Myoelectric activity of the ileum, cecum, and right ventral colon, and cecal emptying of radiolabeled markers in clinically normal ponies

1. The activities of the two key enzymes involved in glutamine metabolism, glutaminase and glutamine synthetase, were measured in mucosal biopsies taken from different sites throughout the human gastrointestinal tract, from oesophagus to rectum. 2. The specific activity of glutamine synthetase was highest in the stomach (4.5 nmol glutamine formed per minute per mg of protein), but both small and large intestine and the oesophagus had little synthesizing capacity (less than 0.3 nmol of glutamine formed per minute per mg of protein). 3. Glutaminase specific activity was highest in the small intestine (53 nmol glutamate formed per minute per mg of protein by duodenal mucosa), intermediate in the large intestine and lowest in the oesophagus and stomach (less than 13 nmol of glutamate formed per minute per mg of protein). 4. The glutamine concentration in the mucosa was lower in the duodenum than in the colon (0.62 and 0.95 mmol/kg wet weight respectively), but both were much lower than the measured K(m) values of glutaminases obtained from these sites (3.8 and 4.0 nmol/kg wet weight respectively). 5. The concentration of glutamine in saliva, stomach juice, bile and duodenal juice suggests that very little glutamine passes into the gastrointestinal tract via these secretions. 6. The study provides the most complete information on the distribution of glutamine synthetase and glutaminase along the human gastrointestinal tract, and suggests that (i) both the small and large intestines have a high potential for glutamine metabolism, but little synthesizing capacity, thus both must derive their glutamine from other sources, and (ii) neither the stomach nor the oesophagus have a high glutaminase activity, although the stomach has substantial capacity to synthesize glutamine. The distribution of the enzymes along the gastrointestinal tract may help rationalize the use of glutamine for treating diseases that affect different parts of the gastrointestinal tract.     

The proposed role of glutamine in some cells of the immune system and speculative consequences for the whole animal

The activity of glutaminase is high in lymphoid organs, lymphocytes and macrophages and increases in the popliteal lymph node in response to an immunological challenge. Consistent with this high activity, glutamine is utilised at a high rate by resting lymphocytes and macrophages in culture. Mitogenic stimulation of lymphocytes increases both glutaminase activity and the rate of glutamine utilisation. The major products of glutamine utilisation by lymphocytes and macrophages in culture are glutamate, aspartate, lactate and ammonia; < 25% of the glutamine used is completely oxidised. It is suggested that the high rate of glutamine utilisation by cells of the immune system serves to maintain a high intracellular concentration of intermediates of biosynthetic pathways such that optimal rates of DNA, RNA and protein synthesis can be maintained. In the absence of glutamine, lymphocytes do not proliferate in vitro; proliferation increases greatly as the glutamine concentration increases. The synthesis of interleukin-2 by lymphocytes and of interleukin-1 by macrophages is glutamine-dependent. Macrophage-mediated phagocytosis is influenced by glutamine availability. Glutamine is synthesized in skeletal muscle. Skeletal muscle and plasma glutamine levels are lowered by sepsis, injury, burns, surgery and endurance exercise and in the overtrained athlete. These observations indicate that a significant depletion of the skeletal muscle glutamine pool is characteristic of trauma and it has been suggested that the lowered plasma glutamine concentration contributes, at least in part, to the immunosuppression which accompanies such situations. Beneficial effects of the provision of glutamine or its precursors have been reported in patients following surgery, radiation treatment or bone marrow transplantation or suffering from injury, sepsis or burns.     

Regulation of hepatic glutaminase in the streptozotocin-induced diabetic rat

The liver of diabetic animals removes increased quantities of glutamine. We therefore examined factors that affect hepatic glutaminase activity in hepatocytes and mitochondria. Glutamine use, through glutaminase, was measured in isolated rat hepatocytes by monitoring the production of 14CO2 from [1-(14)C]glutamine. Hepatocytes from streptozotocin-induced diabetic rats use glutamine more rapidly than do hepatocytes from normal or insulin-maintained diabetic rats. Glutamine use in all of these hepatocytes was stimulated by glucagon and epinephrine. Glutaminase activity, assayed in broken mitochondrial membranes, was increased approximately 2.5-fold in diabetic rats. The sensitivity of glutaminase, measured in intact liver mitochondria, to phosphate was markedly left-shifted in mitochondria from diabetic rats compared with those from controls. In fact, glutaminase was increased 10-fold at 2.5 mmol/l phosphate compared with controls. This increased sensitivity of glutaminase to physiological concentrations of phosphate is characteristic of its hormonal activation. Therefore, activation of glutaminase plays a major role in diabetes and is as important as increases in its total enzyme amount in determining the increased glutamine uptake in diabetes.     

