CAJ | 학술논문

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.
목적 료해을이철매Ⅰ(GLO-Ⅰ)재자궁내막암조직화세포중적표체정황,탐토GLO-Ⅰ대자궁내막암세포증식여조망적영향.　방법응용면역조화항생물소단백-생물소복합물(ABC)법검측자궁내막암(29빈)、정상자궁내막(19빈)조직중GLO-Ⅰ단백적표체,자외분광광도계검측자궁내막암급기암방조직、정상자궁내막조직중GLO-Ⅰ적활성.GLO-Ⅰ소분자RNA(siRNA)전염자궁내막암세포주lshikawa세포후,채용단백인적법급역전록(RT)-PCR기술분별검측기GLO-Ⅰ단백급mRNA적표체수평,채용사갑기우담서람(MTT)비색법、류식세포의분별검측기세포증식급조망정황.결과 (1)자궁내막암조직중GLO-Ⅰ단백적양성표체솔위76%(22/29),정상자궁내막조직위0/19,량자비교,차이유통계학의의(P<0.01).정상자궁내막、자궁내막암암방조직중GLO-Ⅰ활성(분별위1.1、0.8 IU/mg)교약,자궁내막암조직중GLO-Ⅰ활성(92.3 IU/mg)명현승고,차이유통계학의의(P<0.01).(2)Ishikawa세포전염GLO-Ⅰ silRNA후,기GLO-Ⅰ mRNA적표체수평위0.25±0.06,저우음성대조(전염여목적기인무관적siRNA)적0.93±0.10,차이유통계학의의(P<0.01);기GLO-Ⅰ단백적표체수평위0.38±±0.06,저우음성대조적0.94±0.13,차이야유통계학의의(P<0.01).(3)Ishikawa세포전염GLO-Ⅰ siRNA후,기세포증식능력여공백대조(부가전염시제)비교명현강저(P=0.028);기세포조망솔위(6.7±0.8)%,고우음성대조적(1.2±0.4)%화공백대조적(1.4±0.4)%,차이유통계학의의(P<0.01).결론 GLO-Ⅰ재자궁내막암조직화세포중균정과도표체,자궁내막암조직중GLO-Ⅰ활성이상승고.억제GLO-Ⅰ기인표체가촉진자궁내막암세포조망,병억제기증식.
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.