Protein turnover in intestinal mucosal villus and crypt brush border membranes

Following a concomitant oral dose of salicylamide and acetaminophen (5 mg/kg of each) the urinary excretion of glucuronide and sulfate conjugates of the drugs were followed in children (ages seven to ten years) and adults. No significant difference were observed between the two age groups in the half-lives for appearance of salicylamide conjugates in urine. Age-related changes in the metabolic pathways, however, were observed. The mean percentage of dose excreted as salicylamide sulfate was significantly higher in children (78%) than in adults (36%). In contrast, salicylamide glucuronide was the major excretory product in adults. Similar age-related differences were observed for acetaminophen conjugation. Pharmacokinetic analysis indicated that the deficiency in glucuronide conjugation of these drugs in children is accompanied by a higher rate of sulfate formation.     

pH-dependent fluoride transport in intestinal brush border membrane vesicles

Ciliary neurotrophic factor (CNTF) exerts a multiplicity of effects on a broad spectrum of target cells, including retinal neurons. To investigate how this functional complexity relates to the regulation of CNTF receptor alpha (CNTFR alpha) expression, we have studied the developmental expression of the receptor protein in chick retina by using immunocytochemistry. During the course of development, the receptor is expressed in all retinal layers, but three levels of specificity can be observed. First, the expression is regulated temporally with immunoreactivity observed in ganglion cells (embryonic day 8 [E8] to adult), photoreceptor precursors (E8-E12), amacrine cells (E10 to adult), bipolar cells (E12-E18), differentiated rods (E18 to adult), and horizontal cells (adult). Second, expression is restricted to distinct subpopulations of principal retinal neurons: preferentially, large ganglion cells; subpopulations of amacrine cells, including a particular type of cholinergic neuron; a distinctly located type of bipolar cell; and rod photoreceptors. Third, expression exhibits subcellular restriction: it is confined largely to dendrites in mature amacrine cells and is restricted entirely to outer segments in mature rods. These data correlate with CNTF effects on the survival of ganglion cells and mature photoreceptors, the in vitro differentiation of photoreceptor precursors and cholinergic amacrine cells, and the number of bipolar cells in culture described here or in previous studies. Thus, our results demonstrate an exceptional degree of complexity with respect to the regulation of neuronal CNTFR alpha expression in a defined model system. This suggests that the same signaling pathway is used to mediate a variety of regulatory influences, depending on the developmental stage and cell type.     

Short-chain fatty acid-supplemented total parenteral nutrition improves nonspecific immunity after intestinal resection in rats

BACKGROUND: Total parenteral nutrition (TPN) alters both specific and nonspecific immune functions, resulting in immunosuppression. Short-chain fatty acids have been shown to improve the adaptive responses of the gut after surgery. The following study investigates the effects of adding short-chain fatty acids to TPN on the immune system after an 80% small bowel resection. METHODS: Rats (237 +/- 3 g) were infused with either TPN (n = 25) or TPN supplemented with short-chain fatty acids (n = 26) for 3 or 7 days. Hematologic analysis was performed on peripheral blood and splenocytes were isolated to characterize cell phenotypes, natural killer cell cytotoxicity and to estimate proliferative response. RESULTS: The relative percent of T (CD3+) cells increased (p < .05) and the relative percent of macrophages decreased (p < .001, n = 13) in the spleens of the 3-day TPN-fed rats. By day 7, these differences disappeared. The natural killer cells from rats that were supplemented with short-chain fatty acids had higher (p < .0001) cytotoxic activity than the TPN groups at day 3. Mitogenic response did not differ between groups but were depressed compared with sham-treated rats. By day 7, rats on standard TPN had larger (p < .0001) spleens than all other groups. This group also had a higher total white blood cell count because of increased numbers of macrophages and neutrophils (p < .02). CONCLUSION: Short-chain fatty acids improve components of nonspecific immune responses and may be beneficial in reducing certain aspects of TPN-associated immunosuppression after major surgery.     

