Synergistic Effects of Glutamine Deprivation and Metformin in Acute Myeloid Leukemia

Objective The metabolic reprogramming of acute myeloid leukemia(AML)cells is a compensatory adaptation to meet energy requirements for rapid proliferation.This study aimed to examine the synergistic effects of glutamine deprivation and metformin exposure on AML cells.Methods SKM-1 cells(an AML cell line)were subjected to glutamine deprivation and/or treatment with metformin or bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl)ethyl sulfide(BPTES,a glutaminase inhibitor)or cytarabine.Cell viability was detected by Cell Counting Kit-8(CCK-8)assay,and cell apoptosis and reactive oxygen species(ROS)by flow cytometry.Western blotting was conducted to examine the levels of apoptotic proteins,including cleaved caspase-3 and poly(ADP-ribose)polymerase(PARP).Moreover,the human long noncoding RNA(lncRNA)microarray was used to analyze gene expression after glutamine deprivation,and results were confirmed with quantitative RT-PCR(qRT-PCR).The expression of metallothionein 2A(MT2A)was suppressed using siRNA.Cell growth and apoptosis were further detected by CCK-8 assay and flow cytometry,respectively,in cells with MT2A knockdown.Results Glutamine deprivation or treatment with BPTES inhibited cell growth and induced apoptosis in SKM-1 cells.The lncRNA microarray result showed that the expression of MT family genes was significantly upregulated after glutamine deprivation.MT2A knockdown increased apoptosis,while proliferation was not affected in SKM-1 cells.In addition,metformin inhibited cell growth and induced apoptosis in SKM-1 cells.Both glutamine deprivation and metformin enhanced the sensitivity of SKM-1 cells to cytarabine.Furthermore,the combination of glutamine deprivation with metformin exhibited synergistic antileukemia effects on SKM-1 cells.Conclusion Targeting glutamine metabolism in combination with metformin is a promising new therapeutic strategy for AML.

Glutamine transporters as effective targets in digestive system malignant tumor treatment

Glutamine is one of the most abundant non-essential amino acids in human plasma and plays a crucial role in many biological processes of the human body.Tumor cells take up a large amount of glutamine to meet their rapid proliferation requirements,which is supported by the upregulation of glutamine transporters.Targeted inhibition of glutamine transporters effectively inhibits cell growth and proliferation in tumors.Among all cancers,digestive system malignant tumors(DSMTs)have the highest incidence and mortality rates,and the current therapeutic strategies for DSMTs are mainly surgical resection and chemotherapy.Due to the relatively low survival rate and severe side effects associated with DSMTs treatment,new treatment strategies are urgently required.This article summarizes the glutamine transporters involved in DSMTs and describes their role in DSMTs.Additionally,glutamine transportertarget drugs are discussed,providing theoretical guidance for the further development of drugs DSMTs treatment.

Environmental enrichment in combination with Bifidobacterium breve HNXY26M4 intervention amplifies neuroprotective benefits in a mouse model of Alzheimer's disease by modulating glutamine metabolism of the gut microbiome

The gut microbiota-brain axis has emerged as a novel target for Alzheimer's disease(AD),a neurodegenerative disease characterised by behavioural and cognitive impairment.However,most previous microbiome-based intervention studies have focused on single factors and yielded only modest cognitive improvements.Here,we proposed a multidomain intervention strategy that combined Bifidobacterium breve treatment with environmental enrichment(EE)training.In this study,we found that compared with EE or B.breve treatment alone,B.breve intervention combined with EE amplified its neuroprotective effects on AD mice,as reflected by improved cognition,inhibited neuroinflammation and enhanced synaptic function.Moreover,using microbiome and metabolome profiling,we found that the combination of B.breve and EE treatment restored AD-related gut microbiota dysbiosis and reversed microbial metabolite changes.Finally,by integrating behavioural and neurological data with metabolomic profiles,we revealed that the underlying mechanism may involve the modulation of microbiota-derived glutamine metabolism via gut-brain interactions.Collectively,combined B.breve intervention with EE treatment can alleviate AD-related cognitive impairment and improve brain function by regulating glutamine metabolism of the gut microbiome.Our findings provide a promising multidomain intervention strategy,with a combination of dietary microbiome-based and lifestyle-targeted interventions,to promote brain function and delay the progression of AD.