The role of glutamine, serum and energy factors in growth of enterocyte-like cell lines

BACKGROUND: Glutamine is routinely added to most cell cultures. Glutamine has been found to be the preferential nutrient to the rapidly replicating intestinal mucosa, but whether this is a metabolic effect or due to other properties of this amino acid is not determined. To study the importance of glutamine on the growth of two enterocyte-like cell lines, the effects of depriving the media or supplementing it with glutamine were assessed in media with different serum and energy supplements. METHODS: CaCo-2 and HT-29 cells were grown in serum-free medium, with fetal bovine or synthetic serum, and with or without glucose or galactose. The glutamine content was varied between 0 and 4 mM. All growth assays were performed in triplicate by counting in a hemocytometer. RESULTS: Both cell lines were dependent of serum factors for growth, but displayed distinct requirements on glutamine supplementation. Glutamine was an obligate supplement with dose-dependent correlation to growth (r = 0.87, p < 0.01) for CaCo-2 cells cultured in synthetic, but not in fetal bovine serum. In HT-29 cells, the correlation between glutamine and growth was significant (r = 0.68, p < 0.05) only in fetal bovine serum in the absence of galactose. CONCLUSION: This study shows that glutamine has different growth stimulating effects on two enterocyte-like cell lines studied. This could reflect different modes of action of glutamine on proliferation and differentiation in an enterocyte cell population.     

Investigation on glutamine amidohydrolase (EC 3.5.1.2) and glutamine aminotransferase (EC 2.5.1.15) activity in liver and plasma of EAC-bearing mice following glutaminase therapy

The anti-neoplastic activity of bacterial glutaminase on Ehrlich ascites tumor-bearing mice was studied by determining the reduction in the tumor cell count and extension of life span of the host after therapy. The therapeutic effect of glutaminase in relation to change in activity of glutaminolytic enzymes (glutamine amidohydrolase (GNase) and glutamine aminotransferase (GAt)) in liver and plasma were also studied. Bacterial glutaminase was shown to be effective in lowering the tumor burden with increased life span of the host. Glutamine amidohydrolase activity in the liver and plasma was raised significantly with increased tumor burden, whereas GAt activity remained unchanged. Following glutaminase therapy, this high level of GNase activity decreased in comparison to the untreated control. These changes were not seen when normal mice were treated with the same enzyme. Thus alteration in the enzyme levels, particularly GNase was observed to have some correlation with progression of the tumor growth.     

No differences in mucosal adaptive growth one week after intestinal resection in rats given enteral glutamine supplementation or deprived of glutamine

OBJECTIVE: To evaluate the effects of intraluminal glutamine on the adaptation of intestinal mucosa after resection compared with transsection and un-operated on control animals. DESIGN: Open, controlled, experimental study. SETTING: University hospital, Sweden. SUBJECTS: 123 Sprague-Dawley rats. INTERVENTION: Daily isonitrogenous oral diet was given either free of glutamine or supplemented with 4% glutamine for 2 or 7 days to rats subjected to intestinal resection, transection or no operation. MAIN OUTCOME MEASURES: Body weight and protein content, DNA content, and thymidine incorporation in jejunal and ileal mucosa. RESULTS: Resection resulted in a significant growth stimulation evaluated by weight/body weight, protein, and DNA content (p < 0.05-0.001). Glutamine supplementation did not significantly influence this growth response. Thymidine incorporation in jejunum was stimulated by glutamine on day 3 (p < 0.05-0.001). CONCLUSION: The glutamine fortified diet had no growth stimulating effects compared with a glutamine free diet one week after 60% intestinal resection. An early increase in thymidine incorporation indicated that glutamine had a transient proliferative effect.     