Copper metabolism and requirements in total parenteral nutrition

Copper metabolism and requirements in patients receiving total parenteral nutrition were studied in 28 patients with gastrointestinal diseases. During each of the 3 wk of the study period, each of 24 patients received in their total parenteral nutrition solutions, a daily dose of copper amounting to 0.25 mg, 1.05 mg, or 1.85 mg, in a random order. The other 4 patients received a fixed daily dose of 1 mg throughout the 3 wk. Increased losses of copper through the gastrointestinal tract occurred in patients with diarrhea or high-output stomas or fistulas. Patients with abnormalities of liver excretory functions had decreases in gastrointestinal copper losses. Urinary copper excretion was twice that of normal subjects. Copper infused in excess of the requirements was retained and not excreted. Plasma copper did not reflect the copper balance and cannot be used as a guide for copper supplementation. Copper requirements were found to be 0.3 mg/day in patients with normal amounts of gastrointestinal excretion. In the presence of diarrhea or increased fluid loss through gastrointestinal stomas or fistulas, the copper requirements for total parenteral nutrition are 0.4--0.5 mg/day.     

Hepatobiliary toxicity of total parenteral nutrition in adults

The enzymology and clinical manifestations of total parenteral nutrition (TPN)-induced liver abnormalities have been investigated extensively. The cause, pathogenesis, and treatment of TPN-related hepatic and biliary dysfunction in adults still are not well understood, however. The findings of experimental studies in animals has not necessarily correlated with the human data, and there have been few prospective, randomized controlled trials examining the mechanism, cause, or treatment of TPN-induced hepatobiliary toxicity in adults. This article examines the animal models of pathogenesis and treatment of TPN-induced intrahepatic and extrahepatic abnormalities, and provides a discussion of abnormalities seen in humans.     

Efficacy and safety of total parenteral nutrition in pediatric patients

Pediatric patients differ from adult patients because of active musculoskeletal growth and development of visceral organs and because they have a proportionately smaller nutritional reserve, especially premature infants. Measures of outcome of effective nutritional support in pediatric patients who have experienced trauma or medical disease or who have undergone surgical procedures include weight gain, increased height and circumference of the head, increased hepatic synthesis of plasma proteins, immunocompetence, decreased morbidity, improved survival, and fast recovery. If a pediatric patient cannot eat or be tube-fed enterally after 3 days of recovery and support with fluids, parenteral nutrition is indicated. Examples in which this treatment has dramatically decreased morbidity include gastroschisis, short-bowel syndrome, necrotizing enterocolitis, and Hirschsprung's disease. Contraindications to its use include severe congenital (usually genetic) defects and terminal cancer, conditions in which life expectancy and quality of life are severely decreased. The team approach to parenteral and enteral nutrition in pediatric patients is preferred, and stable patients receiving long-term nutritional support, including infants, should be considered for home parenteral nutrition. When administered by protocol, parenteral nutrition is safe in pediatric patients. In properly selected pediatric patients, direct and indirect costs for such therapy may be significantly less than those in adults, and the cost-to-benefit ratio is appreciably higher when life expectancy, parental pleasure, and potential work productivity are considered. Ethical and social issues in initiating and discontinuing parenteral nutrition are best decided during thorough empathic discussions between physicians and parents.     

Glucose dynamics during continuous hemodiafiltration and total parenteral nutrition