Effects of Supplemental Glutamine and Lysine on Growth Performance of Broiler Chickens

The optimum levels of Lysine and Glutamine needed for growth performance and maintenance of the chicken broilers were evaluated in a randomized 3 × 4 factorial arrangement of dietary treatments. The battery cages measured 99 × 66 × 25 cm that can be sufficient for 5 birds. Day old Chicken broilers totaling 180 were assigned to dietary treatments comprising of 3 concentrations of Lysine (0.85, 1.14, and 1.42) each in combination with 4 concentrations of Glutamine (0, 1, 2, and 3). Each dietary treatment was replicated 3 times and each replication had 5 birds. The birds were given feed and water ad libitum with a 23-hour light regimen for a period of 4 weeks. Then, the experimental birds were evaluated for body weight gain, feed consumption, and feed conversion in order to determine their optimum requirement for dietary Lysine and Glutamine. Based on the findings of this study, the highest performance was observed in birds fed the diet supplemented with 1.42 lysine and 1% glutamine, but the highest improvement in feed conversion was observed in diet contain 1.14 and 1.42 with 1% and 3% glutamine, respectively. Birds fed 1.42 lysine and 1% glutamine had the highest total body weight gain and feed consumption. The lysine requirements in the diet for Chicken are between 1.14 and 1.42 with glutamine level of 1%.

Dynamics of glutamine synthetase expression in hepatic ischemiareperfusion injury: Implications for therapeutic interventions

BACKGROUND Hepatic ischemia-reperfusion injury(IRI)poses a great challenge in liver surgery and transplantation because of oxidative stress and inflammatory responses.The changes in glutamine synthetase(GS)expression during hepatic IRI remain unclear.AIM To investigate the dynamic expression of GS during hepatic IRI.METHODS Following hepatic ischemia for 1 h and reperfusion,liver tissue samples were collected at 0.5,6,and 24 hours postreperfusion for fixation,embedding,section-ing.Hematoxylin and eosin staining and GS staining were performed.RESULTS GS expression rapidly decreases in hepatocytes around the central vein after IRI,reaching its lowest point at 6 hours postreperfusion,and then gradually recovers.CONCLUSION GS is highly sensitive to IRI,highlighting its potential role as an indicator of liver injury states and a target for therapeutic intervention.

Glutamine supplementation attenuates intestinal apoptosis by inducing heat shock protein 70 in heatstroke rats

BACKGROUND:Heatstroke is the most hazardous heat-related illness and has a high fatality rate.We investigated whether glutamine supplementation could have a protective effect on heatstroke rats.METHODS:Twenty-five 12-week-old male Wistar rats(weight 305±16 g)were randomly divided into a control group(n=5),heatstroke(HS)group(n=10),and heatstroke+glutamine(HSG)group(n=10).Seven days before heat exposure,glutamine(0.4 g/[kg·d])was administered to the rats in the HSG group by gavage every day.Three hours after heat exposure,serum samples were collected to detect white blood cells,coagulation indicators,blood biochemical indicators,and inflammatory cytokines in the rats.The small intestine tissue was stained to analyze pathological structural changes and apoptosis.Finally,immunohistochemistry and Western blotting were used to analyze the expression levels of heat shock protein 70(HSP70).Multiple comparisons were analyzed by using one-way analysis of variance,and the Bonferroni test was conducted for the post hoc comparisons.RESULTS:After heat exposure,the core temperature of the HS group(40.65±0.31°C)was higher than the criterion of heatstroke,whereas the core temperature of the HSG group(39.45±0.14°C)was lower than the criterion.Glutamine supplementation restored the increased white blood cells,coagulation indicators,blood biochemical indicators,and inflammatory cytokines that were induced by heatstroke to normal levels.The intestinal mucosa was injured,and the structure of tight junctions was damaged in the HS group;however,the structure of intestinal mucosal epithelial cells was stable in the HSG group.Glutamine supplementation alleviated intestinal apoptosis and up-regulated HSP70 expression.CONCLUSION:Glutamine supplementation may alleviate intestinal apoptosis by inducing the expression of HSP70 and have a protective effect on heatstroke rats.

