Cloning and Expression of Rat Liver S-Adenosylmethionine Synthetase

The S-adenosylmethionine synthetase（SAM synthetase） is responsible for in vivo synthesis of S-adenosylmethionine（SAM）, which is a kind of biologically active molecules distributed in all body tissues and fluids and involved in a number of biochemical reactions. In this study, a cDNA containing the coding sequence for rat liver SAM synthetase was cloned into the prokaryotic expression vector pQE30 and expressed in E. coli M15. A major band corresponding to a protein of 48 kDa was detected on SDS-PAGE. The protein was distributed in both the soluble fraction and the insoluble fraction. In soluble fractions the protein was fully active.

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.

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.

Effect of Low Temperature Stress on Sweet Potato S-Adenosyl Methionine Synthetase Gene Expression

[Objective] The mRNA expression level changes of S-adenosylmethionine synthetase （SAMS） under low temperature stress was studied. [Method] Total RNA were extracted from leaves, stem and earthnut of sweet potato 0,12,24,48 and 72 h after low temperature treatement, mRNA expression level was analyzed by reverse expression and Real-time PCR technique. [Result] The expression quality of the gene extracted from leaves, stem and earthnut increased and the expression quality reached the peak point 24,72 and 72 h after low temperature treatment respectively. The expression change of earthnut was the biggest. [Conclusion] Low temperature was good for increasing mRNA expression of relevart genes.

Long-chain acyl-CoA synthetase in fatty acid metabolism involved in liver and other diseases:An update

Long-chain acyl-Co A synthetase(ACSL) family members include five different ACSL isoforms, each encoded by a separate gene and have multiple spliced variants. ACSLs on endoplasmic reticulum and mitochondrial outer membrance catalyze fatty acids with chain lengths from 12 to 20 carbon atoms to form acyl-Co As, which are lipid metabolic intermediates and involved in fatty acid metabolism, membrane modifications and various physiological processes. Gain- or lossof-function studies have shown that the expression of individual ACSL isoforms can alter the distribution and amount of intracellular fatty acids. Changes in the types and amounts of fatty acids, in turn, can alter the expression of intracellular ACSLs. ACSL family members affect not only the proliferation of normal cells, but the proliferation of malignant tumor cells. They also regulate cell apoptosis through different signaling pathways and molecular mechanisms. ACSL members have individual functions in fatty acid metabolism in different types of cells depending on substrate preferences, subcellular location and tissue specificity, thus contributing to liver diseases and metabolic diseases, such as fatty liver disease, obesity, atherosclerosis and diabetes. They are also linked to neurological disorders and other diseases. However, the mechanisms are unclear. This review addresses new findings in the classification and properties of ACSLs and the fatty acid metabolismassociated effects of ACSLs in diseases.

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).

Novel methionyl-tRNA synthetase gene variants/phenotypes in interstitial lung and liver disease: A case report and review of literature

Interstitial lung and liver disease(ILLD) is caused by biallelic mutations in the methionyl-tRNA synthetase(MARS) gene. To date, no genetic changes other than missense variants were reported in the literature. Here, we report a five-month old female infant with typical ILLD(failure to thrive, developmental delay, jaundice, diffuse interstitial lung disease, hepatomegaly with severe steatosis, anemia, and thrombocytosis) showing novel phenotypes such as kidney stones, acetabular dysplasia, prolonged fever, and extreme leukocytosis. Whole exome sequencing revealed a novel truncating variant(c.2158 C>T/p.Gln720 Stop) together with a novel tri-nucleotide insertion(c.893_894 insTCG that caused the insertion of an arginine at amino acid position 299) in the MARS gene.

Modulating effects of acyl-CoA synthetase 5-derived mitochondrial Wnt2B palmitoylation on intestinal Wnt activity

AIM: To investigate the role of acyl-CoA synthetase 5 (ACSL5) activity in Wnt signaling in intestinal surface epithelia.