OBJECTIVE: To determine glucose balance during dextrose-free continuous hemodiafiltration with or without dextrose-containing ultrafiltrate replacement fluid and full nutritional support. DESIGN: Prospective, nonrandomized, observational study. SETTING: A 24-bed multiple trauma critical care unit in a level-I trauma center. PATIENTS: Seventeen multiple trauma patients with multiple organ dysfunction syndrome requiring hemodialysis for acute renal failure. INTERVENTIONS: Continuous hemodiafiltration effluent volume and glucose concentration were measured. Study days were classified according to whether dextrose was used in the ultrafiltrate replacement therapy. Use of dextrose in replacement therapy was determined clinically. Parenteral nutrition was not altered for potential glucose absorption from continuous hemodiafiltration. Ultrafiltrate replacement consisted of 5% dextrose in saline on 21 study days (D5YES) and dextrose-free solutions on 54 study days (D5NO). RESULTS: The D5YES group received 316 +/- 145 g glucose/day from the ultrafiltrate replacement fluid, in addition to glucose in total parenteral nutrition (total glucose intake = 942 +/- 229 g/day in D5YES, 682 +/- 154 g/day in D5NO) (p < 0.05). Glucose loss in continuous hemodiafiltration effluent was 82 +/- 61 g/day in D5YES and 57 +/- 22 g/day in D5NO (P < 0.05), for a net glucose uptake of 8.1 +/- 2.1 mg/kg per min in D5YES and 5.4 +/- 1.5 mg/kg per min in D5NO (p < 0.05). Glucose loss was predictable when dialysate and ultrafiltrate replacement fluids were dextrose-free (R2 = 0.77), but less so when dextrose was used as ultrafiltrate replacement (R2 = 0.47). CONCLUSION: Dextrose-free dialysate promotes glucose loss during continuous hemodiafiltration, but the loss is small and predictable. Use of a dextrose-containing ultrafiltrate replacement fluid results in a significant increase in glucose intake without a commensurate increase in glucose loss, and makes glucose loss in effluent less predictable.     

Effect of vitamin A on small intestinal brush border enzymes in a rat

To determine the utility of the myocardial tracer Tc-99m-tetrofosmin in the examination of patients with left bundle branch block (LBBB) and to investigate Tc-99m-tetrofosmin uptake and retention in the myocardium, early and delayed Tc-99m-tetrofosmin SPECT was performed in 10 patients having LBBB without coronary stenosis. METHODS: After 740 MBq of Tc-99m-tetrofosmin injection in the resting state, the early and delayed SPECT imaging was done at 30 min and 180 min, respectively. RESULTS: Decreased Tc-99m-tetrofosmin uptake in the septal segments was observed in 4 patients (40%) at 30 min and in 9 (90%) at 180 min. Reverse redistribution was seen in 9 of 10 patients. In patients with LBBB, the septal-to-lateral uptake ratio was lower in the delayed images than in the early images (0.80 +/- 0.09 vs. 0.89 +/- 0.09, p < 0.001). In patients with LBBB, the washout rate of Tc-99m-tetrofosmin was higher in the septal segments than in the lateral segments (28.3 +/- 4.3% vs. 22.8 +/- 3.3%, p < 0.001). CONCLUSION: The SPECT data indicate that in LBBB without coronary stenosis, the uptake of Tc-99m-tetrofosmin is decreased in the septal wall, and that reverse redistribution occurs frequently. Our results contribute to the elucidation of both the cellular biokinetics of Tc-99m-tetrofosmin in the myocardium and the hemodynamics of the septum in LBBB, and indicate the possible clinical utility of Tc-99m-tetrofosmin.     

Bolus intravenous infusion of amino acids or lipids does not stimulate gallbladder contraction in neonates on total parenteral nutrition

On the basolateral infoldings of the strial marginal cells in the cochlea, Na K ATPase activity is abundant. To clarify the humoral control by norepinephrine, K-NPPase activity of strial marginal cells in the cochlea was investigated in normal, reserpine, norepinephrine (NE), reserpine plus NE-treated guinea pigs using a cerium-based method. K-NPPase activity was almost completely decreased 3-20 days after reserpine administration. At 10 days after reserpinization and following NE repeated treatment, enzyme activity was detectable. These results suggested that norepinephrine might restore and regulate strial K-NPPase activity.     

