OBJECTIVEThis study investigated the expression of platelet-derived growth factor-B (PDGF-B) and its receptor-β (PDGFR-β) in rat cerebral cortex following sepsis and explored the possible underlying mechanism of neuro-protective effect of glutamine (Gln).

METHODSOne hundred and twenty 10-day-old Wistar rats were randomly divided into three groups: a control group that received an intraperitoneal injection of normal saline (1 mL/kg), a sepsis group that received an intraperitoneal injection of lipopolysaccharide (LPS, 5 mg/kg), and a Gln treatment group that was administered with Gln (1.346 g/kg) 1 hr before LPS injection. The rats were subdivided into 5 groups sacrificed at 2, 6, 12, 24 and 72 hrs after LPS or normal saline injection (n=8). The distribution and expression of PDGF-B and PDGFR-β in the cerebral cortex were ascertained by immunohistochemistry and Western blot.

RESULTSThe immunohistochemistry results showed that the PDGF-B and PDGFR-β expression in the cerebral cortex increased significantly in the Gln treatment group 72 hrs after LPS injection compared with that in the control and the sepsis groups. The Western blot results showed that the PDGF-B expression in the brain tissue in the sepsis and the Gln treatment groups were significantly lower than that in the control group 2, 6, and 12 hrs after LPS injection, while the Gln treatment group had increased PDGF-B expression compared with the sepsis group 12 and 72 hrs after LPS injection. Compared with the control group, the PDGFR-β expression in the brain tissue in the sepsis group increased 2 and 6 hrs after LPS injection but decreased significantly 72 hrs after LPS injection. There were no significant differences in the PDGFR-β expression between the Gln treatment and the control groups at all different time points.

CONCLUSIONSGln can increase the PDGF-B and PDGFR-β expression in the brain tissue of rats with sepsis. The increased PDGF-B and PDGFR-β expression might contribute to neuro-protective effects of Gln.

Animals ; Brain ; Glutamine ; pharmacology ; Lipopolysaccharides ; pharmacology ; RNA, Messenger ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar

OBJECTIVEEndotoxemia is a serious syndrome resulting in multi-organ failure. Once it happens, the penetration of small intestine epithelium increases, body liquid losses, then effective circulating blood decreases and serious metabolic acidosis, serious hypotension, systolic failure, and even shock may occur. In this pathological process, endotoxin, tumor necrosis alpha and systolic dysfunction play important roles. Nowadays, many studies have been done to resolve the systolic dysfunction, but too much attention had been paid to the followings: the depressions of myocardium caused by tumor necrosis alpha, other inflammatory factors, endotoxin and metabolic acidosis; the disturbance of blood vessel-nerve regulations; nitric oxide (NO)/inducible nitric oxide synthase (iNOS) over-synthesis and the decreased density of beta-receptors in the myocardium and/or their activities. Little attention has been paid to the relationship between alpha sarcmeric actin (alpha-SA) and systolic dysfunction during endotoxemia. Glutamine (Gln) can be metabolized into glutathione, an eliminator of free radical. It has been used in preventing myocardial damage from reperfusion. This study aimed to observe the dynamic changes of alpha-SA and mRNA expressions in rats with endotoxemia and examine the effects of Gln on them.

METHODSClassical rat model of endotoxemia was established by intraperitoneal injection of LPS (4 mg/kg, Escherichia coli O55:B5, Sigma). 121 Wistar 18-day-rats were divided into three groups randomly, (1) 0 h control group (normal saline: 1 ml/kg, n = 11). (2) LPS group (LPS: 4 mg/kg, n = 55). (3) Gln group (LPS: 4 mg/kg and immediately 13.64%; Gln: 1 ml/kg, Fresenus, n = 55), Furthermore, LPS and Gln groups were divided into 2, 4, 6, 24 and 72 h time points (n = 11). Each time point of LPS and Gln as well as control rats were anaesthetized at each time point with 1% chloral hydrate injected intraperitoneally at the dosage of 1 ml/kg. Then rats were sacrificed at appoint time, and the hearts were isolated. Eight of them were put in 76 degrees C liquid nitrogen and then frozen in minute 80 degrees C icebox in order to measure the expression of alpha-SA mRNA by RT-PCR. Three of them were fixed in 4% formaldehydum polymerisatum for 12 to 16 h, then the expression of alpha-SA was detected by immunohistochemistry.

