多烯磷脂酰胆碱联合谷胱甘肽治疗妊娠期肝内胆汁淤积症患者的效果及对围产结局的影响

目的探讨多烯磷脂酰胆碱联合谷胱甘肽治疗妊娠期肝内胆汁淤积症(ICP)患者的效果及对围产结局的影响。方法选取2015年1月～2018年12月ICP患者124例,随机分为两组。两组均予以卧床休息、间断吸氧、高渗葡萄糖、维生素C及能量合剂等治疗。对照组在此基础上予以多烯磷脂酰胆碱针15 mL静脉滴注,1次/d;观察组在对照组基础上再加谷胱甘肽针2.4 g静脉滴注,1次/d。两组均连用2周。观察两组治疗后临床症状及血清生化指标改善情况,并比较其围产结局。结果治疗2周后,两组瘙痒评分均较治疗前显著下降(P0.01或P0.05),且观察组下降幅度较对照组更明显(P0.05);观察组黄疸和瘙痒消失时间明显少于或短于对照组(P0.05)。治疗2周后,两组血清ALT、TBA和AST水平均较治疗前明显下降(P0.01或P0.05),且观察组下降幅度较对照组更明显(P0.05)。同时观察组宫内窘迫、早产和新生儿窒息发生率明显低于对照组,分娩出血量明显少于对照组,新生儿体重明显大于对照组(P0.05)。结论多烯磷脂酰胆碱联合谷胱甘肽治疗ICP患者不仅能明显改善其临床症状,降低肝酶指标与胆汁酸水平,而且能降低发生不良围产结局,有利于母婴安全。     

Polymorphism analysis of Glutathione S-transferase A1 in patients with hematological diseases and its effect on GST enzyme activity

Glutathione S-transferases (GSTs) are important drug-metabolizing enzymes that catalyze the binding of glutathione (GSH) to electrophilic substances. GST has genetic polymorphism, and the enzyme activity of GST affects the metabolism of certain drugs in vivo. In the present day, we investigated the GST enzyme activity and GSTA1 gene polymorphism in 170 patients with hematological diseases and explored their relationship. The GSTA1 gene polymorphism of the patient was analyzed by PCR- restriction fragment length polymorphism (PCR-RFLP) technique, and the base sequences of the four mutation sites (-631, -567, -69, and -52) in the promoter region were determined by DNA-Sequencer. The patient's GST enzyme activity was calculated by measuring the rate at which it catalyzed the reaction between 1-chloro-2,4-dinitrobenzene (CDNB) and GSH. The average GST enzyme activities of males and females were 5.20±0.13 and 5.17±0.12 nmol/min/mL, respectively, and the difference was not significant (P = 0.91). The frequencies of genotypes GSTA1*A*A (wild genotype), GSTA1*A*B (heterozygous genotype), and GSTA1*B*B (homozygous mutant genotype) were 75.3%, 22.9%, and 1.8%, respectively. Alleles GSTA1*A and *B were distributed at 86.8% and 13.2%, respectively. The genotype frequency distribution between males and females was no significant difference by Pearson’s chi-square test (P = 0.743). The average GST activity of the heterozygous mutant genotype (4.83±0.76 nmol/min/mL) was lower than the wild genotype (5.34±1.26 nmol/min/mL, P = 0.018), and higher than that of the homozygous mutant genotype (3.32±0.07 nmol/min/mL, P = 0.022). These findings might help us improve the individualized treatment of patients with hematological diseases in the future and promote the development of precision medicine for blood diseases.     

还原型谷胱甘肽对正加速度暴露下急性胃黏膜损伤大鼠胃黏膜的影响

目的研究还原型谷胱甘肽(GSH)对正加速度(+Gz)暴露下急性胃黏膜损伤大鼠胃黏膜的影响及可能机制。方法 40只雄性SD大鼠随机分成4组:无水乙醇对照组(A组)、+5 Gz值暴露组(B组)、+10 Gz值暴露组(C组)、GSH预处理组(D组),每组10只。D组适应性喂养7 d后,连续3 d腹腔注射GSH。4组均于10 d后禁食24 h,禁水12 h,用无水乙醇灌胃1 h后,A组不受+Gz作用,B组暴露于+5 Gz值3 min,C、D组暴露于+10 Gz值3 min,下离心机后观察各组胃黏膜损伤情况,并检测胃黏膜中丙二醛(MDA)、GSH的含量及谷胱甘肽过氧化物酶(GSH-Px)的活性。结果各组大鼠胃黏膜在肉眼观察均有损伤,损伤程度:+10Gz值暴露组+5 Gz值暴露组无水乙醇对照组GSH预处理组,B组与A组相比差异均有统计学意义(P均0.05),C组分别与A、B两组相比差异均有统计学意义(P0.05);D组胃黏膜损伤最轻,与C组相比差异有显著统计学意义(P0.01)。与A组相比,B组胃黏膜中MDA、GSH含量变化不明显,差异均无统计学意义(P均0.05),GSH-Px活性明显升高,差异有显著统计学意义(P0.01);与A、B两组相比,C组胃黏膜中MDA含量升高明显,GSH含量降低明显,GSH-Px活性明显降低,差异均有统计学意义(P0.05);与C组相比,D组胃黏膜MDA含量明显降低,GSH的含量明显升高,GSH-Px的活性明显升高,差异有统计学意义(P0.05)。结论 GSH对+Gz值暴露引起的急性胃黏膜损伤大鼠的胃黏膜有预防及保护作用,该作用可能是通过增加胃黏膜GSH的含量、GSH-Px的活性及抑制MDA的作用而实现的。     