Essential fatty acid deficiency and home total parenteral nutrition patients

The requirements for essential fatty acids in patients on home parenteral nutrition are not well described. We therefore studied the needs of 12 patients receiving parenteral nutrition for at least 4 mo (range: 4 mo-17.3 yr; mean 7.0 +/- 5.2 yr). Prior to the study, each patient had been receiving intravenous lipids either weekly or biweekly and had a triene to tetraene ratio (TTR) on plasma phospholipids performed at least annually. A TTR > or = 0.2 was considered diagnostic for essential fatty acid deficiency (EFAD). The purpose of this study was to determine the required intravenous lipid supplementation in patients on home total parenteral nutrition (HTPN). Patients with an initial TTR of < 0.2 had their intravenous lipid stopped and changes in their serum phospholipid fatty acids were followed every 3-4 wk. Nine of 12 patients had TTRs > 0.2 at some point in the study. Phase I consisted of patients who at initiation of the study had normal TTRs and were taken off lipid supplementation until their TTR became abnormal. Phases II, III, IV, and V consisted of lipid delivered in total nutrient admixtures in biweekly doses of 0.6, 1.2, 1.8, and 2.4 g of fat/kg bodyweight, respectively. Eight patients normalized their TTRs on the biweekly lipid regimens; one patient expired before his ratio normalized; and three patients could not be made deficient in essential fatty acids after 26 or more wk of fat-free parenteral nutrition. Most patients required 1.2 to 2.4 g of lipid/kg bodyweight/biweekly to correct serologic EFAD. The clinical background, as well as the length of small bowel remaining, did not seem to identify those patients who required lipid supplementation nor the final dose of lipid needed to normalize their TTRs.     

Refeeding varying fatty acid and cholesterol diets alters phospholipids in rat intestinal brush border membrane

The epidermis is an attractive site for therapeutic gene delivery because it is accessible and capable of delivering polypeptides to the systemic circulation. A number of difficulties, however, have emerged in attempts at cutaneous gene delivery, and central among these is an inability to sustain therapeutic gene production. We have examined two major potential contributing factors, viral vector stamina and involvement of long-lived epidermal progenitor cells. Human keratinocytes were either untreated or transduced with a retroviral vector for beta-galactosidase (beta-Gal) at > 99% efficiency and then grafted onto immunodeficient mice to regenerate human epidermis. Human epidermis was monitored in vivo after grafting for clinical and histologic appearance as well as for gene expression. Although integrated vector sequences persisted unchanged in engineered epidermis at 10 weeks post-grafting, retroviral long terminal repeat (LTR)-driven beta-Gal expression ceased in vivo after approximately 4 weeks. Endogenous cellular promoters, however, maintained consistently normal gene expression levels without evidence of time-dependent decline, as determined by immunostaining with species-specific antibodies for human involucrin, filaggrin, keratinocyte transglutaminase, keratin 10, type VII collagen, and Laminin 5 proteins out to week 14 post-grafting. Transduced human keratinocytes generated multilayer epidermis sustained through multiple epidermal turnover cycles; this epidermis demonstrated retention of a spatially appropriate pattern of basal and suprabasal epidermal marker gene expression. These results confirm previous findings suggesting that viral promoter-driven gene expression is not durable and demonstrate that keratinocytes passaged in vitro can regenerate and sustain normal epidermis for prolonged periods.     

Sea-blue histiocyte syndrome in bone marrow secondary to total parenteral nutrition

Clinical and hematological abnormalities can occur in patients receiving intravenous fat emulsions as part of a long-term parenteral nutrition; they consist of hepatosplenomegaly and peripheral blood cytopenia(s). These abnormalities lead to bone marrow examination which revealed numerous macrophages laden with blue staining pigment granules and separate lipid vacuoles, presenting the typical histochemical characteristics of sea-blue histiocytes. Thus, long-term parenteral nutrition including fat-emulsion sources may represent a further condition in addition to the wide variety of disorders which can be associated with sea-blue histiocytosis. Moreover, in view of its clinical and morphological presentation, this storage pathological state could be compared with the so-called sea-blue histiocyte syndrome described by Silverstein and colleagues.     