RESULTS(1) Compared to 0 h, the expressions of alpha-SA and mRNA in LPS group were significantly depressed (P < 0.01). In LPS group, the lowest was at 6 - 24 h, while in Gln group, it was postponed to 24 h. At 72 h, there was no difference in expressions of alpha-SA between Gln and 0 h group (P > 0.05). (2) Comparing at same time point, the expressions of alpha-SA were significant higher in Gln group than those in LPS group, while the expressions of alpha-SA mRNA in Gln group were high at 4-72 h. There was, however, no significant difference at early phase (P > 0.05).

CONCLUSIONAlpha-SA and its mRNA expression were depressed in LPS-induced endotoxemia, especially from 6 to 24 h. It could damage the systolic function. alpha-SA decrease in endotoxemia was due to the inhibited synthesis other than the promoted degradation. Glutamine could inhibit the effects of LPS on both alpha-SA and its mRNA expressions.

Actins ; metabolism ; Animals ; Endotoxemia ; metabolism ; Glutamine ; pharmacology ; Lipopolysaccharides ; Myocardium ; metabolism ; RNA, Messenger ; metabolism ; Rats ; Rats, Wistar

Animals ; Burns ; drug therapy ; metabolism ; Glutamine ; pharmacology ; therapeutic use ; Humans ; Wound Healing ; drug effects

Calpain I (mu-calpain) and II (m-calpain) are well known calcium-activated neutral cysteine proteases. Many reports have shown that activation of calpain is related to cataract formation, neuronal degeneration, blood clotting, ischemic injuries, muscular dystrophy and cornified cell envelope (CE) formation. Here, we report that insoluble CE formation was reduced after treatment with calpain I inhibitor (N-acetyl-leucyl-leucyl-norleucinal) on normal human epidermal keratinocytes (NHEK), whereas serine and thiol protease inhibitors had no effect on the reduction of CE. When NHEK cells were confluent, keratinocytes were treated with various concentrations (0.5 microM-0.5 mM) of calpain I inhibitor or serine and thiol protease inhibitors under calcium induced differentiation. Insoluble CE formation was reduced about 90% in the 50 microM calpain inhibitor I treated group by day 9 of culture, whereas insoluble CE was reduced only 10% in the same condition. Interestingly TGase activity was blocked by 90% in the 0.5 mM calpain inhibitor treated group within 72 h, whereas TGase activity was retained by 80% in the 0.5 mM serine protease inhibitor treated group at 7 day treatment. Therefore it can be suggested that cysteine protease calpains might be responsible for the activation of the TGase 1 enzyme to complete insoluble CE formation during epidermal differentiation.
Calcium/pharmacology ; Calpain/metabolism* ; Calpain/antagonists & inhibitors* ; Cell Differentiation ; Dose-Response Relationship, Drug ; Epidermis/metabolism ; Human ; In Vitro ; Keratinocytes/metabolism ; Keratinocytes/enzymology ; Protease Inhibitors/pharmacology ; Protein-Glutamine gamma-Glutamyltransferase/metabolism* ; Protein-Glutamine gamma-Glutamyltransferase/antagonists & inhibitors* ; Tissue Culture

Animals ; Cadmium/toxicity* ; Dysbiosis/metabolism* ; Gastrointestinal Microbiome ; Glutamine/pharmacology* ; Methionine ; Mice ; Mice, Inbred C57BL ; Ostreidae ; Protein Hydrolysates ; Tyrosine