Characterization of prognosis and immune infiltration by a novel glutamine metabolism-related model in cutaneous melanoma

Glutamine metabolism(GM)plays an important role in tumor growth and proliferation.Skin cutaneous melanoma(SKCM)is a glutamine-dependent cancer.However,the molecular characteristics and action mechanism of GM on SKCM remain unclear.Therefore,we aimed to explore the effects of GM-related genes on survival,clinicopathological characteristics,and the tumor microenvironment in SKCM.In this study,682 SKCM samples were obtained from the Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)databases.Consensus clustering was used to classify SKCM samples into distinct subtypes based on 41 GM-related genes.Differences in survival,immune infiltration,clinical characteristics,and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathways as well as differentially expressed genes(DEGs)between subgroups were evaluated.A prognostic model was constructed according to prognostic DEGs.Differential analyses in survival,immune infiltration,tumor microenvironment(TME),tumor mutation burden(TMB),stemness,and drug sensitivity between risk groups were conducted.We identified two distinct GM-related subtypes on SKCM and found that GM-related gene alterations were associated with survival probability,clinical features,biological function,and immune infiltration.Then a risk model based on six DEGs(IL18,SEMA6A,PAEP,TNFRSF17,AIM2,and CXCL10)was constructed and validated for predicting overall survival in SKCM patients.The results showed that the risk score was negatively correlated with CD8+T cells,activated CD4+memory T cells,M1 macrophages,andγδT cells.The group with a low-risk score was accompanied by a better survival rate with higher TME scores and lower stemness index.Moreover,the group with high-and low-risk score had a significant difference with the sensitivity of 75 drugs(p<0.001).Overall,distinct subtypes in SKCM patients based on GM-related genes were identified and the risk model was constructed,which might contribute to prognosis prediction,guide clinical therapy,and develop novel therapeutic strategies.

Glutamine addiction and therapeutic strategies in pancreatic cancer

Pancreatic cancer remains one of the most lethal diseases worldwide owing to its late diagnosis,early metastasis,and poor prognosis.Because current therapeutic options are limited,there is an urgent need to investigate novel targeted treatment strategies.Pancreatic cancer faces significant metabolic challenges,principally hypoxia and nutrient deprivation,due to specific microenvironmental constraints,including an extensive desmoplastic stromal reaction.Pancreatic cancer cells have been shown to rewire their metabolism and energy production networks to support rapid survival and proliferation.Increased glucose uptake and glycolytic pathway activity during this process have been extensively described.However,growing evidence suggests that pancreatic cancer cells are glutamine addicted.As a nitrogen source,glutamine directly(or indirectly via glutamate conversion)contributes to many anabolic processes in pancreatic cancer,including amino acids,nucleobases,and hexosamine biosynthesis.It also plays an important role in redox homeostasis,and when converted toα-ketoglutarate,glutamine serves as an energy and anaplerotic carbon source,replenishing the tricarboxylic acid cycle intermediates.The present study aims to provide a comprehensive overview of glutamine metabolic reprogramming in pancreatic cancer,focusing on potential therapeutic approaches targeting glutamine metabolism in pancreatic cancer.

Methods for the Determination of Stable Isotopes of Carbon and Nitrogen Directly in Valine, Proline, Glutamine, and Glutamic Acid

Amino acids are very important compounds for the body and are involved in important functions that keep us healthy. Amino acids are essential components such as valine, proline, glutamine and glutamic acid. They can be synthesized either naturally or artificially. To examine the metabolism and regulate the synthesis process, compounds labeled with nitrogen or carbon isotopes need to be used. These isotopic compounds allow for more extensive research and enable studies that would otherwise be impossible. However, their use is dependent on the availability of simple, efficient methods for isotopic analysis. Currently, the determination of the atomic fraction of carbon and nitrogen isotopes is only possible through their conversion into molecular nitrogen or carbon monoxide or carbon dioxide. This leads to the loss of information about isotopic enrichment in specific centers of the molecule. This article explores a new direct approach to determining the atomic fraction of carbon and nitrogen isotopes in the isotope-modified or identical centers of these compounds. This method eliminates the transfer process and dilution due to nitrogen and carbon impurities. It is now possible to simultaneously determine the atomic fraction of nitrogen and carbon isotopes in the research substance. This method can be applied to amino acids, making it an effective tool for proposing new research methods. Several articles [1] [2] [3] have proposed similar methods for organic compounds and amino acids.