大鼠肝纤维化肝组织中抗衰老标志蛋白和抗增殖蛋白的表达研究

目的 研究猪血清所致肝纤维化大鼠肝组织中抗衰老标志蛋白(RGN)、抗增殖蛋白(PHB)表达及谷胱甘肽复方注射液(CGII)对其的干预. 方法 健康雄性Wistar大鼠40只,随机分为正常组、造模组.正常组予等渗盐水腹腔注射0.5 ml/只,造模组予腹腔注射无菌猪血清0.5ml/只,每周两次,连续8周,随机处死造模大鼠4只行病理学检查,验证造模成功后,停止注射猪血清,剩余造模组大鼠随机分为纤维化组和CGII干预组.CGII干预组大鼠给予浓度60 mg/mlCGII肌肉注射;正常组和纤维化组均予等单位体质量容积的等渗盐水肌肉注射,每日一次,连续6周.取大鼠肝组织行HE及Masson染色.RT-PCR和免疫组织化学检测肝组织中RGN、PHB mRNA及蛋白质表达. 计量资料采用单因素方差分析,两两比较采用LDS检验,病理学半定量结果采用秩和检验分析. 结果 纤维化组大鼠肝组织RGN、PHB mRNA相对表达量分别为75.99±12.8、64.54±12.35较正常组的182.09±17.84、192.20±17.12明显减低,F值分别为105.646、347.232,P值均＜0.001,差异均有统计学意义;RGN、PHB相对蛋白表达量分别为10.85±2.81、14.5±2.75较正常组的61.08±7.10、55.86±4.23表达均明显降低,F值分别为226.343、525.889,P值均＜ 0.001,差异均有统计学意义.CGII干预后大鼠肝纤维化程度较纤维化组明显减轻(P值均＜ 0.001),肝组织RGN、PHB的mRNA和蛋白表达均较纤维化组明显增高,差异有统计学意义(P值均＜ 0.001).结论 RGN及PHB在猪血清所致大鼠肝纤维化肝组织表达降低.     

两种不同盖髓剂对人牙髓细胞毒性的体外研究

目的:研究丁香酚及Dycal对人牙髓细胞的毒性。方法:体外组织块法培养人牙髓细胞,采用噻唑蓝比色法(MTT法)及台盼蓝染色活细胞计数法来评价不同浓度及不同时间段丁香酚及Dycal对牙髓细胞细胞毒性,比色法测定丁香酚干预组细胞内总谷胱甘肽含量。结果:0.1mmol/L浓度的丁香酚及0.1%浓度的Dycal对人牙髓细胞毒性明显,其对细胞毒性反应成浓度及时间依赖性改变。各丁香酚组细胞内谷胱甘肽含量低于对照组,第3天丁香酚最低浓度组细胞内谷胱甘肽含量高于高浓度组。结论:高浓度丁香酚及Dycal对牙髓细胞毒性明显,细胞毒性与浓度及作用时间有关。低浓度丁香酚可能通过谷胱甘肽的参与抵抗氧化应激,从而起到对细胞的保护作用。Dycal可能更加适合用于间接盖髓。     

谷胱甘肽硫转移酶GSTO1基因Glu155△Glu位点多态性与燃煤污染型地方性砷中毒的关系

目的 探讨谷胱甘肽硫转移酶GSTO 1基因Glu155△Glu位点多态性与贵州省燃煤污染型地方性砷中毒发生的关系.方法 采用双相引物-聚合酶链反应(polymerase chain reaction-with confronting twopair primers,PCR-CTPP)技术检测贵州省130例燃煤污染型地方性砷中毒患者和130例健康对照人群的GSTO1 Glu155△Glu基因型,DNA测序验证检测结果.用非条件Logistic回归模型分析GSTO1 Glu155△Glu多态性与燃煤污染型砷中毒发病风险的关系.结果 PCR-CTPP法检测出的Glu/Glu和Glu/△Glu基因型结果与DNA测序结果一致.砷中毒组GSTO1 Glu155△Glu位点Glu/Glu和Glu/△Glu基因型分布频率分别为94.85％(92/97)和5.15％(5/97),对照组分别为99.15％(117/118)和0.85％(1/118),二者比较差异有统计学意义(x2=3.896,P＜0.05),两组均未检测到△Glu/△Glu基因型,调整年龄和性别后发现GSTO1 Glu155△Glu的多态性是燃煤污染型砷中毒发病的危险因素[比值比(OR)=1.85,95％可信区间(CI):1.39～17.48].结论 GSTO1 Glu155△Glu多态性与燃煤污染型砷中毒的发病风险有一定关联,两者之间的关系值得进一步扩大样本量进行深入研究.     