Changes of Levels of Glutamine Synthetase Isoforms in Roots and Leaves in Responseto Nitrogen Fertilizer Application at Different Growth Stages in Irrigated Rice

Nitrogen is a key element to control the growth and yield of crops. Fertilizer urea nitrogen (N) 60,45, and 30 kg/hm2 was applied at three different stages, midtillering, panicle initiation, and flowering, of the growth and development of rice plants, respectively. At both midtillering and panicle initiation, the total activity of glutamine synthetase (GS) in rice roots and leaves was incrased remarkably as a result of a large amount of ammonia absorbed by roots. Native-PAGE and activity staining showed that the increase of total activity in rice roots and leaves was due to the synthesis of GSrb in roots and GS2 in leaves and that the activity of GSra in roots and GS1 in leaves remained constant. The results showed that the assimilation of external nitrogen was carried out by GSrb but not GSra in rice roots and that the activitry of GS2 was induced also by the external nitrogen, and that GSrb played main role in meeting the needs of the rapid tillering for nitrogen. At flowering, the activity of GS in rice roots and leaves did not change almost after topdressing. These rssults suggest that the change of GS activity in rice roots may use as a measure of the utilization efficiency of the fertilizer.

Intestinal acyl-CoA synthetase 5: Activation of long chain fatty acids and behind

The intestinal mucosa is characterized by a high complexity in terms of structure and functions and allows for a controlled demarcation towards the gut lumen.On the one hand it is responsible for pulping and selective absorption of alimentary substances ensuring the immunological tolerance,on the other hand it prevents the penetration of micro-organisms as well as bacterial outgrowth.The continuous regeneration of surface epithelia along the crypt-villus-axis in the small intestine is crucial to assuring these various functions.The core phenomena of intestinal epithelia regeneration comprise cell proliferation,migration,differentiation,and apoptosis.These partly contrarily oriented processes are molecularly balanced through numerous interacting signaling pathways like Wnt/β-catenin,Notch and Hedgehog,and regulated by various modifying factors.One of these modifiers is acyl-CoA synthetase 5(ACSL5).It plays a key role in de novo lipid synthesis,fatty acid degradation and membrane modifications,and regulates several intestinal processes,primarily through different variants of protein lipidation,e.g.,palmitoylation.ACSL5 was shown to interact with proapoptotic molecules,and besides seems to inhibit proliferation along the crypt-villus-axis.Because of its proapoptotic and antiproliferative characteristics it could be of significant relevance for intestinal homeostasis,cellular disorder and tumor development.

Effect of Nitrate on Activities and Transcript Levels of Nitrate Reductase and Glutamine Synthetase in Rice

Real-time polymerase chain reaction analysis was used to compare the effect of NO3^- on the activities of nitrate reductase （NR） and glutamine synthetase （GS）, and the transcript levels of two NR genes, OsNial and OsNia2, two cytosolic GS1 genes, OsGln1;1 and OsGln1;2, and one plastid GS2 gene OsGln2, in two rice （Oryza sativa L.） cultivars Nanguang （NG） and Yunjing （Y J）. Both cultivars achieved greater biomass and higher total N concentration when grown in a mixed N supply than in sole NH4^＋ nutrition. Supply of NO3^- increased NR activity in both leaves and roots. Expression of both NR genes was also substantially enhanced and transcript levels of OsNia2 were significantly higher than those of OsNial. NO3 also caused an increase in GS activity, but had a complex effect on the expression of the three GS genes. In roots, the OsGln1;1 transcript increased, but OsGln1;2 decreased. In leaves, NO3^- had no effect on the GS1 expression, but the transcript for OsGln2 increased both in the leaves and roots of rice with a mixed supply of N. These results suggested that the increase in GS activity might be a result of the complicated regulation of the various GS genes. In addition, the NO3-induced increase of biomass, NR activity, GS activity, and the transcript levels of NR and GS genes were proportionally higher in NG than in Y J, indicating a stronger response of NG to NO3^- nutrition than YJ.

Changes in Activities of Glutamine Synthetase during Grain Filling and Their Relation to Rice Quality

Four japonica rice varieties differed in cooking and eating qualities were used in a pot experiment to study the relationship between the activities of glutamine synthetase during grain filling and rice quality. The activities of glutamine synthetase gradually increased and then declined as a single peak curve in the course of grain filling. The 15th day after heading was a turning point, before which the enzymatic activities in the inferior rice varieties with high protein content were higher than those in the superior rice varietie with low protein content, and after which it was converse. The activity of glutamine synthetase in grain was correlated with the taste meter value, peak viscosity and breakdown negatively at the early stage of grain filling whereas positively at the middle and late stages. Moreover, it was correlated with the protein content of rice grain and setback positively at the early stage and negatively at the middle and late stages. The correlation degree varied with the course of grain filling. From 15 days to 20 days after heading was a critical stage, in which the direction of correlation between the activity of glutamine synthetase and taste meter value and RVA properties of rice changed.