Prognostic model and treatment plan analysis of hepatocellular carcinoma based on genes related to glutamine metabolism

Objective:To identify the prognosis of hepatocellular carcinoma(HCC)and the effect of anti-cancer drug therapy by screening glutamine metabolism-related signature genes because glutamine metabolism plays an important role in tumor development.Methods:We obtained gene expression samples of normal liver tissue and hepatocellular carcinoma from the TCGA database and GEO database,screened for differentially expressed glutamine metabolismrelated genes(GMRGs),constructed a prognostic model by lasso regression and step cox analysis,and assessed the differences in drug sensitivity between high-and low-risk groups.Results:We screened 23 differentially expressed GMRGs by differential analysis,and correlation loop plots and PPI protein interaction networks indicated that these differential genes were strongly correlated.The four most characterized genes(CAD,PPAT,PYCR3,and SLC7A11)were obtained by lasso regression and step cox,and a risk model was constructed and confirmed to have reliable predictive power in the TCGA dataset and GEO dataset.Finally,immunotherapy is better in the high-risk group than in the low-risk group,and chemotherapy and targeted drug therapy are better in the low-risk group than in the high-risk group.Conclusion:In conclusion,we have developed a reliable prognostic risk model characterized by glutamine metabolism-related genes,which may provide a viable basis for the prognosis and Treatment options of HCC patients.

Effects of Neuromuscular Electrical Stimulation in Combination with Glutamine Administration on Skeletal Muscle Atrophy in Colon-26 Tumor-Bearing Mice

The depressed protein synthetic response,a phenomenon termed anabolic resistance,has been shown to be involved in muscle wasting induced by cancer cachexia.Moreover,a positive relationship between the protein synthetic rate and intracellular glutamine(GLN)concentration has been found in skeletal muscles.This study investigated the effects of neuromuscular electrical stimulation(ES)and GLN administration on muscle wasting and GLN metabolism in colon-26(C-26)tumor-bearing mice.CD2F1 mice were divided into 8 groups:control(CNT),CNT+ES,CNT+GLN,CNT+ES+GLN,C-26,C-26+ES,C-26+GLN,C-26+ES+GLN.Cancer cachexia was induced by subcutaneous injection of C-26 cells and developed for four weeks.ES was performed on the left plantar flexor muscles every other day,and GLN(1 g/kg)was administered daily intraperitoneally starting one day after the C-26 injection.Tumor-free body mass and fast-twitch gastrocnemius(Gas)muscle weight were lower in the C-26 group than in the CNT group(-19%and-17%,respectively).Neither ES training nor GLN administration,alone or in combination,ameliorated the loss of Gas muscle weight in the C-26 mice.However,ES training in combination with GLN administration inhibited the increased expression of GLN synthetase(GS)in the C-26 muscles.Thus,it is likely that GLN plays a critical role in muscle protein metabolism and,therefore,can be targeted as a tentative treatment of cancer cachexia.

Comparative Study of Immunological Properties on Glutamine Synthetase Isozymes in Rice Plants

In rice (Oryza sativa L.) roots two glutamine synthetase (GS) isozymes, GSra and GSrb, were identified recently in the author's experiments, but the homology of both GSra and GSrb as well as their localization in the rice roots are unclear. In the present study, the purified GSra and GSrb from rice roots were used to immunize rabbits to obtain the respective antibodies. The immunodiffusion and immunoblotting experiments showed that the antibody against GSra or GSrb was specific for GS and its isozymes. The immunoprecipitation test indicated that the antibody of GSra or GSrb not only recognized its respective antigen, but also well recognized each other's antigen. GSra or GSrb antibody recognized also better cytosolic GS1 of rice leaves, but the recognization for chloroplast GS2 from rice or spinach (Spinacia oleracea Mill.) leaves was weaker. Our results indicate that GSra and GSrb from rice roots are quite similar in antigenicity and are extremely similar proteins and that both GSra and GSrb may also be a form of cytosolic GS just as the cytosolic GS1 of rice leaves.

