凋亡蛋白抑制因子Livin在子宫内膜癌中的表达及其对肿瘤细胞侵袭和迁移能力的影响
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摘要
目的研究凋亡蛋白抑制因子Livin在子宫内膜癌中的表达及其对肿瘤细胞侵袭和迁移能力的影响。方法选取承德医学院附属医院妇科2015年3月—2016年10月手术切除的子宫内膜标本共118例,其中子宫内膜癌组织62例,非典型增生内膜组织26例,正常子宫内膜组织30例。采用荧光定量PCR法检测并比较Livin mRNA在子宫内膜癌、非典型增生和正常内膜组织中,以及在不同临床病理特征子宫内膜癌组织中的表达,分析其差异性表达与子宫内膜癌临床病理分期的关系。采用脂质体法用Livin的小分子干扰RNA(Livin-siRNA)和阴性对照序列(Livin-NC)转染Ishikawa和HEC-1-A细胞,分为ISK-I组和HEC-I组(均转染Livin-siRNA)、ISK-NC组和HEC-NC组(均转染Livin-NC)、ISK-B组和HEC-B组(均仅以10%FBS培养细胞)。转染后48 h,采用Western印迹法检测并比较Livin蛋白在各组细胞中的表达。进行Transwell小室侵袭和迁移实验,观察干扰Livin对细胞侵袭和迁移能力的影响。结果 Livin mRNA在子宫内膜癌组织中的相对表达量(8.624±0.264)显著高于非典型增生内膜组织(3.497±0.186)和正常内膜组织(1.000±0.098,P值均<0.05)。比较Livin mRNA在不同年龄(<60岁与≥60岁)、不同肌层浸润深度(<1/2与≥1/2)、不同病理分级(G_1级与G_2+G_3级)、不同国际妇产科联盟(FIGO)分期(Ⅰ或Ⅱ期与Ⅲ或Ⅳ期)和有无淋巴结转移的患者中的表达,结果显示,Livin mRNA仅在不同肌层浸润深度(相对表达量分别为6.392±0.149和9.227±0.216,t=7.325,P=0.016)和不同病理分级(相对表达量分别为7.068±0.103和9.392±0.125,t=5.578,P=0.027)的患者中表达的差异均有统计学意义。转染后48 h,ISK-I组(0.395±0.012)和HEC-I组(0.514±0.022)的Livin蛋白相对表达量分别显著低于ISK-NC组(1.108±0.048)、HEC-NC组(1.426±0.062)、ISK-B组(1.164±0.054)和HEC-B组(1.448±0.071,P值均<0.05)。侵袭实验结果显示,ISK-I组[(20.2±3.6)个]和HEC-I组[(28.8±3.2)个]的穿膜细胞数分别显著少于ISK-NC组[(42.8±4.2)个]和HEC-NC组[(56.7±3.8)个,P值均<0.05]。迁移实验结果显示,ISK-I组[(22.5±2.2)个]和HEC-I组[(30.8±2.4)个]的穿膜细胞数分别显著少于ISK-NC组[(52.8±3.6)个]和HEC-NC组[(58.9±4.5)个,P值均<0.05)]。结论 Livin在子宫内膜癌组织中表达上调,且与肌层浸润深度和病理分级有关。下调Livin表达可抑制子宫内膜癌细胞的侵袭和迁移能力。
Objective To study the expression of apoptosis protein inhibitor—Livin in endometrial carcinoma and its effect on invasion and migration of tumor cells. Methods A total of 118 cases of endometrial specimens were obtained from the Department of Gynecology, Affiliated Hospital of Chengde Medical College from March 2015 to October 2016, including 62 cases of endometrial carcinoma, 26 cases of atypical proliferative endometrium, and 30 cases of normal endometrial tissue. Fluorescence quantitative polymerase chain reaction(PCR) was used to detect the expression of Livin mRNA in endometrial carcinoma, atypical hyperplasia and normal endometrial tissue, and in different clinicopathological features of endometrial carcinoma tissues. The relationship between the expression of Livin mRNA and clinicopathological staging of endometrial carcinoma was analyzed. Livin-siRNA and negative control sequence(Livin-NC) were transfected into Ishikawa and HEC-1-A cells by liposome method. They were divided into ISK-I group, and HEC-I group(transfected by Livin-siRNA), ISK-NC group, HEC-NC group(transfected by Livin-NC), ISK-B group, HEC-B group(only cultured with 10% fetal bovine serum). The expression of Livin protein in each group was detected by Western Blot 48 h after transfection. Transwell chamber invasion and migration experiments were performed to observe the effect of interference with Livin on cell invasion and migration. Results The relative expression of Livin mRNA in endometrial carcinoma tissues was 8.624±0.264, which was significantly higher than that in atypical hyperplasia tissues(3.497±0.186) and normal endometrial tissues(1.000±0.098, both P<0.05). Livin mRNA in different ages(<60 years and ≥60 years), depths of muscle layers(<1/2 and ≥1/2), pathological grades(G_1 and G_2+G_3), international