Glutamine deprivation induces apoptosis in intestinal epithelial cells

BACKGROUND: Glutamine is the most abundant amino acid in the blood, and its deprivation leads to gut mucosal atrophy. The small intestinal mucosa is maintained by a balance between cell proliferation and cell death by apoptosis. We reported that glutamine is required for nitrogen-stimulated proliferation in intestinal epithelial cells. We do not know whether glutamine regulates apoptosis in the gut. The purpose of this study is to determine whether glutamine deprivation induces apoptosis in rat intestinal epithelial (RIE-1) cells and to compare the effect of glutamine starvation with that of methionine and cysteine (Met/Cys) starvation. METHODS: RIE-1 cells were deprived of either glutamine or Met/Cys for 24 hours. Cell numbers were determined by cell counting and tetrazolium enzymatic assay. Apoptosis was quantified by Annexin V assay and confirmed by DNA gel electrophoresis and Hoecsht nuclear staining. RESULTS: Deprivation of glutamine or Met/Cys resulted in decreased cell numbers. However, only the glutamine-deprived group showed significant induction of apoptosis with increased Annexin V staining, DNA laddering, and nuclear condensation. CONCLUSIONS: This study provides biochemical and morphologic evidence that glutamine deprivation induces apoptosis in rat intestinal epithelial cells. In contrast, Met/Cys starvation suppresses cell number without induction of apoptosis. These results suggest that glutamine serves as a specific survival factor in enterocytes.     

Effect of L-glutamine and n-butyrate on the restitution of rat colonic mucosa after acid induced injury

BACKGROUND: L-glutamine and n-butyrate are important nutrients for colonocytes affecting both their structure and function. The effect of these epithelial substrates on resealing of rat distal colon after acid induced injury was studied. METHODS: Isolated colonic mucosa of 32 rats was mounted in Ussing chambers and exposed to Krebs-Ringer solution for four hours. Epithelial injury was induced by short-term exposure to luminal hydrochloric acid and resealing was studied with or without added glutamine or butyrate. RESULTS: Glutamine (luminal and serosal) reduced tissue conductance, mannitol and lactulose permeability, and permeation of enteropathogenic Escherichia coli. Glutamine (serosal) diminished conductance and mannitol permeability. Both interventions stimulated bromodeoxyuridine incorporation in nuclei of colonocytes. Luminal butyrate had no measurable effect on these parameters. CONCLUSIONS: These data suggest that L-glutamine stimulates repair mechanisms of rat colonic mucosa after acid injury. This effect on the gut barrier is associated with a stimulation of crypt cell proliferation. The addition of glutamine to parenteral solutions may be beneficial for patients under intensive care whose intestinal barrier is weakened in the course of sepsis and trauma.     

Effect of physiological hyperinsulinemia on blood flow and interstitial glucose concentration in human skeletal muscle and adipose tissue studied by microdialysis

Glutamine supplementation is beneficial for preventing intestinal atrophy and maintaining mucosal functions in metabolically stressed patients. The mechanisms by which glutamine prevents mucosal atrophy remain unclear. In particular, the role of glutamine in the survival of cells under stress is unknown. Intestinal epithelial cells (IEC-6) were cultured in media with or without supplementation of L-glutamine. A low concentration of L-glutamine (1.0 mmol/L) was sufficient to minimize the percentage of floating cells under basal conditions. Heat shock at 43 degrees C for 90 min decreased (P < 0. 001) the number of attached cells, while increasing (P < 0.001) the number of floating cells, which is a measurement of the extent of cell death in these cultures. Glutamine enhanced attached cell count and diminished heat shock-induced cell death in a dose-dependent manner. Of note, 2 mmol/L was suboptimal in both respects, thus indicating that heat-shocked cells require higher concentrations of glutamine for optimal cell survival. Maximal effect was achieved with 8 mmol/L glutamine, which increased (P < 0.001) cell growth (indicated by the number of attached cells) and diminished (P < 0. 001) cell death (indicated by the number of floating cells). Further increase of L-glutamine concentration to 12 or 20 mmol/L did not provide additional benefit in minimizing cell death. Heat shock protein 70 (hsp 70) mRNA was induced by heat shock only in cultures supplemented with L-glutamine, and the induction was more consistent and greater in cultures containing higher concentrations of glutamine. Thus, glutamine supplementation reduced heat shock-induced cell death. This effect, together with the maintenance of cell growth, may play a key role in the prevention of intestinal mucosal atrophy.     

