Expression of glyoxalase Ⅰ and its effect on cell proliferation and apoptosis in endometrial carcinoma

Objective To examine the expressions of glyoxalase Ⅰ (GLO-Ⅰ ) in endometrial cancer tissues and cell lines and to investigate the roles of GLO-Ⅰ on proliferation and apoptosis in endometrial cancer cells. Methods Immunohistochemistry, western blot and RT-PCR were used to investigate the expressions of GLO-Ⅰ protein and mRNA in endometrial cancer tissues and Ishikawa cell lines ;enzyme activity of GLO-Ⅰ in normal endometrium, endometrial cancer and paraneoplastic tissue samples was detected with spectrophotometer; proliferation and apoptosis of Ishikawa cell before and after RNA interference (RNAi) procedure were detected by the methyl thiazolyl tetrazolium (MTT) and flow cytometry, respectively. Results (1)There were significant differences of GLO-Ⅰ expression between normal endometrium (0/19) and endometrial cancer tissues ( 76%, 22/29 ); these were also significant differences of enzyme activity of GLO-Ⅰ among normal endometrium, paraneoplastic and endometrial cancer tissues( 1.1,0.8 vs 92.3 IU/mg; P <0.01 ). Enzyme activity of GLO-Ⅰ in fresh normal endometrium and paraneoplastic tissues was weak, while that of fresh endometrial cancer tissues was as high as 92. 3 IU/mg in average. (2)The expression of GLO-Ⅰ mRNA in Ishikawa cell transfected with GLO-Ⅰ siRNA was significantly lower than that in negative group (0.25 ± 0.06 vs 0.93 ± 0.10, P < 0.0l ), and the similar results that in the expression of GLO-Ⅰ protein (0.38 ±0.06 vs 0.94 ±0.13, P <0.01 ). (3) Proliferation in Ishikawa cell was significantly inhibited after silencing RNA expression of GLO-Ⅰ ( P = 0.028 ). The apoptosis rate of cells transfected with GLO- Ⅰ siRNA was significantly higher than that of negative control group and blank control group [ ( 6.7 ± 0.8 ) % vs ( 1.2 ± 0.4) %, ( 1.4 ± 0.4 ) %; P < 0.01 ]. Conclusion The expression and enzyme activity of GLO- Ⅰ is significantly increased in endometrial cancer, which could promote abnormal proliferation and inhibit apoptosis in endometrial cancer cells.     

乙二醛酶Ⅰ在子宫内膜癌组织和细胞中的表达及其对细胞增殖和凋亡的影响

目的 了解乙二醛酶Ⅰ(GLO-Ⅰ)在子宫内膜癌组织和细胞中的表达情况,探讨GLO-Ⅰ对子宫内膜癌细胞增殖与凋亡的影响.　方法应用免疫组化抗生物素蛋白-生物素复合物(ABC)法检测子宫内膜癌(29份)、正常子宫内膜(19份)组织中GLO-Ⅰ蛋白的表达,紫外分光光度计检测子宫内膜癌及其癌旁组织、正常子宫内膜组织中GLO-Ⅰ的活性.GLO-Ⅰ小分子RNA(siRNA)转染子宫内膜癌细胞株lshikawa细胞后,采用蛋白印迹法及逆转录(RT)-PCR技术分别检测其GLO-Ⅰ蛋白及mRNA的表达水平,采用四甲基偶氮唑蓝(MTT)比色法、流式细胞仪分别检测其细胞增殖及凋亡情况.结果 (1)子宫内膜癌组织中GLO-Ⅰ蛋白的阳性表达率为76%(22/29),正常子宫内膜组织为0/19,两者比较,差异有统计学意义(P<0.01).正常子宫内膜、子宫内膜癌癌旁组织中GLO-Ⅰ活性(分别为1.1、0.8 IU/mg)较弱,子宫内膜癌组织中GLO-Ⅰ活性(92.3 IU/mg)明显升高,差异有统计学意义(P<0.01).(2)Ishikawa细胞转染GLO-Ⅰ silRNA后,其GLO-Ⅰ mRNA的表达水平为0.25±0.06,低于阴性对照(转染与目的基因无关的siRNA)的0.93±0.10,差异有统计学意义(P<0.01);其GLO-Ⅰ蛋白的表达水平为0.38±±0.06,低于阴性对照的0.94±0.13,差异也有统计学意义(P<0.01).(3)Ishikawa细胞转染GLO-Ⅰ siRNA后,其细胞增殖能力与空白对照(仅加转染试剂)比较明显降低(P=0.028);其细胞凋亡率为(6.7±0.8)%,高于阴性对照的(1.2±0.4)%和空白对照的(1.4±0.4)%,差异有统计学意义(P<0.01).结论 GLO-Ⅰ在子宫内膜癌组织和细胞中均呈过度表达,子宫内膜癌组织中GLO-Ⅰ活性异常升高.抑制GLO-Ⅰ基因表达可促进子宫内膜癌细胞凋亡,并抑制其增殖.     

The relationship between single nucleotide polymorphisms in estrogen-metabolizing genes CYP1A1,CYP17,COMT and estrogen receptor alpha and the risk of endometrial adenocarcinoma among the Chinese women

<正>Objective:To explore whether polymorphisms of the genes responsible for catechol estrogen(CE)formation via estrogen biosynthesis(CYP17)and hydroxylation (CYP1A1)and CE inactivation(COMT)and ERa are associated with an elevated risk for en- dometrial adenocarcinoma in Chinese women.Methods:A multigenic case-control study was conducted,eighty-seven endometrial adenocarcinoma patients and ninety controls were recrui- ted.PCR-RFLP assays were used to determine the genotypes of estrogen-metabolizing genes and ERa gene.Results:The endometrial adenocarcinoma risk associated with individual susceptibili- ty genotypes varied among the six polymorphic sites and was the highest for CYP17,followed by CYP1 A1 Ile-Val,CYP1A1 MspI,COMT,ERa XhaI and ERa PvuII.Multivariate logistic regres- sion showed the CYP1A1 MspI genotype was the most significant determinant for endometrial adenocarcinoma development and was associated with a 3.61 fold increase in risk(95% confi- dence interval,1.73～7.55).Furthermore,a trend of increasing risk for developing endometrial adenocarcinoma was found in women harboring higher numbers of high-risk genotypes.Conclu- sion:The CYP1A1,CYP17 and ERa XbaI genotypes are related to the susceptibility of endome- trial adenocarcinoma,they may be useful markers for predicting endometrial adenocarcinoma susceptibility.The allele encoding for low acticity COMT,ERa PvuII may not be a genetic risk factor for endometrial adenocarcinoma.