obstetrics and gynecology alliances(FIGO) stage(Ⅰ or Ⅱ and Ⅲ or Ⅳ) and expression in patients with and without lymph node metastasis were compared. The results showed that Livin mRNA was only statistically significant in patients with different depths of myometrial invasion(relative expression levels were 6.392±0.149 and 9.227±0.216, t=7.325, P=0.016) and different pathological grades(relative expression levels were 7.068±0.103 and 9.392±0.125, t=5.578, P=0.027). At 48 hours after transfection, the relative expressions of Livin protein in ISK-I(0.395±0.012) and HEC-I(0.514±0.022) groups were significantly lower than that in ISK-NC(1.108±0.048), HEC-NC(1.426±0.062), ISK-B(1.164±0.054) and HEC-B(1.448±0.071) groups(all P<0.05). The results of the invasion test showed that the number of transmembrane cells in the ISK-I group(20.2±3.6) and HEC-I group(28.8±3.2) were significantly less than those in the ISK-NC group(42.8±4.2) and HEC-NC group(56.7±3.8, both P<0.05). The results of the migration test showed that the number of transmembrane cells in the ISK-I group(22.5±2.2) and HEC-I group(30.8±2.4) were also significantly less than those in the ISK-NC group(52.8±3.6) and HEC-NC group(58.9±4.5, both P<0.05). Conclusion The expression of Livin in endometrial carcinoma is up-regulated and related to the depth of myometrial invasion and pathological grade. Down-regulation of Livin expression can inhibit the invasion and migration of endometrial carcinoma cells.(Shanghai Med J, 2019, 42: 222-226)
Objective To study the expression of apoptosis protein inhibitor—Livin in endometrial carcinoma and its effect on invasion and migration of tumor cells. Methods A total of 118 cases of endometrial specimens were obtained from the Department of Gynecology, Affiliated Hospital of Chengde Medical College from March 2015 to October 2016, including 62 cases of endometrial carcinoma, 26 cases of atypical proliferative endometrium, and 30 cases of normal endometrial tissue. Fluorescence quantitative polymerase chain reaction(PCR) was used to detect the expression of Livin mRNA in endometrial carcinoma, atypical hyperplasia and normal endometrial tissue, and in different clinicopathological features of endometrial carcinoma tissues. The relationship between the expression of Livin mRNA and clinicopathological staging of endometrial carcinoma was analyzed. Livin-siRNA and negative control sequence(Livin-NC) were transfected into Ishikawa and HEC-1-A cells by liposome method. They were divided into ISK-I group, and HEC-I group(transfected by Livin-siRNA), ISK-NC group, HEC-NC group(transfected by Livin-NC), ISK-B group, HEC-B group(only cultured with 10% fetal bovine serum). The expression of Livin protein in each group was detected by Western Blot 48 h after transfection. Transwell chamber invasion and migration experiments were performed to observe the effect of interference with Livin on cell invasion and migration. Results The relative expression of Livin mRNA in endometrial carcinoma tissues was 8.624±0.264, which was significantly higher than that in atypical hyperplasia tissues(3.497±0.186) and normal endometrial tissues(1.000±0.098, both P<0.05). Livin mRNA