Use of ultravist to examine the gastrointestinal tract in patients with emergency abdominal pathology

The paper deals with the results of using the nonionic water soluble contrast agents ultravist 300 and ultravist 370 (Schoring, Germany) to examine the gastrointestinal tract (GIT) in 21 patients with acute abdominal abnormality. GIT contrasting was made in 9 patients in the early postoperative period and in 12 patients on their admission to the Institute. The examinations revealed the high contrast rate of the above agent when administered into the stomach and upper small intestine, which excluded failure of gastroenteroanastomic sutures and the sutured gastric wall in 2 patients, established, in terms of gastric displacement and deformity, left-sided subdiaphragmatic abscess, and in terms of transposition of a portion of the gastric fornix into the pleural cavity, rupture of the left diaphragm, and to exclude diaphragmatic rupture with closed abdominal injury. The revealed important quality of ultravist within a short time (1-2 hours) to contrast the small intestine and enter the colon enabled differential diagnosis to be made between complete and partial small intestinal ileus and between early comissural small intestinal ileus and postoperative intestinal paresis. Ultravist contrast studies allowed the authors to avoid an emergency operative intervention and to follow up the resolution of ileus during medical therapy.     

Adaptive alterations in cellular metabolism with malignant transformation

OBJECTIVE: The authors studied the differences between glutamine and glucose utilization in normal fibroblasts and in fibrosarcoma cells to gain insights into the metabolic changes that may occur during malignant transformation. SUMMARY BACKGROUND DATA: The process of malignant transformation requires that cells acquire and use nutrients efficiently for energy, protein synthesis, and cell division. The two major sources of energy for cancer cells are glucose and glutamine. Glutamine is also essential for protein and DNA biosynthesis. We studied glucose and glutamine metabolism in normal and malignant fibroblasts. METHODS: Studies were done in normal rat kidney fibroblasts and in rat fibrosarcoma cells. We measured glutamine transport across the cell membrane, breakdown of glutamine by the enzyme glutaminase (the first step in oxidation), glutamine and glucose oxidation rates to CO2, rates of protein synthesis from glutamine, and glutamine-dependent growth rates. RESULTS: Glutamine transport rates were increased more than sixfold in fibrosarcomas compared to normal fibroblasts. In fibroblasts, glutamine transport was mediated by systems ASC and A. In malignant fibrosarcomas, only system ASC was identifiable, and its Vmax was 15 times higher than that observed in fibroblasts. Despite an increase in transport, glutaminase activity was diminished and glutamine oxidation to CO2 was reduced in fibrosarcomas versus normal fibroblasts. In fibroblasts, glutamine oxidation was 1.8 times higher than glucose oxidation. In contrast, glucose oxidation was 3.5 times greater than glutamine oxidation in fibrosarcomas. Protein synthesis from glutamine transported by fibrosarcomas was threefold greater than that observed in normal fibroblasts. Despite marked increases in glutamine utilization and glucose oxidation in fibrosarcoma cells, growth rates were higher in the normal fibroblasts. CONCLUSIONS: The process of malignant transformation is associated with a marked increase in cellular glutamine transport, which is mediated by a single high-affinity, high-capacity plasma membrane carrier protein. In normal fibroblasts, the transported glutamine is used primarily for energy production via oxidation of glutamine carbons to CO2. In fibrosarcomas, glutamine oxidation falls and glutamine is shunted into protein synthesis; simultaneously, the malignant cell switches to a glucose oxidizer. The increased glutamine transport and glucose oxidation in fibrosarcomas appears to be related to the malignant phenotype and not merely to an increase in cell growth rates.     

