Systematic analysis and functional verification of citrus glutathione S-transferases reveals that CsGSTF1 and CsGSTU18contribute negatively to citrus bacterial canker

Citrus bacterial canker(CBC) is resulted from Xanthomonas citri subsp. citri(Xcc) infection and poses a significant threat to citrus production.Glutathione S-transferases(GSTs) are critical in maintaining redox homeostasis in plants, especially in relation to abiotic and biotic stress responses. However, the function of GSTs in resisting CBC remains unclear. Here, citrus glutathione S-transferases were investigated applying a genome-wide approach. In total, 69 CsGSTs belonging to seven classes were identified, and the phylogeny, chromosomal distribution, gene structures and conserved motifs were analyzed. Several CsGSTs responded to Xcc infection, as observed in the upregulation of CsGSTF1 and CsGSTU18 in the CBC-sensitive ‘Wanjincheng' variety but not in the resistant ‘Kumquat' variety. CsGSTF1 and CsGSTU18 were localized at the cytoplasm. Transient overexpression of CsGSTF1 and CsGSTU18 mediated reactive oxygen species(ROS) scavenging, whereas the virus-induced gene silencing(VIGS) of CsGSTF1 and CsGSTU18 caused strong CBC resistance and ROS burst. The present study investigated the characterization of citrus GST gene family, and discovered that CsGSTF1 and CsGSTU18 negatively contributed to CBC through modulating ROS homeostasis. These findings emphasize the significance of GSTs in infection resistance in plants.

Glutathione S-transferase(GST) gene expression profiles in two marine bivalves exposed to BDE-47 and their potential molecular mechanisms

Glutathione S-transferases （GSTs） are phase II enzymes that facilitate the detoxification of xenobioties and play important roles in antioxidant defense. We investigated the expression patterns of seven Venerupis philippinarum GSTs （VpGSTs） and four Mytilus galloprovincialis GSTs （MgGSTs） following exposure to BDE-47. Differential expressions of the seven VpGSTs and four MgGSTs transcripts were observed, with differences between the hepatopancreas and gills. Among these GSTs, the sigma classes （VpGSTS1, VpGSTS2, VpGSTS3, MgGST1, and MgGST3） were highly expressed in response to BDE-47 exposure, demonstrating their potential as molecular biomarkers for environmental biomonitoring studies. We obtained the three-dimensional crystal structures of VpGSTs and MgGSTs by homologous modeling. A model to elucidate the binding interactions between the ligands and receptors was defined by molecular docking, Hydrophobic and n were the most often observed interactions between BDE-47 and the GSTs.

The potential of carcinoembryonic antigen,p53,Ki-67 and glutathion Stransferase-π as clinico-histopathological markers for colorectal cancer

Objective： Colorectal cancer is one of the major contributors to cancer death worldwide. Lack of reliable colorectal cancer markers has hampered the management of these cancer patients. Our main purpose was to study the correlation between histopathological variables of colorectal adenocarcinomas and identify histopathological markers that are of prognostic value in patients with colorectal cancer. Methods： In the present study, we examined the expression of carcinoembryonic antigen （CEA）, p53, Ki-67 and glutathion Stransferase （GST） -n by using immunohistochemical staining methods in 126 colorectal carcinoma patients and evaluated the lymph node metastasis status in these patients by histopathological examination. Results： The positive rates of CEA, p53, Ki-67 and GST-π expression in the colorectal cancer tissue specimens examined were 95.23%, 55.56%, 53.38% and 82.30%, respectively. Expression of p53 and Ki-67 was significantly correlated with the Dukes stages of the tumor, with higher levels of these proteins in Dukes＇ C and D tumors than those in Dukes＇ A and B tumors. Furthermore, the expression of p53, GST-π and Ki-67 correlated with prognosis of patients with colorectal cancer. Additionally, the expression of p53 in colorectal cancer was closely related to the expression of Ki-67 and the expression of GST-π was directly correlated with that of p53. Conclusion： The expression of CEA, p53, Ki-67 and GST-π was correlated with various clinical features of patients with colorectal cancer. The combined use of these histopathological markers appeared to be a promising tool in predicting the prognosis of patients with this type of cancer.