Iactulose和glutamine对梗阻性黄疸大鼠空肠黏膜的影响

目的:研究梗阻性黄疸时空肠黏膜的变化及lactulose和glutamine对梗阻性黄疸大鼠空肠黏膜的影响．方法:Wistar大鼠84只,随机分为4组．通过手术结扎切断大鼠胆总管得到梗阻性黄疸模型．对梗阻性黄疸大鼠分别经胃灌注lactulose和glutamine药物,比较给药前及给药后5,10 d各组大鼠空肠黏膜绒毛高度变化,同时与未行胆管结扎的假手术对照组进行比较．结果:无论胆总管结扎与否,给药前各组空肠黏膜的绒毛高度无明显差异．胆总管结扎后大鼠空肠黏膜高度减低(5 d:q=4．32,P<0．01;10 d:q=11．03,P<0．01);应用生理盐水组大鼠的空肠黏膜绒毛高度明显低于应用lactulose和glutamine组大鼠的空肠黏膜绒毛高度,且应用glutamine组与胆总管未结扎组相近(5 d:q= 3．62,P>0．05;10 d:q=3．83,P>0．05);而应用lactulose和glutamine的2组大鼠空肠黏膜绒毛高度无明显差异(P>0．05)．结论:结扎大鼠胆总管可导致其空肠黏膜萎缩．经胃肠道应用lactulose或glutamine对胆道梗阻所致的大鼠空肠黏膜萎缩均具有保护作用,且二者对肠黏膜的保护作用无明显差异．

Protective effect of glutamine on intestinal injury and bacterial community in rats exposed to hypobaric hypoxia environment

AIM: To investigate the protective effect of glutamine (Gln) on intestinal injury and the bacterial community in rats exposed to hypobaric hypoxia environment.

Glutamine:aprecursor of glutathione and its effect on liver

AIM To investigate the relationship between alanyl-glutamine (ALA-GLN) and glutathione (GSH) biosynthesis in hepatic protection.METHODS Twenty male Wistar rats were randomly divided into two groups: one receiving standard parenteral nutrition (STD) and the other supplemented with or without ALA-GLN for 7 days. The blood and liver tissue samples were examined after 5-fluorouracil (5-FU) was injected peritoneally.RESULTS The concentration measurements were significantly higher in ALA-GLN group than in STD group in serum GLN (687 μmol/ L±50 μmol/ L vs 505 μmol/ L±39 μmol/ L, P＜0.05), serum GSH (14 μmol/ L±5 μmol/ L vs 7 μmol/ L±3 μmol/ L, P＜0.01) and in liver GSH content (6.9 μmol/ g±2.5 μmol/ g vs 4.4 μmol/ g±1.6 μmol/ g liver tissue, P＜0.05). Rats in ALA-GLN group had lesser elevations in hepatic enzymes after 5-FU administration.CONCLUSION The supplemented nutrition ALA-GLN can protect the liver function through increasing the glutathione biosynthesis and preserving the glutathione stores in hepatic tissue.

Effects of creep feeding and supplemental glutamine or glutamine plus glutamate (Aminogut) on pre- and post-weaning growth performance and intestinal health of piglets

Background： Creep feeding is used to stimulate piglet post-weaning feed consumption.L-Glutamine（GLN） is an important source of fuel for intestinal epithelial cells.The objective of this study was to determine the impact of creep feeding and adding GLN or AminoGut（AG;containing glutamine ＋ glutamate） to pre-and post-weaning diets on pig performance and intestinal health.Litters（N = 120） were allotted to four treatments during 14–21 d of lactation： 1） No creep feed（NC,n = 45）;2） creep fed control diet（CFCD,n = 45）;3） creep fed 1% GLN（CFGLN,n = 15）;4） creep fed.88% AG（CFAG,n = 15）.After weaning,the NC and CFCD groups were sub-divided into three groups（n = 15 each）,receiving either a control nursery diet（NC-CD,CFCD-CD） or a diet supplemented with either GLN（NC-GLN,CFCD-GLN） or with AG（NC-AG,CFCD-AG）.Litters that were creep fed with diets containing GLN or AG also were supplemented with those amino acids in the nursery diets（CFGLN-GLN,CFAG-AG）.Glutamine was added at 1% in all three post-weaning diet phases and AG was added at.88% in phase 1 and 2 and at.66% in phase 3.Results： Feed conversion（feed/gain） showed means among treatment means close to significance（P = 0.056） and Tukey＇s test for pairwise mean comparisons showed that Pigs in the CFGLN-GLN group had the best feed conversion（feed/gain） in the first three-week period post-weaning,exceeding（P = 0.044） controls（CFCD-CD） by 34%.The NC-AG group had（P = 0.02） the greatest feed intake in the last three week of the study,exceeding controls（CFCD-CD） by 12%.CFGLN-GLN,CFCD-GLN and sow reared（SR） pigs had the greatest（P = 0.049） villi height exceeding the CFCD-AG group by 18%,20% and 19% respectively.The CFAG-AG group had the deepest（P = 0.001） crypts among all treatments.CFGLN-GLN,CFCD-GLN and SR groups had the greatest（P = 0.001） number of cells proliferating（PCNA） exceeding those in the NC-CD group by 43%,54% and 63% respectively.Sow reared pigs showed the greatest（P = 0.001） intestinal absorption capacity for xylose and mannitol.Conclusion： Supplementation of creep feed and nursery diets with GLN and/or AminoGut in the first three week improved feed conversion possibly due to improved intestinal health.