Inverse relationship between fenfluramine-induced prolactin release and blood pressure in humans

OBJECTIVES: Up to 20% of patients with AIDS have abnormal intestinal permeability (IP). Glutamine seems to play an important role in preventing the increase in IP and loss of intestinal mucosal mass associated with total parenteral nutrition, and may be superior to glucose for oral rehydration in the setting of intestinal infection. This study was designed to see if supplemental glutamine could alter the abnormal IP of AIDS. METHODS: Randomly chosen patients with AIDS from the Jacobi Medical Center human immunodeficiency virus (HIV) clinic underwent IP testing using lactulose and mannitol. Those with abnormal IP were enrolled. Duodenal biopsies were performed with a Crosby capsule and the patients were randomized in a double-blind fashion to receive placebo or glutamine (4 g/day or 8 g/day) for 28 days, after which intestinal permeability tests and duodenal biopsies were repeated. Intestinal morphology was graded by ratio of villus height to crypt depth, and by degree of inflammation. RESULTS: All patients complied with the therapy and there were no dropouts or reported side effects. The results showed less worsening of IP with the 4 g/day dose, compared with placebo. At the 8 g/day dose, there was stabilization of IP and improved absorption of mannitol. Intestinal morphology and inflammation did not change in any group. CONCLUSIONS: These results, although not significant, suggest a trend towards improved IP and enhanced intestinal absorption with glutamine. Glutamine doses of at least 20 g/day may be necessary to improve IP. We recommend further studies at higher doses and for longer durations.     

Oral glutamine reduces the duration and severity of stomatitis after cytotoxic cancer chemotherapy

BACKGROUND: Mouth sores and/or difficulty swallowing are common and painful consequences of cytotoxic chemotherapy for cancer. In previous studies oral glutamine was found to protect animals from the effects of whole abdominal radiation and methotrexate-induced enteritis. Glutamine also was found to reduce oral mucositis in a nonrandomized pilot study in humans. Therefore, the authors attempted to determine the efficacy of oral glutamine in a randomized, double blind, crossover trial in cancer patients receiving chemotherapy. METHODS: Twenty-four patients (16 children and 8 adults) received glutamine or placebo (glycine) suspension (2 g amino acid/M2/dose twice daily) to swish and swallow on days of chemotherapy administration and for at least 14 additional days. Patients completed a calendar indicating days of mouth pain associated with each chemotherapy course and the effect of mouth pain on oral intake. RESULTS: Paired data indicated significant amelioration of stomatitis associated with glutamine administration after chemotherapy. The duration of mouth pain was 4.5 days less in chemotherapy courses in which glutamine supplementation was compared with placebo (Wilcoxon's signed rank test, P=0.0005). The severity of oral pain also was reduced significantly when glutamine was provided with chemotherapy (the amount of days mucositis restricted oral intake to soft foods [> or =Grade 2; Modified Eastern Cooperative Oncology Group grading system] was 4 days less with glutamine compared with placebo; Wilcoxon's signed rank test, P=0.002). CONCLUSIONS: Low dose oral glutamine supplementation during and after chemotherapy significantly reduced both the duration and severity of chemotherapy-associated stomatitis. Oral glutamine appears to be a simple and useful measure to increase the comfort of many patients at high risk of developing mouth sores as a consequence of intensive cancer chemotherapy.     

In vitro mucosal digestion of synthetic gliadin-derived peptides in celiac disease

Two celiac-active synthetic peptides derived from the A-gliadin structure corresponding to residues 8-19 (LQPQNPSQQQPQ) and to 11-19 were digested in vitro with small intestinal mucosa from children with celiac disease in remission and from normal children. The products of digestion were separated into two fractions on the basis of M(r) < 400 and M(r) > 400 by gel permeation chromatography and subjected to amino acid analysis. After digestion of the dodecapeptide with celiac mucosa, 71 +/- 14% (molar) of the total digestion products remained in the M(r) > 400 fraction. Glutamine, proline, serine, and asparagine were the major amino acids present. Glutamine, proline, and leucine were the major amino acids in the M(r) < 400 fraction. The M(r) > 400 fraction from the celiac mucosal digestion of the nonapeptide was of similar composition to the corresponding fraction from the dodecapeptide and represented 78 +/- 15% of the total products. Digestion of the two peptides with normal mucosa gave lower amounts of products in the M(r) > 400 fraction, but they were of similar composition to the corresponding fractions from the celiac mucosal digestion. Peptides such as NPSQQP and QNPSQQQ may be present in the M(r) > 400 fractions since glutamine and proline are present in the approximate ratio of 2:1, respectively. The results indicate a defect in the mucosal digestion of peptides which are active in an animal model of celiac disease.     