Polymorphisms of the oxidant enzymes glutathione S-transferase and glutathione reductase and their association with resistance of Plasmodium falciparum isolates to antimalarial drugs

Objective:To investigate the association between amplification of the two regulatory genes controlling glutathione(GSH) levels,glutathione reductase(PfGR) and glutathione S-transferase (PfGST) genes and sensitivity of Plasmodium falciparum(P.falciparum) isolates collected from different malaria endemic areas of Thailand to standard antimalarial drugs.Methods:A total of 70 P.falciparum isolates were collected from endemic areas of multi-drug resistance (Tak,Chantaburi and Ranong Provinces) during the year 2008-2009.The in vitro assessment of antimalarial activity of P.falciparum clones(K1- and Dd2 chloroquine resistant and 3D7- chloroquine sensitive) and isolates to chloroquine,quinine,mefloquine and arteusnate was performed based on SYBR Green modified assay.Results:68(97.14%),11(15.71%) and 28(40%) isolates respectively were classified as chloroquine-,quinine- and mefloquine-resistant isolates. With this limited number of P.falciparum isolates included in the analysis,no significant association between amplification of PfGST gene and sensitivity of the parasite to chloroquine, quinine,mefloquine and quinine was found.Based on PCR analysis,Dd2,Kl and 3D7 clones all contained only one copy of the PfGST gene.All isolates(70) also carried only one copy number of PfGST gene.There appears to be an association between amplification of PfGR gene and chloroquine resistance.The 3D7 and Dd2 clones were found to carry only one PfGR gene copy, whereas the K1 clone carried two gene copies.Conclusions:Chloroquine resistance is likely to be a consequence of multi-factors and enzymes in the GSH system may be partly involved. Larger number of parasite isolates are required to increase power of the hypothesis testing in order to confirm the involvement of both genes as well as other genes implicated in glutathione metabolism in conferring chloroquine resistance.

Cloning, characterization and expression analysis of a microsomal glutathione S-transferase gene from the seagrass Zostera marina

The response of glutathione S-transferase(GST)in Zostera marina to temperature variation was analyzed at molecular level by cloning the microsomal GST gene and texting the microsomal GST expression regularity under different temperature.Specific speaking,express ZmGST in Escherichia coli,then purify the recombinant protein and make the thermal stability analysis.Therefore,the experiments were carried out to provide a theoretical basis for the further elaboration to the population degradation mechanisms of Z.marina.In conclusion,the thermostability and the response of ZmGST gene to temperature changes can determine its temperature tolerance range,and affect its resilience in turn.

CircRNA_0084927 promotes colorectal cancer progression by regulating miRNA-20b-3p/glutathione S-transferase mu 5 axis