Enteral nutrition combined with glutamine promotes recovery after ileal pouch-anal anastomosis in rats

AIM To assess the effect of enteral nutrition(EN) supplemented with glutamine on recovery after ileal pouch-anal anastomosis(IPAA) in rats, to provide an experimental basis for nutritional support in patients with ulcerative colitis(UC) after IPAA. METHODS Male Sprague-Dawley(SD) rats were randomly divided into three groups(n = 8) after IPAA operation using a microsurgical technique. From the third postoperative day, rats in the control group, EN group, and immune nutrition(IN) group were fed standard rat chow, short peptide EN, and short peptide EN combined with glutamine ad libitum, respectively. The rats' general condition was observed throughout the study. Serum levels of total protein(TP), albumin(ALB), prealbumin(PA), and transferrin(TF) were detected on the 30 th postoperative day, using an automatic biochemical analyzer. The ileal pouch mucosa was stained with hematoxylin and eosin(HE), and occludin protein levels were detected by immunohistochemistry.RESULTS The body weight of rats in the EN group(359.20 ± 10.06 g) was significantly higher than that in the control group(344.00 ± 9.66 g)(P < 0.05) and lower than that in the IN group(373.60 ± 9.86 g)(P < 0.05) on the 30 th postoperative day. The levels of serum TP, ALB, PA, and TF in the EN group were significantly higher than those in the control group(P < 0.01 for all) and lower than those in the IN group(P < 0.05 for all). Histopathological score(EN: 0.80 ± 0.37; IN: 0.60 ± 0.40; control group: 2.29 ± 0.18) and expression level of occludin protein(EN: 0.182 ± 0.054; IN: 0.188 ± 0.048; control group: 0.127 ± 0.032) were significantly lower in the control group compared with the EN and IN groups(P < 0.05 for all), but there were no significant differences between the latter two groups(P > 0.05 for all). CONCLUSION EN combined with glutamine may effectively improve nutritional status after IPAA. Our results suggest a benefit of glutamine supplementation in EN for UC patients undergoing IPAA, although human studies are required to confirm this finding.

Influences of Mo on Nitrate Reductase, Glutamine Synthetase and Nitrogen Accumulation and Utilization in Mo-Efficient and Mo-Inefficient Winter Wheat Cultivars

The objective is to study whether the accumulation and utilization of plant N are controlled by Mo status in winter wheat cultivars. Mo-efficient cultivar 97003 （eft） and Mo-inefficient cultivar 97014 （ineff） were grown in severely Mo-deficient acidic soil （Tamm-reagent-extractable Mo 0.112 mg kg^-1） with （＋Mo） and without （-Mo） the application of 0.13 mg kg^-1 Mo. The accumulation and use efficiency of plant total N were significantly higher in ＋Mo than that in -Mo and in eft than that in ineff under Mo deficiency. N use efficiency was remarkably higher in maturity but it was forwarded to jointing stage after Mo supply, thus indicating that Mo supply promoted the N use efficiency besides N uptake and eff was efficient in N uptake and utilization. The overall activity of nitrate reductase （NR, EC 1.6.6.1） was significantly higher in ＋Mo than in -Mo and ratio of ＋Mo/-Mo was even to 14.8 at filleting stage for ineff. Activity of glutamine synthetase （GS, EC 6.3.1.2） was significantly lower in ＋Mo than in -Mo. Concentration of nitrate and glutamate were also significantly lower in ＋Mo than in -Mo, thus provided evidences for enhancing N use efficiency by Mo supply. Activities of NR and GS were significantly higher and concentrations of nitrate and glutamate were significantly lower in eff than ineff under Mo deficiency, thus indicated eff was more efficient in N reduction and utilization. It is therefore concluded that Mo could promote N accumulation and utilization in winter wheat which was directly related to NR and feedback regulated by GS. Higher Mo status also results in higher accumulation and utilization of plant N in eft.