Glutamine metabolism is changed in lymphocytes from aged rats

Changes in the protein content, maximal activity, and Km of phosphate-dependent glutaminase were measured in the lymphoid organs (thymus, spleen, and mesenteric lymph nodes) from just-weaned, mature (3 months), and aged rats (15 months). Also, [U-14C] glutamine transport and decarboxylation and the production of glutamate and aspartate from 2 and 20 mM glutamine were measured in incubated mesenteric lymph node lymphocytes. The ageing process induced a reduction in the protein content of the thymus and spleen, as well as the phosphate-dependent glutaminase activity in the thymus and isolated lymphocytes. The Km of phosphate-dependent glutaminase, however, was not affected by the process. Ageing reduced [U-14C] glutamine decarboxylation and increased glutamate and aspartate production in incubated lymphocytes. The results indicate that the ageing process does modify several aspects of glutamine metabolism in lymphocytes: reduces maximal glutaminase activity and [U-14C] glutamine decarboxylation and increases the Km for [U-14C] glutamine uptake and the production of glutamate and aspartate.     

Parenteral glutamine dipeptide supplementation does not ameliorate chemotherapy-induced toxicity

BACKGROUND: Glutamine-supplemented total parenteral nutrition (TPN) improved the nitrogen balance in catabolic situations. In animal studies, parenteral glutamine supplementation appeared to maintain gut integrity. This study was performed to evaluate the possible positive effects of glutamine supplementation in catabolic hematologic patients. METHODS: This was a prospective double-blind placebo-controlled pilot study, in which 20 treatment cycles in unselected hematologic patients with intensive chemotherapy were studied. Glutamine was given as a dipeptide. Patients were randomized per treatment cycle to receive isonitrogenous TPN (0.272 g nitrogen/kg of body weight) and isoenergetic TPN (2200 kcal NPE/day) without or with 40 g L-alanyl-L-glutamine (26 g glutamine) until the neutrophil count was greater than 0.5 x 10(9)/L. The daily oral food intake was recorded and analyzed carefully. Toxicity grades for performance status, mucositis, and diarrhea were scored according to the World Health Organization classification. RESULTS: No differences in neutropenic period, fever, extra antibiotics, and toxicity scores were observed, except for a gain in body weight per treatment cycle in favor of the glutamine-supplemented TPN. No side effects or allergic reactions were noted after the dipeptide administration. CONCLUSION: Supplementation of glutamine dipeptide was safe but had no significant positive clinical effect.     

Barrett esophagus--an increasing problem

There has been a considerable increase in the incidence of adenocarsinoma in the proximal stomach (cancer of the cardia) and distal oesophagus (Barrett's cancer) for the past 20 years. There is probably also a parallel increase in the pathogenetically related conditions reflux oesophagitis and Barrett's oesophagus. In patients with classical Barrett's oesophagus, i.e., metaplastic changes in the mucosa more than 3 cm up from the gastro-oesophageal junction, a follow-up programme with endoscopy and adequate biopsies is recommended in cases where a finding of premalignant changes or malignancy will have therapeutic consequences. In "short segment" Barrett's oesophagus it is still not clear how extensive the biopsy and follow-up programme should be. It is also not clear whether other tests should be performed. Screening for malignancy, possibly by means of cancer-markers, and local treatment modalities of (pre-) malignant changes, are interesting possibilities that are being investigated.     

Role of glutamine in immunologic responses

Glutamine has traditionally been thought of as a nonessential amino acid, but laboratory and clinical data suggests that it may be essential during certain inflammatory conditions, such as infection and injury. Glutamine is a necessary nutrient for cell proliferation, serves as a specific fuel for inflammatory cells and enterocytes, and, when present in appropriate concentrations, enhances cell function. During inflammatory states, glutamine consumption may outstrip endogenous production and a relative glutamine deficiency state may exist. Animal and clinical studies suggest that improved outcome may be possible by providing the appropriate dose of this nutrient by the appropriate route to achieve adequate tissue concentrations. Such an approach prevents patients from being exposed to some of the inadequacies of present day conventional nutrition. The overall benefit of providing an appropriate glutamine-supplemented diet to all metabolically compromised patients arises from the multiple anabolic and host protective effects of this amino acid, of which immunomodulation is only one important facet of glutamine's essential nature.