The relevance of transglutaminases with neural function and several disorders has been emphasized recently. Especially, many polypeptides associated with neurodegenerative diseases are suggested to be putative transglutaminase substrates such as beta amyloid protein of Alzheimer's disease, microtubule-associated proteins and neurofilaments, etc. In addition, the CAG repeated gene products with probable polyglutamine tract, putative transglutaminase substrates, were identified in several neurodegenerative disorders. However, the identity of the brain transglutaminase has not been confirmed, because of enzymic stability and low activity. In the present experiment, we have isolated brain-specific transglutaminases, designated as TGase NI and TGase NII, which are different from other types of transglutaminases in respects of molecular weights (mw. 45 kDa, 29 kDa respectively), substrate affinity, elution profile on ion-exchange chromatography, sensitivity to proteases and ethanol, and immunological properties. The enzymes were localized specifically in the brain tissues but not in the liver tissue. And neural cells such as pheochromocytoma cell, glioma cell, primary neuronal and glial cells were shown to be enriched with TGase NI and TGase NII. The possible biological roles of the enzymes were discussed not only on the aspect of crosslinking activity but also of signal transducing capacity of the enzyme in the brain.
Animal ; Astrocytes/enzymology ; Blotting, Western ; Brain/enzymology* ; Calcium/metabolism ; Chromatography, Ion Exchange ; Endopeptidases/pharmacology ; Enzyme Stability ; Ethanol/pharmacology ; Glioma ; Immunoblotting ; Immunohistochemistry ; Male ; Molecular Weight ; Neurons/enzymology ; PC12 Cells ; Protein-Glutamine gamma-Glutamyltransferase/isolation & purification* ; Protein-Glutamine gamma-Glutamyltransferase/immunology ; Protein-Glutamine gamma-Glutamyltransferase/chemistry* ; Rats ; Rats, Sprague-Dawley ; Trypsin/pharmacology ; Tumor Cells, Cultured

BACKGROUND/AIMS: NSAIDs induce gut damage throughout the entire gastrointestinal tract and bacterial translocation. The aim of this study was to examine if administration of glutamine was able to prevent the NSAID-induced gut damages and bacterial translocation in the animal models. METHODS: Rats were utilized into 5 groups; control group, diclofenac group, and diclofenac with glutamine 0.8, 1.6, and 3.2 g/kg/day group. The animals with glutamine were fed with L-glutamine for 4 days before diclofenac administration. Gut injury was induced by administration of a single dose of diclofenac (80 mg/kg orally). Intestinal permeability (24 hour urinary excretion of phenolsulfonphthalein), enteric aerobic bacterial counts, serum biochemical profiles and bacterial translocation to mesenteric lymph nodes, liver and spleen were measured. RESULTS: Diclofenac caused the increase in intestinal permeability, enteric bacterial count, enteric protein and albumin loss and bacterial translocation. Administration of glutamine reduced the increase in intestinal permeability, protein losing enteropathy, enteric bacterial overgrowth and bacterial translocation induced by diclofenac. CONCLUSIONS: Glutamine may have beneficial effects on NSAID-induced gut damage and bacterial translocation.
Animals ; Anti-Inflammatory Agents, Non-Steroidal/*pharmacology/toxicity ; Bacterial Translocation/*drug effects ; Diclofenac/*pharmacology/toxicity ; English Abstract ; Glutamine/*pharmacology ; Intestines/drug effects/*microbiology ; Male ; Rats

OBJECTIVETo evaluate the effect of combination of glutamine (GLN) and mitomycin C (MMC) on the human gastric carcinoma cell line MGC-803 in vitro.

METHODSThe effects of GLN and MMC were measured by MTT assay, and the interaction between the two agents was evaluated by the median-effect principle. Flow cytometry was used for cell cycle analysis.