Effects of Ganoderma lucidum spore powder on astrocyte expression and glutamine synthetase activity in the hippocampal region of epileptic rats

BACKGROUND： Recent studies have demonstrated that astrocyte dysfunction plays a central role in inhibiting epileptic seizures and that regulation of astrocyte function may be a new target for treatment of epilepsy. OBJECTIVE： To observe the effects of Ganoderma lucidum spore powder （GLSP） on astrocyte morphology and glutamine synthetase （COS） activity in the hippocampal region of epileptic rats. DESIGN, TIME AND SETTING： A randomized, controlled animal experiment was performed at the Function Laboratory, College of Basic Medicine, Jiamusi University between October and December 2006. MATERIALS： A total of 30 Sprague Dawley （SD） rats were randomized to three groups （n = 10）： control, model, and GLSP. GLSP was sourced from Jiamusi Wild Ganoderma Lucidum Planting Base and prepared to 30 g/L with physiological saline before use. Pentylenetetrazol （PTZ） （10 g/L） was provided by Sigma Company, USA. METHODS： The control group received intraperitoneal （i.p.） and intragastric （i.g.） physiological saline. Following epilepsy induction by i.p. administration of PTZ （35 mg/kg）, rats from the mode/and GLSP groups were ig injected with physiological saline and GLSP （300 mg/kg）, respectively. Each compound was administered once per day, for a total of 28 successive days. Epileptic seizure convulsions were graded 0-5. A higher grade indicated more severe epilepsy. Only those rats showing stage 2 or higher convulsions at least 5 times successively were included in further experiments. MAIN OUTCOME MEASURES： Immediately alter injection, seizure activity was monitored for 30 minutes to determine the latent period and seizure duration; simultaneously, astrocyte numbers and GS activity in the hippocampal region of rats with epilepsy were detected by immunohistochemistry. RESULTS： All 30 rats were included in the final analysis. On day 28, following PTZ administration epileptic seizures were not found in the control group. In the GLSP group, rats exhibited rhythmic head nodding or facial spasms, and the latent period was significantly longer than that of the model group （P 〈 0.05）. In the model group, the majority of rats exhibited myoclonus or generalized convulsions, and epileptic seizure duration was slightly （but not significantly） longer than that in the GLSP group （P 〉 0.05）. Within 2-3 minutes of PTZ injection, the model group exhibited grade 4 or 5 epileptic seizures, and the GLSP group mostly showed grade 2 or 3, and occasionally grade 4 or 5, epileptic seizures. All rats recovered within 30 minutes. The model group exhibited significantly increased astrocytes （P 〈 0.05）, with thicker cellular processes and a higher number of cellular processes, and significantly decreased GS activity （P 〈 0.05） tban the control group. The astrocyte count was significantly decreased but GS activity was significantly increased in the GLSP group when compared with the model group （P 〈 0.05）; however, astrocyte appearance was similar in both groups （P 〈 0.05）. CONCLUSION： GLSP can effectively inhibit astrocyte numbers and elevate GS activity in the hippocampal region of rats with epilepsy, thereby reducing epileptic seizures.

Expression of Arginyl-tRNA Synthetase in Rats with Focal Cerebral Ischemia

Aminoacyl-tRNA syntheses （AARS） can catalyze the adenosine triphosphate （ATP）-dependent acylation of their cognate tRNA（s） with a specific amino acid. They can be seen as an index to reflect the energy metabolic rate of ischemic brain cells in ischemic penumbra. This study ex- amined the relationship between arginyl-tRNA synthetase （ArgRS）, one of the AARS, and cerebral ischemia in rats. The model of middle cerebral artery occlusion （MCAO） was established in rats. The expression levels of ArgRS protein and mRNA were detected in rat brain tissues at different time points following MCAO by Western blotting and RT-PCR, respectively. The results showed that the MCAO model was successfully established. Western blotting and RT-PCR analysis revealed that the ArgRS protein and mRNA were expressed in brain cells in both ischemic and normal penumbra tissues. The expression levels of ArgRS protein and mRNA peaked at 6 h after MCAO and decreased gradually. At 24 h, the expression levels of ArgRs protein and mRNA in ischemic penumbral tissues were lower than those in normal tissues. The expression levels of ArgRS mRNA and protein in ischemic penumbra var- ied with ischemic time, suggesting that the energy metabolism of brain cells in penumbra changed dy- namically after ischemia to ensure the endogenous self-protection of the body. The brain oxygen supply should be improved as soon as possible, especially within 6-12 h after ischemia, so as to meet the de- mand for energy metabolism in ischemic penumbra and make sure the cell structure remains stable.