BACKGROUND Colorectal cancer(CRC)is the third most common cancer and the second most common cause of cancer-related death worldwide.The 5-year survival rate of patients with early-stage CRC could reach 90%,but it is very low in patients with advanced-stage CRC.Recent studies have shown that circular RNAs play important roles in regulating the migration and invasion of CRC cells.AIM To elucidate the role of circRNA_0084927(circ_0084927)in the migration and invasion of CRC cells and its underlying mechanism.METHODS Clinical tissue samples and cells were collected,and the expression of circ_0084927 was detected by quantitative polymerase chain reaction(qPCR).The diagnostic performance of circ_0084927 was assessed by receiver operating characteristic curve analysis.The role of circ_0084927 in CRC cell proliferation,migration,and invasion was determined using cell counting kit-8 assay,wound healing assay,and transwell assay,respectively.The regulatory relationship among circ_0084927,miRNA-20b-3p(miR-20b-3p),and glutathione S-transferase mu 5(GSTM5)was identified using databases,luciferase reporter assay,qPCR,and Western blot analysis.AKT-mTOR signaling was also verified after circ_0084927 knockdown or miR-20b-3p mimic treatment.RESULTS The expression of circ_0084927 was significantly increased in CRC tissues and cells,and it was higher in advanced-stage CRC compared with early-stage CRC.The area under the curve(AUC)of circ_0084927 was 0.806[95%confidence interval(CI):0.683-0.896].In addition,the AUC was 0.874(95%CI:0.738-0.956)in patients with advanced-stage CRC and 0.713(95%CI:0.555-0.840)in those with early-stage CRC.Knockdown of circ_0084927 inhibited the migration and invasion of HCT116 cells.Moreover,circ_0084927 was found to act as a sponge of miR-20b-3p.MiR-20b-3p activation reduced the circ_0084927 level,whereas miR-20b-3p inhibition increased the circ_0084927 level.But the effect was not found after circ_0084927 mutation.In addition,miR-20b-3p expression in CRC patients was also reduced and negatively correlated with circ_0084927 expression.The function of circ_0084927 in HCT116 cells with circ_0084927 knockdown was rescued by miR-20b-3p.Moreover,GSTM5 expression was significantly decreased after overexpressing miR-20b-3p or inhibiting circ_0084927,but its expression was rescued when circ_0084927 and miR-20b-3p were both inhibited.Finally,AKTmTOR signaling was markedly regulated by circ_0084927 and miR-20b-3p.CONCLUSION The expression of circ_0084927 is significantly increased in CRC and higher in advanced-stage CRC than in early-stage CRC.Moreover,circ_0084927 potentially regulates CRC cell migration and invasion via the miR-20b-3p/GSTM5/AKT/mTOR pathway.

Comparative Studies of Substrate and Inhibitor Specificity of Glutathione S-Transferases in Six Tissues of Oxya chinensis(Thunberg)(Orthoptera:Acrididae)

Specific activity, substrate specificity, and kinetic parameters （Km and Vmax） of glutathione S-transferases （GSTs） towards three substrates, 1-chloro-2,4-dinitrobenzene （CDNB）, 1,2-dichloro-4-nitrobenzene （DCNB）, and p-nitrobenzene chloride （pNBC） were investigated in six tissues （foregut, midgut, hindgut, fat body, hemolymph, and muscle） of Oxya chinensis. In addition, the inhibition in vitro （ethacrynic acid, and Cibacron Blue 3GA） of Oxya chinensis in the six tissues was also investigated. Glutathione S-transferase activity was detected in all the six tissues examined. The rank order of GST activities towards CDNB was fat body 〉 midgut 〉 hindgut 〉 muscle 〉 foregut 〉 hemolymph both in females and males. Glutathione S-transferase activities in the fat body in females and males were 1.3- to 10.4-fold and 1.1- to 10.0- fold higher than those in the other tissues. The rank order of GST activities towards the other substrates changed slightly. From these results, it was inferred that GSTs in the fat body and midgut played important roles in detoxifying xenobiotics including insecticides and plant allelochemicals in O. chinensis. In the three substrates examined, CDNB seemed to be the best substrate, followed by pNBC and DCNB. The kinetic parameters of GSTs were different among the six tissues. This suggested that GSTs in different tissues have various affinities and catalytic efficiency to substrates. In vitro inhibition study showed that the median inhibition concentration （IC50） values of the two inhibitors to GSTs from the six tissues were different. The results suggested that the two inhibitors have different inhibition potency to GSTs from the different tissues. The observed changes in kinetic parameters and inhibition in vitro among the six tissues of the insect might suggest that the number and structure of isoenzymes and their rate of expression varied for the different tissues.