RESULTSGLN did not significantly stimulate the cell growth in vitro. High-concentration of GLN could inhibit the cell growth. MMC could effectively inhibit the cell growth in a time-dependent manner. The interaction of these two agents showed a weak antagonistic activity (1 < CI < 1.2703). MMC induced remarkable S-phase arrest. Low-dose GLN has limited effect on the S-phase arrest of MMC, while high-dose GLN significantly attenuated the S-phase arrest and lowered the proliferation index of MGC-803 cell.

CONCLUSIONSCombination of GLN and MMC has a a weak and dose-dependent antagonistic activity in the treatment of gastric carcinoma cell line MGC-803. The combination of high-dose MMC and low-dose GLN may achieve better efficacy.

Adenocarcinoma ; pathology ; Antibiotics, Antineoplastic ; pharmacology ; Cell Line, Tumor ; Dose-Response Relationship, Drug ; Drug Synergism ; Glutamine ; pharmacology ; Humans ; Mitomycin ; pharmacology ; Stomach Neoplasms ; pathology

OBJECTIVETo investigate the mechanism that glutamine (Gln) downregulates the cytokine expression in lipopolysaccharide (LPS)-stimulated human peripheral blood mononuclear cells (PMBCs).

METHODSPMBCs were extracted from healthy volunteer by density gradient centrifugation, the cells were divided into two parts. The first part of PMBCs was pretreated with Gln of the concentration of 0, 8, 15 mmol/L for 0.5 h and 2.0 h respectively, then stimulated by LPS for 4.0 h. Cells and supernatants were collected. The second part of PBMCs was divided into group A, B and C. Group A and B were pretreated with HSP70 blocker (Quercetin) for 1.0 h, then were stimulated by LPS for 4.0 h. Cells and supernatants were also collected. The release of TNF-alpha and IL-10 was analyzed via enzyme-linked immunosorbent assay (ELISA) and HSP70 via Western Blot. In this experiment, the effect of Quercetin on TNF-alpha, IL-10 and HSP70 expression in human PBMCs was assessed.

RESULTSGln led to an increase in HSP70 expression, and decreased TNF-alpha, IL-10 release at 4.0 h after LPS stimulation when 8 mmol/L glutamine pretreated for 0.5 h and 2.0 h, 15 mmol/L glutamine pretreated for 0.5 h (P < 0.05). The expression level of HSP70 was significantly decreased, however, the expression of TNF-alpha and IL-10 was enhanced in Quercetin group (P < 0.05).

CONCLUSIONThe effect of glutamine attenuating cytokine release in PBMCs is related to the enhancement of HSP70 expression.

Cells, Cultured ; Cytokines ; metabolism ; Glutamine ; pharmacology ; HSP70 Heat-Shock Proteins ; metabolism ; Humans ; Interleukin-10 ; metabolism ; Leukocytes, Mononuclear ; drug effects ; metabolism ; Lipopolysaccharides ; pharmacology ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo study the effects of resveratrol on amino acid release in cultured astrocytes with stretch injury and explore its possible mechanism for central nervous system protection.

METHODSCultured rat astrocytes were treated with different concentrations of resveratrol for 12 h and the subsequent changes in Glu and Gln release and lactate dehydrogenase(LDH) leakage were examined after stretch injury.

RESULTSHigh-performance liquid chromatography showed that stretch injury increased Glu release from the astrocytes (P<0.05), and at the concentration of 1 micromol/L, resveratrol further increased Glu release (P<0.05), whereas at 100 micromol/L, resveratrol obviously inhibit Glu release (P<0.05). LDH leakage increased significantly after stretch injury of the astrocytes (P<0.05), and resveratrol acted to further increase LDH leakage at 1 micromol/L but inhibited LDH leakage at 100 micromol/L (P<0.05).

CONCLUSIONResveratrol can inhibit the release of Glu and LDH leakage from rat astrocytes to protect the cells from stretch injury.

Animals ; Animals, Newborn ; Astrocytes ; cytology ; metabolism ; Cells, Cultured ; Glutamine ; metabolism ; Glutathione ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Neuroprotective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Stilbenes ; pharmacology ; Stress, Mechanical