, respectively. During cyclic TPN, frank negative calcium balance was observed in 3 of 5 patients studied compared to 2 of 5 patients during continuous TPN. The pattern of urinary calcium loss during cyclic TPN was such that approximately 80% of the daily urinary calcium losses occurred during the 12 hours of TPN infusion. Cyclic administration of TPN increased the urinary calcium losses in all patients suggesting that an intermittent TPN infusion schedule, as typically utilized in home TPN programs, increases the risk of developing negative calcium balance, at least during the early phase of cyclic TPN administration.
Nutritional aspects of inflammatory bowel disease.
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Intestinal absorption of 1,25-dihydroxyvitamin D3 in the rat.
Intestinal absorption of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] from in vivo jejunal sacs was studied in rats with thoracic duct and bile duct cannulas. In 6 h 86.5% of the administered 1,25(OH)2D3 was absorbed, but only 7.3% was recovered in thoracic duct lymph. Appearance in plasma of [3H]-1,25(OH)2D3 after intrajejunal administration was identical in rats with and without diverting lymph cannulas, indicating that the 1,25(OH)2D3 is absorbed almost entirely via portal blood. When 1,25(OH)2D3 was instilled into the jejunum in a fat suspension without bile salts rather than a mixed micellar solution, less was recovered in lymph, but total intestinal absorption was unchanged. High-pressure liquid chromatography of a lymph extract demonstrated that 1,25(OH)2D3 was present only as the unchanged secosteroid. In lymph, only 4.1% of the 1,25(OH)2D3 was in the chylomicrons, with the remainder bound to the plasma protein fraction of lymph. Treatment of rats with cycloheximide to block chylomicron synthesis did not decrease absorption of 1,25(OH)2D3. These results indicate that intestinal absorption of 1,25(OH)2D3 is effective and not very dependent on luminal bile salts. Almost all 1,25(OH)2D3 is released from the intestine directly into portal blood and does not require packaging in chylomicrons for transport into intestine lymph.
Calciuria in total parenteral nutrition: effects of amino acids and glucose in rats.
Total parenteral nutrition (TPN) is known to cause hypercalciuria and negative calcium balance in some patients. We have now shown that the administration of TPN to rats causes marked increases in urinary calcium losses. Moreover, urinary calcium excretion in the TPN rat responds to changes in the infusate concentration of calcium and amino acids similarly to what has been observed in TPN patients. For any given increase in the amount of calcium infused 130% more calcium was excreted in the urine by rats receiving TPN compared to rats receiving saline alone. At a fixed level of calcium infused, urinary calcium increased linearly when the amino acid content of the infusate was increased from 0 to 2.75 to 4.25%. However, a reduction in the glucose load, via isocaloric substitution with lipid by 60%, had no effect on urinary calcium excretion. The TPN rat appears to be a promising animal model in which to investigate the hypercalciuretic effect of intravenous nutrition, particularly as calcium homeostasis may be affected by various nutritional components of the TPN solution.