in different ages(<60 years and ≥60 years), depths of muscle layers(<1/2 and ≥1/2), pathological grades(G_1 and G_2+G_3), international obstetrics and gynecology alliances(FIGO) stage(Ⅰ or Ⅱ and Ⅲ or Ⅳ) and expression in patients with and without lymph node metastasis were compared. The results showed that Livin mRNA was only statistically significant in patients with different depths of myometrial invasion(relative expression levels were 6.392±0.149 and 9.227±0.216, t=7.325, P=0.016) and different pathological grades(relative expression levels were 7.068±0.103 and 9.392±0.125, t=5.578, P=0.027). At 48 hours after transfection, the relative expressions of Livin protein in ISK-I(0.395±0.012) and HEC-I(0.514±0.022) groups were significantly lower than that in ISK-NC(1.108±0.048), HEC-NC(1.426±0.062), ISK-B(1.164±0.054) and HEC-B(1.448±0.071) groups(all P<0.05). The results of the invasion test showed that the number of transmembrane cells in the ISK-I group(20.2±3.6) and HEC-I group(28.8±3.2) were significantly less than those in the ISK-NC group(42.8±4.2) and HEC-NC group(56.7±3.8, both P<0.05). The results of the migration test showed that the number of transmembrane cells in the ISK-I group(22.5±2.2) and HEC-I group(30.8±2.4) were also significantly less than those in the ISK-NC group(52.8±3.6) and HEC-NC group(58.9±4.5, both P<0.05). Conclusion The expression of Livin in endometrial carcinoma is up-regulated and related to the depth of myometrial invasion and pathological grade. Down-regulation of Livin expression can inhibit the invasion and migration of endometrial carcinoma cells.(Shanghai Med J, 2019, 42: 222-226)
引文
[1]冯凤芝,范辰辰,林仲秋.子宫内膜癌的靶向治疗[J].中国实用妇科与产科杂志,2017,33(5):472-476.
[2]杨媛,王建六,魏丽惠.Notch信号通路与子宫内膜癌关系的研究进展[J].中华妇产科杂志,2015,50(5):391-393.
[3]LIU C,WU X,LUO C,et al.Antisense oligonucleotide targeting Livin induces apoptosis of human bladder cancer cell via a mechanism involving caspase 3[J/OL].J Exp Clin Cancer Res,2010,29:63[2018-04-20].https:∥doi.org/10.1186/1756-9966-29-63.
[4]陈小刚,罗鹏程,张青汉,等.小干扰RNA联合介导Livin和Survivin基因沉默对人肾癌细胞786-0增殖抑制及凋亡的影响[J].中华实验外科杂志,2014,31(6):1199-1201.
[5]周留正,陈兵海,孙浩,等.前列腺癌组织Livin表达与PTEN相关性研究[J].中华肿瘤防治杂志,2017,24(23):1651-1655.
[6]武亚运,张志,王进,等.Livin在胃癌中表达及与临床相关性的Meta分析[J].中国医药导报,2017,14(17):73-79.
[7]史芳涛,王庆锋.凋亡抑制蛋白基因Livin及肿瘤转移相关基因在肺癌组织中的表达及其与肺癌临床特征的关系分析[J].癌症进展,2017,15(7):794-797,805.
[8]HIRD A W,AQUILA B M,HENNESSY E J,et al.Small molecule inhibitor of apoptosis proteins antagonists:apatent review[J].Expert Opin Ther Pat,2015,25(7):755-774.
[9]CHEN L,REN G S,LI F,et al.Expression of Livin and vascular endothelial growth factor in different clinical stages of human esophageal carcinoma[J].World J Gastroenterol,2008,14(37):5749-5754.
[10]LIANG Y Z,FANG T Y,XU H G,et al.Expression of CD44v6and Livin in gastric cancer tissue[J].Chin Med J(Engl),2012,125(17):3161-3165.
[11]WANG Y,LI Y,ZHOU B,et al.Expression of the apoptosis inhibitor Livin in colorectal adenoma-carcinoma sequence:correlations with pathology and outcome[J].Tumour Biol,2014,35(12):11791-11798.
[12]YE L,SONG X,LI S,et al.Livin-αpromotes cell proliferation by regulating G1-S cell cycle transition in prostate cancer[J].Prostate,2011,71(1):42-51.