Glucose response to abrupt initiation and discontinuation of total parenteral nutrition

Plasma glucose was studied during the initiation of total parenteral nutrition (TPN) and the discontinuation of TPN without a tapering schedule. Blood was sampled every 5 minutes for 2 hours after the start of TPN and 1 week later as TPN was discontinued. A total of 14 initiations and 14 discontinuations were studied in 18 patients. Severity of illness in patients ranged from stable condition postoperatively to multiple-system failure; six patients had diabetes mellitus. The TPN solution was a 3:1 admixture that provided a caloric intake equal to 1.2 times the resting energy expenditure, with 40% fat and 60% carbohydrate calories. An average of 1963 kcal was provided per day (340 g of glucose, 79 g of fat). During the initiation phase, the mean increase in plasma glucose was 60 mg/dL. The increase for diabetic patients was 79 +/- 14 mg/dL compared with 52 +/- 23 mg/dL for the nondiabetics. During the discontinuation phase, the mean plasma glucose decreased 40 +/- 20 mg/dL; two patients with high concentrations of regular insulin (50 and 100 units) showed an increase in plasma glucose when the TPN was stopped. Plasma glucose returned to the preinfusion baseline after discontinuation. During both initiation and discontinuation, plasma glucose showed little change after the first 60 minutes. No clinical symptoms of hypoglycemia were observed. In conclusion, TPN as a 3:1 admixture can be safely started as full nutrition support and stopped abruptly without a tapering schedule. Plasma glucose response is rapid, predictable, and mostly complete within 60 minutes.     

Amplitude-dependent modulation of brush border enzymes and proliferation by cyclic strain in human intestinal Caco-2 monolayers

Little is known about the effects of repetitive deformation during peristaltic distension and contraction or repetitive villus shortening on the proliferation and differentiation of the intestinal epithelium. We sought to characterize the effects of repetitive deformation of a physiologically relevant magnitude and frequency on the proliferation and differentiation of human intestinal epithelial Caco-2 cells, a common cell culture model for intestinal epithelial biology. Human intestinal epithelial Caco-2 cells were cultured on collagen-coated membranes deformed by -20 kPa vacuum at 10 cycles/minute, producing an average 10% strain on the adherent cells. Proliferation was assessed by cell counting and 3H-thymidine incorporation. Alkaline phosphatase and dipeptidyl dipeptidase specific activity were measured in cell lysates. Since cells at the membrane periphery experience higher strain than cells in the center, the topography of brush border enzyme histochemical and immunohistochemical staining was analyzed for strain-dependence. Cyclic strain stimulated proliferation compared to static cells. Proliferation was highest in the membrane periphery where strain was maximal. Strain also modulated differentiation independently of its mitogenic effects, selectively stimulating dipeptidyl dipeptidase while inhibiting alkaline phosphatase. Strain-associated enzyme changes were also maximal in areas of greatest strain. The PKC inhibitors staurosporine and calphostin C ablated strain mitogenic effects while intracellular PKC activity was increased by strain. The strain-associated brush border enzyme changes were attenuated but not blocked by PKC inhibition. Thus, strain of a physiologically relevant frequency and magnitude promotes proliferation and modulates the differentiation of a well-differentiated human intestinal epithelial cell line in an amplitude-dependent fashion. PKC may be involved in coupling strain to increased proliferation.     

Effect of lipid-free total parenteral nutrition on hepatic drug conjugation in rats

The effect of total parenteral nutrition (TPN) on drug conjugation in male Sprague-Dawley rats was examined using a nutrition solution composed of amino acids and glucose. The overall disposition of acetaminophen including the formation kinetics of the sulfate and glucuronide metabolites was used as an in vivo probe. Selected drug metabolizing enzyme activities also were examined in vitro. TPN, 200 kcal/kg/day, was administered by continuous i.v. infusion for 14 days and changes elicited were compared to control animals allowed free access to rat chow. TPN decreased the total clearance of acetaminophen by 34% and the formation clearance to acetaminophen sulfate by 47%. The formation clearance of acetaminophen to acetaminophen glucuronide was unaffected by TPN. Cytochrome P450 concentration and oxidative demethylase activity toward p-nitroanisole were decreased in parallel, 47 and 53%, respectively, and UDP-glucuronosyltransferase activity with p-nitrophenol and acetaminophen as the acceptor aglycones was decreased 44 and 25%, respectively in the animals receiving TPN. Sulfotransferase activity toward both p-nitrophenol and acetaminophen decreased 28% in animals receiving TPN vs. ad libitum rat chow. Administration of the parenteral nutrition solution as a continuous enteral infusion via a doudenal catheter slightly decreased p-nitroanisole demethylase activity (26%), but had no other significant effects on either cytochrome P450 concentration or on drug conjugating enzyme activities determined in vitro. These results show that parenteral nutrition administered i.v. depresses drug conjugation and suggest that alterations in both hepatic oxidative and conjugative biotransformation arising from total parenteral nutrition are largely attributable to bypassing the intestinal route for nutrient intake.     

