去甲基化药物逆转结肠癌细胞株sw620/L-OHP耐药性的作用及机理研究  

Effect and Mechanism of DNMT Inhibitor on the Reversal of Multidrug Resistance in Human Colon Cancer Cell Line sw620/L-OHP

作　　者：邓茜[1] 李志平[1] 黄春梅[1] 刘桢[1] 钟仁明[2] 李爱军[2] 毕锋[3] 唐秋琳[3] 

机构地区：[1]四川大学华西医院肿瘤中心腹部肿瘤科,成都610041 [2]四川大学华西医院肿瘤放射治疗中心,成都610041 [3]四川大学华西医院信号转导与分子靶向治疗实验室,成都610041

出　　处：《四川大学学报（医学版）》2010年第6期975-979,共5页Journal of Sichuan University(Medical Sciences)

摘　　要：目的探讨甲基转移酶抑制剂5-氮杂-2’-脱氧胞苷(5-aza-dC)对结肠癌奥沙利铂(L-OHP)耐药细胞株sw620/L-OHP耐药性的逆转作用及其作用机制。方法应用5-aza-dC作用于结肠癌细胞sw620与其L-OHP耐药细胞sw620/L-OHP,CCK-8法检测其逆转耐药作用;应用Westernblot检测5-aza-dC使用前后sw620/L-OHP细胞中BNIP3基因及耐药基因编码的P糖蛋白(P-GP)、多药耐药相关蛋白(MRP)表达水平的变化;应用同位素微量示踪法检测5-aza-dC使用前后sw620/L-OHP细胞中甲基转移酶(DNMT)的活性。结果 1.4μmol/L5-aza-dC能使L-OHP对sw620/L-OHP细胞IC50由(0.335±0.043)μg/mL降到(0.069±0.023)μg/mL,逆转耐药倍数为8.26倍(P<0.001);5-aza-dC能使sw620/L-OHP细胞中P-GP、MRP蛋白表达增加(P<0.05),但增加程度与5-aza-dC剂量变化无关(P>0.05);另外5-aza-dC还能使表达低下的BNIP3蛋白重新表达,且其表达水平随5-aza-dC浓度的增加而增加(P<0.001);5-aza-dC能够降低sw620/L-OHP细胞中DNMT的活性(P<0.001)。结论 5-aza-dC通过降低DNMT活性来上调BNIP3的表达,并协同某种机制抑制P-GP、MRP的进一步表达,从而逆转sw620/L-OHP细胞耐药性。Objective To explore whether demethylation inhibitor 5-aza-2 ＇-deoxycytidine （5-aza-dC） could reverse multidrug resistacne （MDR） of human colon cancer and its impacts on the expressions of Bel-2/adenovirus E1B 19 KDa protein interacting protein 3 （BNIP3）, P-glyeoprotein （P-GP） and muhidrug resistanceassoeiated protein （MRP）, as well as the activity of DNA methyhransferase （DNMT）. Methods Oxaliplatin （L-OHP） resistant human colon carcinoma cells （sw620/L-OHP） and its parental cells （sw620） were used to determine the effect of 5-aza-dC. CCK-8 assay was adopted to evaluate the cytotoxicity of 5-aza-dC. Western blot was performed to observe the expressions of BNIP3, P-GP and MRP. 3 H microassay was used to detect the activity of DNMT. Results After the addition of 1.4 μmol/L 5-aza-dC, the 50% inhibitory concentration （IC50） of L-OHP to sw620/ L-OHP cells was decreased from （0. 335± 0. 043） μg/mL to （0. 069 ± 0. 023） μg/mL, and the chemosensitivity increased 8. 26 times （P〈0. 001）. Western blot analysis showed that the expression of BNIP3 was up-regulated by 5-aza-dC in a concentration-dependent manner （P〈0. 001）, and P-GP and MRP expression were also significantly changed （P〈0.05） but not in a concentration-dependent manner （P〉0.05）. DNMT activity was down-rugulated by 5-aza-dC in sw620/L-OHP ceils （P〈0. 001）. Conclusions 5-aza-dC could reversed multi drug resistance of sw620/L-OHP, which may due to the demethylation to upregulate BNIP3 expression, and inhibite the expression of resistance-associated protein P-GP and MRP furtherly by some unknown mechanism.

关 键 词：BNIP3 P糖蛋白 多药耐药相关蛋白 甲基转移酶5-氮杂-2’-脱氧胞苷 逆转耐药 

分 类 号：R735.3[医药卫生—肿瘤]