还原型谷胱甘肽联合安络化纤丸治疗酒精性脂肪肝100例

目的探索还原型谷胱甘肽联合安络化纤丸治疗酒精性脂肪肝的疗效。方法将200例酒精性脂肪肝患者随机分为两组,100例用还原型谷胱甘肽联合安络化纤丸为治疗组,100例单用还原型谷胱甘肽为对照组,疗程均为1个月。结果治疗组有效率91.00%(91/100),对照组有效率70.00%(70/100),两组比较有显著性差异(P<0.001)。结论还原型谷胱甘肽联合安络化纤丸治疗酒精性脂肪肝有良好效果,值得临床推广应用。     

Proteomic analysis of glutathione S-transferase isoforms in mouse liver mitochondria

AIM: To survey glutathione (GSH) S-transferase (GST) isoforms in mitochondria and to reveal the isoforms’ biological significance in diabetic mice.METHODS: The presence of GSTs in mouse liver mitochondria was systematically screened by two proteomic approaches, namely, GSH affinity chromatography/two dimensional electrophoresis (2DE/MALDI TOF/TOF MS) and SDS-PAGE/LC ESI MS/MS. The proteomic results were further confirmed by Western blotting using monoclonal antibodies against GSTs. To evaluate the liver mitochondrial GSTs quantitatively, calibration curves were generated by the loading amounts of individual recombinant GST protein vs the relative intensities elicited from the Western blotting. An extensive comparison of the liver mitochondrial GSTs was conducted between normal and db/db diabetic mice. Student’s t test was adopted for the estimation of regression and significant difference.RESULTS: Using GSH affinity/2DE/MALDI TOF/TOF MS, three GSTs, namely, alpha3, mu1 and pi1, were identified; whereas five GSTs, alpha3, mu1, pi1, kappa1 and zeta1, were detected in mouse liver mitochondria using SDS-PAGE/LC ESI MS/MS, of these GSTs, GST kappa1 was reported as a specific mitochondrial GST. The R² values of regression ranged between values of about 0.86 and 0.98, which were acceptable for the quantification. Based on the measurement of the GST abundances in liver mitochondria of normal and diabetic mice, the four GSTs, alpha3, kappa1, mu1 and zeta1, were found to be almost comparable between the two sets of animals, whereas, lower GST pi1 was detected in the diabetic mice compared with normal ones, the signal of Western blotting in control and db/db diabetic mice liver mitochondria is 134.61 ± 53.84 vs 99.74 ± 46.2, with P < 0.05.CONCLUSION: Our results indicate that GSTs exist widely in mitochondria and its abundances of mitochondrial GSTs might be tissue-dependent and disease-related.     

Impaired GSH biosynthesis disrupts eye development,lens morphogenesis and PAX6 function

PurposeThe purpose of this study was to elucidate the role and molecular consequences of impaired glutathione (GSH) biosynthesis on eye development.MethodsGSH biosynthesis was impaired in surface ectoderm-derived ocular tissues by crossing Gclc^f/f mice with hemizygous Le-Cre transgenic mice to produce Gclc^f/f/Le-Cre^Tg/- (KO) mice. Control mice included Gclc^f/f and Gclc^wt/wt/Le-Cre^Tg/- mice (CRE). Eyes from all mice (at various stages of eye development) were subjected to histological, immunohistochemical, Western blot, RT-qPCR, RNA-seq, and subsequent Gene Ontology, Ingenuity Pathway Analysis and TRANSFAC analyses. PAX6 transactivation activity was studied using a luciferase reporter assay in HEK293T cells depleted of GSH using buthionine sulfoximine (BSO).ResultsDeletion of Gclc diminished GSH levels, increased reactive oxygen species (ROS), and caused an overt microphthalmia phenotype characterized by malformation of the cornea, iris, lens, and retina that is distinct from and much more profound than the one observed in CRE mice. In addition, only the lenses of KO mice displayed reduced crystallin (α, β), PITX3 and Foxe3 expression. RNA-seq analyses at postnatal day 1 revealed 1552 differentially expressed genes (DEGs) in the lenses of KO mice relative to those from Gclc^f/f mice, with Crystallin and lens fiber cell identity genes being downregulated while lens epithelial cell identity and immune response genes were upregulated. Bioinformatic analysis of the DEGs implicated PAX6 as a key upstream regulator. PAX6 transactivation activity was impaired in BSO-treated HEK293T cells.ConclusionsThese data suggest that impaired ocular GSH biosynthesis may disrupt eye development and PAX6 function.