Ischemic Preconditioning Inhibits Over-expression of Arginyl-tRNA Synthetase Gene Rars in Ischemia-injured Neurons

The expression changes of Rars gene in ischemia-injured neurons were investigated by detecting its translational product arginyl-t RNA synthetase（Arg RS）, and the inhibitory effects of ischemic preconditioning（IPC） on Rars gene were explored. Both IPC model and prolonged ischemia（PI） model were established by using the classic oxygen glucose deprivation（OGD） method. The primary cultured neurons were assigned into the following groups： the experimental group（IPC＋PI group）, undergoing PI after a short period of IPC; the conditional control group（PI control group）, subjected to PI without IPC; blank control group, the normally cultured neurons. The Rars transcriptional activities and Arg RS expression levels were measured at different time points after re-oxygenation（3 h/6 h/12 h/24 h）. Data were collected and statistically analyzed. Compared to the blank control group, the Rars activities and Arg RS levels were significantly increased in PI control group, peaking at the time point of 6 h after re-oxygenation. Rars activities and Arg RS levels were significantly lower in the experimental group than in the PI control group at different time points after re-oxygenation. PI insult can induce an escalating activity of Rars and lead to Arg RS over-expression in primary cultured neurons. IPC can inhibit the increased Rars activity and down-regulate Arg RS expression of ischemia-insulted neurons. This mechanism may confer ischemic tolerance on neurons.

THE IMMUNOHISTOCHEMISTRY AND IN SITU cDNA-mRNA HYBRIDIZATION OF CARBAMYL PHOSPHATE SYNTHETASE I IN ENZYME-ALTERED LIVER CELLS DURING CARCINOGENESIS

The changes of carbamyl phosphate synthetase I(CPS 1)in diethylnitrosamine-(DEN)-inducedenzyme-altered liver cells were studied by means of immunohistochemical(PAP)and in situcDNA-mRNA hybridization methods.The experimental rats were treated with DEN,2-acetylaminofluorene(2-AAF)and 2/3 hepatectomy according to Solt-Farber’s protocol andwere further promoted by oral daily administration of 0.05% phenobarbital in drinking water.The results showed that the average number of lesions showing abnormal expression of CPS1 was relatively constant over the course of the experiment(8 months),while the numberof normally expressing lesions gradually decreased.The former lesions were also largerin volume than the latter ones.We conclude that in DEN-initiated lesions the abnormallyexpressed CPS 1 lesions may grow continuously,thus leading to the formation of largernodules.We also suspect that some of these lesions have increased tendencies to developinto tumors.

S-adenosylmethionine in treatment of cholestasis after total parenteral nutrition: laboratory investigation and clinical application

Objective: To observe the effects of S-adenosylmethio-nine (SAMe) in the treatment of cholestasis after totalparenteral nutrition (TPN).Methods: Thirty SD rats were randomly divided intocontrol group, hypercalorie group, hypercalorie+SAMegroup, sepsis group and sepsis+SAMe group to com-pare their states of cholestasis. Sixteen patients re-ceived SAMe because of cholestasis after prolongedTPN, and the therapeutic efficacy was observed.Results: Bile flow was obviously decreased and theserum levels of total bile acid and gamma-glutamyltranspeptidase(γ-GT) were markedly increased in thehypercalorie and sepsis groups. Meanwhile, hepatocytefatty degeneration, dilatation of cholangioles, and bilesludge could be seen microscopically. SAMe adminis-tration in the hypercalorie+SAMe and sepsis+SAMegroups could increase the bile flow, decrease theserum levels of total bile acid and γ-GT, reduce thepathological damage to the liver, and clear the bilesludge in the cholangioles. Cholestasis and abnormalliver function were the main manifestations of the 16patients before SAMe administration. After SAMe treat-ment for 3 weeks, serum levels of total bilirubin, al-kaline phosphatase(AKP), γ-GT, alanine aminotrans-ferase(ALT), and aspartate aminotransferase(AST)were obviously decreased, and normalized in the 4thweek.Conclusion: SAMe could prevent and treat cholestasiswithout discontinuation of TPN.

