膜联蛋白A1基因的表达与胆囊癌临床病理的关系及调控信号转导与转录因子3信号通路对胆囊癌细胞增殖凋亡的影响

The relationship between the expression of ANXA1 gene and the clinical pathology of gallbladder carcinoma and the effect of STAT3 signaling pathway on the proliferation and apoptosis of gallbladder cancer cells

目的探讨膜联蛋白A1(ANXA1)基因的表达与胆囊癌临床病理的关系及调控信号转导与转录因子3(STAT3)信号通路对胆囊癌细胞增殖凋亡的影响及其机制。方法应用免疫组化法检测100例胆囊癌组织中ANXA1基因的阳性表达,分析其与临床病理特征的关系。培养人胆囊癌GBC-SD细胞,分别将ANXA1小干扰RNA(ANXA1-siRNA)和阴性对照(siRNA-NC)转染细胞,以空脂质体转染的细胞作为对照组,蛋白质印迹法(Western blot)检测转染效果;细胞转染后培养48h,细胞计数试剂盒(CCK8)检测细胞增殖;流式细胞仪检测细胞凋亡;蛋白质印迹法检测活化的含半胱氨酸的天冬氨酸蛋白水解酶3(Cleaved Caspase-3)、STAT3、磷酸化的信号转导与转录因子3(p-STAT3)蛋白表达。结果 ANXA1基因在胆囊癌中的阳性表达率显著高于癌旁组织(69.0%比7.5%,χ2=43.261,P<0.01);ANXA1在胆囊癌中的表达与性别和年龄不相关(P>0.05),与肿瘤的分化程度、临床分期、淋巴转移显著相关(P<0.01)。siRNA-NC组中ANXA1、Cleaved caspase3、STAT3、p-STAT3蛋白表达、细胞存活率、细胞凋亡率与对照组比较差异无统计学意义(P>0.05);ANXA1-siRNA组ANXA1蛋白表达、细胞存活率及p-STAT3蛋白表达显著低于对照组(P<0.01),细胞凋亡率及Cleaved caspase3蛋白表达显著高于对照组(P<0.01),STAT3蛋白表达在三个组之间差异无统计学意义(P>0.05)。结论 ANXA1基因的表达参与了胆囊癌的生长、分化和迁移过程,下调ANXA1的表达能抑制人胆囊癌GBC-SD细胞的增殖,并通过下调Cleaved caspase3蛋白促进细胞凋亡,其机制与STAT3信号通路的调控有关。

Objective To investigate the relationship between the expression of annexin A 1 （ANXA1） gene and the clinical pathology of gallbladder carcinoma and the effect of signal transduction and transcription factor 3 （STAT3） signaling pathway on the proliferation and apoptosis of gallbladder cancer cells .Methods The expression of ANXA1 in 100 cases of gallbladder carcinoma was detected by immunohistochemical method,and the relationship between the expression of the ANXA 1 gene and the clinical pathological characteris -tics was analyzed.Human gallbladder carcinoma GBC -SD cells were cultured,ANXA1 small interfering RNA （ANXA1-siRNA） and negative control （siRNA-NC） were transfected respectively into cells ,and with empty liposomes transfected cells as the control group , Western blot was used to detect the transfection effect ;cells were cultured for 48h,cell proliferation was detected by CCK -8;cell apopto-sis was detected by flow cytometry;the expressions of Cleaved Caspase3,p-STAT3,STAT3 protein were detected by Western blot . Results The positive expression of ANXA1 gene in gallbladder carcinoma was significantly higher than those in the adjacent tissues （69.0% vs 7.5%,χ2 =43.261,P 〈0.01）;the expression of ANXA1 in gallbladder carcinoma was not related to gender and age （P 〉0.05）,and the degree of tumor differentiation ,clinical stage,lymph node metastasis was significantly correlated （P 〈0.01）;the ex-pression of ANXA1,Cleaved Caspase3,STAT3,p-STAT3 protein ,cell survival rate,and the apoptosis rate in siRNA-NC group were not significantly different compared with control group （P 〉0.05） and the protein expression of p-STAT3 in ANXA1-siRNA group were significantly lower than the control group （P 〈0.01）,the apoptosis rate and the expression of Cleaved Caspase 3 protein were signifi-cantly higher than the control group （P 〈0.01）;the expression of STAT3 protein was not significantly different among the three group（P 〉0.05）.Conclusions The expression of ANXA1 gene is involved in the growth,differentiation and migration process of gallblad -der carcinoma.Down-regulating the expression of ANXA1 can inhibit the proliferation of GBC-SD cells in human gallbladder carcinoma , and promote cell apoptosis by down -regulating Cleaved Caspase3 protein,and its mechanism is related to the regulation of STAT3 signa-ling pathway.