[13]OH D Y,BANG Y J.Adjuvant and neoadjuvant therapy for gastric cancer[J].Curr Treat Options Oncol,2013,14(3):311-320.
[14]张生军,常琦,刘勇峰,等.Livin通过AKt信号途径促进胃癌SGC7901细胞的上皮细胞间质转换[J].基础医学与临床,2016,36(5):586-589.
[15]CHO S B,LEE W S,PARK Y L,et al.Livin is associated with the invasive and oncogenic phenotypes of human hepatocellular carcinoma cells[J].Hepatol Res,2015,45(4):448-457.
[16]MYUNG D S,PARK Y L,CHUNG C Y,et al.Expression of Livin in colorectal cancer and its relationship to tumor cell behavior and prognosis[J/OL].PLoS One,2013,8(9):e73262[2018-04-20].https:∥doi.org/10.1371/journal.pone.0073262.
[2]杨媛,王建六,魏丽惠.Notch信号通路与子宫内膜癌关系的研究进展[J].中华妇产科杂志,2015,50(5):391-393.
[3]LIU C,WU X,LUO C,et al.Antisense oligonucleotide targeting Livin induces apoptosis of human bladder cancer cell via a mechanism involving caspase 3[J/OL].J Exp Clin Cancer Res,2010,29:63[2018-04-20].https:∥doi.org/10.1186/1756-9966-29-63.
[4]陈小刚,罗鹏程,张青汉,等.小干扰RNA联合介导Livin和Survivin基因沉默对人肾癌细胞786-0增殖抑制及凋亡的影响[J].中华实验外科杂志,2014,31(6):1199-1201.
[5]周留正,陈兵海,孙浩,等.前列腺癌组织Livin表达与PTEN相关性研究[J].中华肿瘤防治杂志,2017,24(23):1651-1655.
[6]武亚运,张志,王进,等.Livin在胃癌中表达及与临床相关性的Meta分析[J].中国医药导报,2017,14(17):73-79.
[7]史芳涛,王庆锋.凋亡抑制蛋白基因Livin及肿瘤转移相关基因在肺癌组织中的表达及其与肺癌临床特征的关系分析[J].癌症进展,2017,15(7):794-797,805.
[8]HIRD A W,AQUILA B M,HENNESSY E J,et al.Small molecule inhibitor of apoptosis proteins antagonists:apatent review[J].Expert Opin Ther Pat,2015,25(7):755-774.
[9]CHEN L,REN G S,LI F,et al.Expression of Livin and vascular endothelial growth factor in different clinical stages of human esophageal carcinoma[J].World J Gastroenterol,2008,14(37):5749-5754.
[10]LIANG Y Z,FANG T Y,XU H G,et al.Expression of CD44v6and Livin in gastric cancer tissue[J].Chin Med J(Engl),2012,125(17):3161-3165.
[11]WANG Y,LI Y,ZHOU B,et al.Expression of the apoptosis inhibitor Livin in colorectal adenoma-carcinoma sequence:correlations with pathology and outcome[J].Tumour Biol,2014,35(12):11791-11798.
[12]YE L,SONG X,LI S,et al.Livin-αpromotes cell proliferation by regulating G1-S cell cycle transition in prostate cancer[J].Prostate,2011,71(1):42-51.
[13]OH D Y,BANG Y J.Adjuvant and neoadjuvant therapy for gastric cancer[J].Curr Treat Options Oncol,2013,14(3):311-320.
[14]张生军,常琦,刘勇峰,等.Livin通过AKt信号途径促进胃癌SGC7901细胞的上皮细胞间质转换[J].基础医学与临床,2016,36(5):586-589.
[15]CHO S B,LEE W S,PARK Y L,et al.Livin is associated with the invasive and oncogenic phenotypes of human hepatocellular carcinoma cells[J].Hepatol Res,2015,45(4):448-457.
[16]MYUNG D S,PARK Y L,CHUNG C Y,et al.Expression of Livin in colorectal cancer and its relationship to tumor cell behavior and prognosis[J/OL].PLoS One,2013,8(9):e73262[2018-04-20].https:∥doi.org/10.1371/journal.pone.0073262.