Ammonia metabolism capacity of HepG2 cells with high expression of human glutamine synthetase

BACKGROUND: Currently, one of the tough problems for the application of bioartificial liver (BAL) is the shortage of suitable hepatocytes. There are reports on different types of BAL assistance developed with porcine hepatocytes and HepG2 C3A cells, but their defects are obvious. In recent years, some studies focus more on liver cells with features of human origin and improved detoxification. In this study, a hepatocyte line with high expression of human glutamine synthetase (hGS) was raised and its capacity for ammonia metabolism was investigated. METHODS: hGS cDNA and alpha-fetoprotein transcription regulatory element (AFP-TRE) were cloned with the designed primers. The eukaryotic expression vectors, pLNChGS and pLNAFhGS, were constructed and transfected into PA317 cells. Recombinant retroviruses (Retro-hGS and Retro-AFhGS) were produced and then infected into HepG2 cells. G418-resistant cell clones, HepG2/pLNChGS and HepG2/pLNAFhGS, were selected and amplified. Then hGS mRNA was measured by semi-quantitative RT-PCR; hGS enzymatic activity and ammonia metabolism analysis in different concentration of NH(4)(+) were detected with a quantitative biochemistry kit. The cell proliferation was also detected by MTT chromatometry. RESULTS: The expression of hGS mRNA in HepG2/pLNChGS cells (8.306+/-0.336) and HepG2/pLNAFhGS cells (21.358+/-1.716) was much stronger than in control cells (P<0.05), and that in HepG2/pLNAFhGS cells was markedly stronger than in HepG2/pLNChGS cells (P<0.05). The hGS enzymatic activities of HepG2/pLNChGS cells (3.279+/-0.328 U/mg prot) and HepG2/pLNAFhGS cells (4.557+/-0.253 U/mg prot) were higher than those of control cells (P<0.05), and those of HepG2/ pLNAFhGS cells were also higher than the activities of HepG2/pLNChGS cells (P<0.05). In addition, the effect of hGS introduction on HepG2 cell proliferation was not significant. The amount of glutamine synthesis in HepG2/pLNChGS or HepG2/pLNAFhGS cells in three different concentrations of NH(4)(+) was higher than in the two control cells (P<0.05). The amount of glutamine synthesis and cell proliferation in the higher concentrations of NH(4)(+) (5 or 10 mmol/L) in HepG2/pLNAFhGS cells increased more than those in HepG2/pLNChGS cells (P<0.05). NH(4)(+) at a high concentration (10 mmol/L) was toxic to HepG2 and HepG2/pLNCX cells, but less toxic to HepG2/pLNChGS and HepG2/pLNAFhGS cells. CONCLUSION: The constructed hepatocytes (HepG2 cells) with specific high-expression of hGS have a powerful ability to degrade ammonia in vitro, and provide necessary experimental data for the selection of biomaterials in BAL.

Effect of Exogenous Ammonium on GlutamineSynthetase, Glutamate Synthase, and Glutamate Dehydrogenase in the Root of Rice Seedling

Root biomass of rice seedlings was increased at lower concentration of exogenous NH 4 + , but it was decreased at higher concentration of exogenous NH 4 + . The level of free NH 4 + in the roots was accumulated gradually with the increase of NH 4 + concentration in the nutrient solution. The content of the soluble proteins was essentially constant at higher NH 4 + . The activities of glutamine synthetase (GS), NADH-dependent glutamate synthase (NADH-GOGAT), and NADH-dependent glutamate dehydrogenase (NADH-GDH) were risen with exogenous NH 4 + concentration at the lower NH 4 + concentration range. But the activities of GS and NADH-GOGAT were declined, and the level of NADH-GDH activity was kept constant under higher NH 4 + concentration. The GS/GDH ratio suggested that NH 4 + was assimilated by GS-GOGAT cycle under lower NH 4 + concentration, but NADH-GDH was more important for NH 4 + assimilation and detoxifying NH 4 + to the tissue cells at the higher NH 4 + level. According to the growth and the activity changes of these ammonium-assimilating enzymes of rice seedling roots, 10. 0 μg/mL NH 4 + -N in nutrient solution was more suitable to the rice growth.