Antioxidant Effect of Selenium-containing Glutathione S-Transferase in Rat Cardiomyocytes

As one of the most important antioxidant enzymes, glutathione peroxidase（GPX） protects cells and tissues from oxidative damage, and plays an important role in cardiovascular and cerebrovascular injuries induced by oxida- tive stress. The antioxidant effect of selenium-containing glutathione S-transferase（Se-GST）, a mimic of GPX was investigated on rat cardiomyocytes. To explore the protection function of Se-GST in hydrogen peroxide（H202） chal- lenged rat cardiomyocytes, we examined malondialdehyde（MDA）, lactate dehydrogenase（LDH）, superoxide dismu- tase（SOD） and cell apoptosis. The results demonstrate exposure of rat cardiomyocytes to H202 for 6 and 12 h induced the significant increases of MDA, LDH and apoptosis rate of cardiomyocytes, but pretreatment of rat cardiomyocytes with Se-GST at 0.0005 or 0.001 unit/mL prevents oxidative stress induced by H202 with the decreases of cell apopto- sis. All the results hint Se-GST has antioxidant activity for oxidative stress challenged rat cardiomyocvtes.

Expression and Characterization of a Sigma-Class Glutathione S-transferase of the Oriental Migratory Locust, Locusta migratoria manilensis (Meyen)

A cDNA encoding a sigma-class glutathione S-transferase of the locust, Locusta migratoria manilensis （LmGSTs1）, was cloned by reverse transcriptase-polymerase chain reaction. The 830 bp-long cDNA encoded a 615 bp open reading frame （204 amino acid polypeptide）, which exhibited the structural motif and domain organization characteristic of GST sigma-class. It revealed 59, 57, 57, and 56% identities to sigma-class GSTs from Blattella germanica, Gryllotalpa orientalis, Nasonia vitripennis, and Pediculus humanus corporis, respectively. A recombinant protein （LmGSTs1） was functionally expressed in Escherichia coli cells in a soluble form and purified to homogeneity. LmGSTs1 was able to catalyze the biotranslation of glutathione with 1-chloro-2,4-dinitrobenzene, a model substrate for GSTs, as well as with p-nitro-benzyl chloride. Its optimal activity was observed at pH 8.0 and at 30℃. Incubation for 30 min at temperatures below 50℃ scarcely affected the activity. The I50 of reactive blue （RB） was 18.5 μmol L-1. In the presence of 0.05 mmol L-1 ethacrynic acid （ECA）, LmGSTs1 showed （81±3）% of the original activities.

Expression of Glutathione S-transferase π in Human Bladder Cancer

The purpose of this study was to investigate the expression of glutathione stransferase (GST) and its clinical significance in human bladder cancer. GST immunoreactivity was assessed respectively in 49 bladder cancers and 30 normal bladder mucosas by avidin biotin peroxidase complex (ABC) techniques. The corelationship of GST expression and clinical and biological feature of bladder cancer was studied. Positive GST was observed in 44 cases of bladder cancer mucosa (89.8%) and in 18 cases of normal bladder mucosa (60%). In 42 cases of stage G\-\{12\} and 7 cases of stage G\-3 cancer patients the positive GST expression rate was 80.9%(34/42) and 100%(7/7) respectively. In 14 cases of recurrent bladder cancer the total positive expression rate was 92.9% (13/14), meanwhile in 19 cases of stage T23 the positive nuclear staining was seen in 10. Higher rate of GST expression was found in bladder cancer than in normal bladder mucosa, which was implied that the resistance to chemotheraputic drugs in bladde r cancer might be related to the expression of GST. GST expression was correlated with tumor grade. Furthermore, increased intranuclear GST expression might be associated with bladder cancer progression.

ENZYME-LINKED IMMUNOSORBENT ASSAY OF HUMAN PLACENTA TYPE GLUTATHIONE S-TRANSFERASE AND ITS APPLICATION IN THE DIAGNOSIS OF HEPATOCARCINOMA

GST-π was purified from human placenta and its antiserum was raised in rabbits. The antibody IgC was purified and degraded into Fab' fragment which was conjugated with horseradish peroxidase (HRP) using N-succinimidyl-4-(N-maleimido-methyl) cyclo-hexane-1-carboxylate (SMCC) as crosslinking reagent to produce Fab'-HRP conjugate. A sandwich ELISA was established for the microquantitative determination of GST-π. The sensitivity was 11 pg/tube, which was far more sensitive than the radioimmunoassay so far reported. Using this method, the serum GST-π of 41 cases normal adult was found to be 1.06±0.94 ng/ml. The upper limit of the normal value was 2.6 ng/ml. In 30 cases of primary hepatocarcinoma, the level of serum GST-π was 24.4± 17.4 ng/ml, which was 23 times higher than the normal average value (P<0.01). The positive rate was 90%. In contrast, serum GST-π in 25 cases of chronic hepatitis was determined to be 1.74±1.16 ng/ml, which was not significantly different from the normal value (P>0.05). The pseudo-positive rate was 12.0%.