Glutamine synthetase as an early marker for hepatocellular carcinoma based on proteomic analysis of resected smal hepatocel ular carcinomas

BACKGROUND:Hepatocellular carcinoma(HCC)is a highly malignant tumor with a poor prognosis.Because small HCCs possess most of the characteristics of early HCC,we investigated small HCCs to screen potential biomarkers for early diagnosis.METHODS:Proteins were extracted from 10 sets of paired tissue samples from HBV-infected small-HCC patients.The extracted proteins were well resolved by two-dimensional electrophoresis.These HCC-associated proteins were then identified by MALDI-TOF/TOF MS following image analysis.Western blotting and immunohistochemistry were used to assess glutamine synthetase(GS)and phenazine biosynthesislike domain-containing protein(PBLD)expression in liver tissue.Enzyme-linked immunosorbent assays in 152 serum samples(from 49 healthy donors,24 patients with liver cirrhosis,and 79 with HCC)were used to further assess the significance of GS clinically.RESULTS:Fifteen up-regulated and three down-regulated proteins were identified.Western blotting confirmed GS overexpression and decreased PBLD expression in liver tissue.Immunohistochemistry showed that GS was expressed in 70.0%(84/120)of HCCs and 35.8%(43/120)of nontumor tissues;PBLD was expressed in 74.2%(89/120) of nontumor tissues and 40.8%(49/120)of HCCs.The Chi-square test showed significant expression differences between HCCs and adjacent tissues.Consistent with this,serum GS levels in HCC patients were significantly higher than those in liver cirrhosis patients and healthy donors,while the latter two groups were also significantly different.In addition, a diagnostic cutoff value of 2.6 mg/ml was used for GS;it was elevated in 19(76.0%)of 25 HCC patients with AFP≤20 ng/ml and 47(88.7%)of 53 HCC patients with AFP≤200 ng/ml.CONCLUSION:GS and PBLD are abnormally expressed in most HCCs.GS may be a novel serum marker for early HCC, especially for those patients with low AFP levels(≤200 ng/ml).

Antioxidant properties of glutamine and its role in VEGF-Akt pathways in portal hypertension gastropathy

AIM: To investigate the effects of glutamine on oxidative/nitrosative stress and the vascular endothelial growth factor (VEGF)-Akt-endothelial nitric oxide synthase (eNOS) signaling pathway in an experimental model of portal hypertension induced by partial portal vein ligation (PPVL). METHODS: Portal hypertension was induced by PPVL. The PPVL model consists of a partial obstruction of the portal vein, performed using a 20 G blunt needle as a guide, which is gently removed after the procedure. PPVL model was performed for 14 d beginning treatment with glutamine on the seventh day. On the fifteenth day, the mesenteric vein pressure was checked and the stomach was removed to test immunoreactivity and oxidative stress markers. We evaluated the expression and the immunoreactivity of proteins involved in the VEGF-Akt-eNOS pathway by Western blotting and immunohistochemical analysis. Oxidative stress was measured by quantification of the cytosolic concentration of thiobarbituric acid reactive substances (TBARS) as well as the levels of total glutathione (GSH), superoxide dismutase (SOD) activity, nitric oxide (NO) production and nitrotyrosine immunoreactivity. RESULTS: All data are presented as the mean ± SE. The production of TBARS and NO was significantly increased in PPVL animals. A reduction of SOD activity was detected in PPVL + G group. In the immunohistochemical analyses of nitrotyrosine, Akt and eNOS, the PPVL group exhibited significant increases, whereas decreases were observed in the PPVL + G group, but no difference in VEGF was detected between these groups. Western blotting analysis detected increased expression of phosphatidylinositol-3-kinase (PI3K), P-Akt and eNOS in the PPVL group compared with the PPVL + G group, which was not observed for the expression of VEGF when comparing these groups. Glutamine administration markedly alleviated oxidative/nitrosative stress, normalized SOD activity, increased levels of total GSH and blocked NO overproduction as well as the formation of peroxynitrite. CONCLUSION: Glutamine treatment demonstrated to reduce oxidative damage but does not reduce angiogenesis induced by PH in gastric tissue, demonstrating a beneficial role for the PI3K-Akt-eNOS pathway.