Effect of methionine-deprived total parenteral nutrition on tumor protein turnover in rats

BACKGROUND: Previous studies have shown that a methionine-lacking diet inhibited tumor growth in rats. The aim of this study was to determine how methionine free total parenteral nutrition (MTPN) can result in the inhibition of tumor growth on tumor protein metabolism in rats. METHODS: On day 0, AH109A rat ascites hepatoma cells were implanted subcutaneously into male Donryu rats (n = 68, body weight, 200-225 gm). On day 10, a catheter for total parenteral nutrition (TPN) was placed and either MTPN or standard TPN solution was given for 5 days. On day 15, 1-14C-leucine was infused continuously to measure tumor protein synthesis. Tumor proteolysis was calculated from tumor regional blood flow, using the 85Sr-microsphere injection method. RESULTS: 1) Tumor weight was reduced with MTPN. 2) MTPN did not affect tumor protein synthesis, probably because endogenous methionine production was increased with MTPN (87.3 +/- 13.5 mumole methionine/kg/hour vs. 218.6 +/- 29.5, P < 0.01); however, MTPN caused an increase of tumor proteolysis (2.68 +/- 0.53 mumole leucine/g/hour vs. 3.79 +/- 0.73, P < 0.05). CONCLUSION: The enhanced tumor protein breakdown contributed to the inhibition of tumor growth that was found with the rats given the methionine free diet.     

Adhesive properties of Vibrio cholerae: adhesion to isolated rabbit brush border membranes and hemagglutinating activity

Adhesion of vibrios to the small intestine may occur (i) by association of the bacteria with secreted mucus gel or (ii) by adherence of the bacteria to the surface of epithelial cells. In the present study, vibrios readily adhered to isolated brush border membranes obtained from rabbit intestinal epithelial cells. Adhesion was temperature dependent and required the presence of divalent cations such as calcium. The agglutination of human O erythrocytes by Vibrio cholerae was observed also, and the hemagglutination test appeared to detect the same mechanism that was involved in the adhesion of vibrios to brush borders. When the bacteria were grown in broth they were adhesive and hemagglutinating, but vibrios grown on agar plates or suspended in buffer for 15 min at 37 C lacked these abilities, even though they retained undiminished motility. These two model systems differed, however, in that strontium promoted only adhesion to brush borders. The significance of this difference remains to be determined. Vibrios were observed to penetrate intestinal mucus gel and occasionally to become entrapped in it. However, there was no evidence that vibrios attached to mucus gel.     

Home parenteral nutrition in the management of patients with chronic idiopathic intestinal pseudo-obstruction

Home parenteral nutrition is indicated in all those patients who are unable to cover all their needs orally or enterally during prolonged periods of time, and who do not require any other general care other than the parenteral nutrition. Our objective is to prove the use of home parenteral nutrition as a nutritional support in patients with severe forms of chronic idiopathic intestinal pseudo-obstruction. In our unit, three patients with this disease, have received home parenteral nutrition between 1993 and the present date. One patient received it during four months, with the catheter being removed due to a fungemia. At present she is being maintained with oral and enteral nutrition. The other two patients continue in the program: one since October 93 and the other since July 94. The hydroelectric alterations caused during the episodes of sub-occlusion make more frequent changes in the composition of the parenteral nutrition necessary, compared to other types of patients. The low incidence of complications and the degree of acceptance by the patient makes this technique an ideal method for the long term nutritional support.