Antibodies against ribosomal protein S29(RPS29)fused with glutathione's transferase specially react with native RPS29 in mouse and human cells

The ribosomal protein S29 also known as RPS29, is not only a component of the 40S subunit of ribosome, but also involved in embryonic development, oncogenesis and other pathologic conditions. However, rare commercial antibody against RPS29 restricts the discovery of precise physiological and pathological function of this protein. In this study, the whole RPS29 gene was inserted into plasmid pGEX-6p-1 to express glutathione＇s transferase （GST） fusion proteins in Escherichia eoli （E. coli） strain BL21. High yields of soluble recombinant proteins were obtained. Mice were immunized with the recombinant RPS29 protein. The serum from the immunized mice could specially react with purified recombinant RPS29 proteins and native RPS29 proteins in CCE cells by western blotting, immunofluorescence staining and flow cytometric analysis. Further more the polyclonal antibodies also reacted specifically with human cell strain ECV304, which showed typical cytoplasmatic fluorescence. The polyclonal antibodies we prepared would be an available tool for studying the roles of RPS29 in embryonic development and human diseases.

Frequency of Null Phenotypes of Glutathione S-Transferase M1 and T1 among the Populations of Tabuk (Northwestern Part of Saudi Arabia)

Background: The variability in the distribution of the null phenotypes of GSTM1 and GSTT1, due to total or partial gene deletion resulting in the lack of the active enzyme, has been reported in different populations, especially in ethnically well-defined groups but not in Tabuk. This study investigated the variability in the distribution of the null phenotypes of GSTM1 and GSTT1 in the population of Tabuk (northwestern part of Saudi Arabia). Method: This study was conducted on 200 subjects of Tabuk—northwestern part of Saudi Arabia among which 100 were chronic smokers and 100 were nonsmokers. The subjects were reporting to hospital for routine checkup. All were without past history of any chronic disease and no significant abnormality. GST genotyping was done by multiplex PCR-based methods. The smoker and control groups were compared using a chi-square test with P GSTM1 deletion homozygosity of 14% and 1% was reported among non smokers and smokers, respectively whereas GSTT1 deletion homozygosity of 28% and 6% was reported among non smokers and smokers, respectively. Our results indicate that there are major differences in allelic distribution of GSTM1 and GSTT1 genes between the two groups investigated. Combined analysis of both genes revealed that 15% of smokers and non smokers harbor the deleted genotype of GSTM1 and 34% of smokers and non smokers harbor the deleted genotype of GSTT1 with significant differences. Conclusion: This study enables selecting subgroups among the general population who are more susceptible to DNA damage and will help genetic studies on the association of GST polymorphisms with disease risks and drug effects in Arab population. Studies with a larger sample size are needed to evaluate and confirm the validity of our results.

小麦抗白粉病相关基因GST克隆与表达

以从小麦抗白粉病相关基因差异表达分析中获得的EST-3(Genbank序列号EX567360)为标签,采用电子克隆的方法对其进行延伸,并对电子克隆结果进行半定量RT-PCR验证,最后对白粉菌不同侵染时间进行了表达分析。经RT-PCR扩增,EST-3表达的带型变化趋势与其在抑制性消减杂交SSH-cDNA的差异显示情况一致,且RT-PCR获得的序列与电子克隆的序列一致性达98%。生物信息学分析表明,该序列是由875bp核苷酸组成的,具有完整的开放阅读框架,编码蛋白为229个氨基酸,GenBank序列号JK841279,含有一个N端和C端谷胱甘肽硫转移酶结构域,该序列与小麦谷胱甘肽硫转移酶基因(GST)一致性较高,达97%。表达分析结果显示,白粉菌侵染24h表达受到抑制,48h开始表达,侵染72h表达最强,96h又开始下降,表明GST基因属于白粉菌诱导型相关基因,参与小麦对白粉病的应答反应。

p63、P-gp、GST-π及Topo-Ⅱ在食管鳞癌组织中的表达

目的:观察比较p63、P-糖蛋白(P-gp)、谷甘胱肽-S-转移酶π(GST-π)、DNA拓扑异构酶(Topo-Ⅱ)在食管癌中的表达情况,并探讨其关系和临床意义。方法:采用免疫组化方法对168例食管鳞癌组织中p63、P-gp、GST-π、Topo-Ⅱ蛋白进行检测,分析其与病理特征的关系及协同表达的意义。结果:p63、P-gp、GST-π、Topo-Ⅱ蛋白在食管鳞癌中的阳性表达率分别为81.0%、57.1%、71.4%、78.6%。p63的表达与肿瘤的临床分期、浸润深度、淋巴结转移相关(P<0.05),P-gp和TopoⅡ的表达与淋巴转移有关(P<0.05),GST-π的表达与肿瘤分化程度、淋巴结转移有关(P<0.05)。p63的阳性表达与P-gp、GST-π呈正相关(r=0.193、0.197,P<0.05),P-gp与GST-π表达呈正相关(r=0.224,P<0.05),P-gp与TopoⅡ表达呈负相关(r=-0.218,P<0.05)。结论:p63、P-gp、GST-π及Topo-Ⅱ均与食管癌发生发展有关,对肿瘤化疗方案的选择和预后判断有重要的临床意义。

日本血吸虫重组28GST T细胞表位谱的预测及鉴定

目的 :用软件预测日本血吸虫重组 2 8GST抗原分子T细胞表位谱 ,并鉴定其Th1型细胞表位。方法 :用软件预测重组2 8GSTT细胞表位谱 ,并筛选几种较好的T细胞表位 ,人工合成表位肽或用基因工程制备重组表位肽融合蛋白。体外刺激经照射的尾蚴感染并用 2 8GST加强免疫的C5 7BL/ 6小鼠 (H 2 b)脾细胞 ,通过淋巴细胞增殖试验、ELISA及流式细胞术等 ,分析各种表位的免疫刺激作用 ,鉴定Th1型细胞表位。结果 :在 9个候选的表位中 ,P6 (73～ 86aa)是刺激作用最强的Th1型细胞表位。结论 :重组 2

杀虫单对二化螟谷胱甘肽转移酶(GSTs)的抑制作用

以二化螟为材料研究了杀虫单、甲胺磷和顺丁烯二酸二乙酯 (DEM)对谷胱甘肽转移酶 (GSTs)的抑制作用。离体抑制试验结果表明 ,杀虫单和DEM都对二化螟GSTs活性有强烈的抑制作用 ,在实验设置的反应体系中 ,2 5mmol·L-1杀虫单和DEM与酶液保温 2min后 ,对GSTs的抑制率达到 70 %以上 ;6min后 ,达到 90 %以上。杀虫单和DEM的抑制中浓度 (IC50 )分别是 1 33和 1 71mmol·L-1,而甲胺磷没有抑制作用。活体实验结果显示 ,用 0 1mol·L-1杀虫单、甲胺磷和DEM 0 0 5 μL分别处理二化螟 4龄幼虫 48h后 ,杀虫单和DEM对GSTs活性的抑制率分别达 10 3%和 14 5 % ,甲胺磷则表现出 8 6 %的诱导作用。酶动力学实验显示 ,杀虫单和DEM都是GSTs的可逆性抑制剂 ,这种抑制作用不是竞争性的。由此表明 ,杀虫单是GSTs的强抑制剂 ,这在杀虫单的毒杀机制中具有重要的意义。

陆地棉GST基因家族全基因组分析

谷胱甘肽转移酶(glutathione-S-transferase,GST)是一种普遍存在的具有多功能的超家族蛋白,在植物初次生代谢、逆境胁迫、胞间信号传递等方面具有重要作用;同时,作为配体其在植物激素代谢以及物质转运方面也发挥作用。为了解析陆地棉(Gossypium hirsutum L.)GST基因家族的信息,本研究对该基因家族成员的种类、进化关系、物理定位、基因结构和保守基序以及表达模式进行了分析。结果显示,在陆地棉全基因组中共含有70个GST基因,进化树和基因结构分析将该家族分为U族、F族、T族、Z族、EF1Bγ族和TCHQD族。基因定位分析发现,除了AD/At2、AD/At4、AD/At5、AD/Dt5、AD/Dt10号染色体上没有GST基因外,其他染色体上都有GST基因,并且在AD/At9、AD/Dt7、AD/Dt12、AD/Dt13这4条染色体上出现基因簇。对F族(Phi类)9个GST基因进行荧光定量分析,结果表明,除Gh GSTF1可能为假基因外,Gh GSTF2~9等8个基因在陆地棉根、茎、叶以及各个发育时期的纤维中均有表达;结合生物信息学分析,推测Gh GSTF8可能参与原花青素/花青素的转运和积累;Gh GSTF4、Gh GSTF6和Gh GSTF9可能在调节陆地棉的生长和胁迫反应中起作用,而Gh GSTF2、Gh GSTF3、Gh GSTF5和Gh GSTF7的功能还有待进一步研究。本研究为陆地棉GST基因家族的分子进化及功能研究提供了理论依据。

GSTM1与GSTP1基因多态性对红细胞GST酶活性的影响

目的研究单独或结合GSTM1纯合缺失基因型时,不同GSTP1基因型对红细胞GST酶活性的影响。方法GST酶活性参照Habig等报道的方法用紫外分光光度法测定。用多重PCR分析GSTM1基因多态性,PCRRFLP检测GSTP15号外显子105位密码子基因多态性。结果汉族人的GSTM1纯合缺失频率为56.1%,GSTP1105I/I、I/V和V/V基因型频率分别为60.7%、35.2%和4.1%。杂合I/V基因型组的平均GST酶活性(3.53±0.63U·g-1Hb)比野生I/I基因型的GST酶活性低(4.25±1.07U·g-1Hb,P=0.000),比突变V/V基因型的GST酶活性高(2.44±0.67U·g-1Hb,P=0.004)。在GSTM1(-)基因组,GSTM1(-)/GSTP1I/I基因型携带者的GST酶活性比GSTM1(-)/GSTP1I/V或V/V携带者的高,而在GSTM1(+)组,两组间的酶活性无差异。不同年龄组平均GSTs活性无差异,而女性的平均GSTs活性比男性的平均值高,但差异无显著性。结论尽管其它GST酶可能会稀释GSTP1基因型对GST酶活性的效果,GST酶活性仍与GSTP1105Val基因型呈很强的相关性。

CpG寡核苷酸的筛选及其对rSj28GST的免疫佐剂作用

目的 研究 Cp G寡核苷酸 (Cp G ODN)对日本血吸虫重组 2 8k Da GST(r Sj2 8GST)抗原分子的免疫佐剂作用和诱导 Th1型反应的效应。方法 用淋巴细胞增殖试验筛选对鼠有良好免疫刺激作用的 Cp G ODN;用基因工程方法生产 r Sj2 8GST抗原分子 ;EL ISA检测小鼠抗 r Sj2 8GST的同型抗体 Ig G1 、Ig G2 a水平。结果 筛选出的 Cp G ODN如 182 6、2 0 0 2、182 6 4、2 10 2等具有良好的免疫刺激活性 ,其 SI>2 ;纯化的 r Sj2 8GST抗原分子 ,呈单一电泳条带 ;以 Cp G ODN为 r Sj2 8GST免疫佐剂 ,其免疫小鼠产生的抗体滴度显著高于对照组 ,且同型抗体 Ig G2 a与 Ig G1 比值也明显升高。结论 Cp G ODN182 6 4是一种良好的免疫佐剂 ,并可显著诱导