人结直肠癌耐药细胞株HT-29/CPT-11的构建及其生物学特性探讨-东南大学学报医学版

人结直肠癌耐药细胞株HT-29/CPT-11的构建及其生物学特性探讨

单位：1. 南京医科大学,江苏南京 210029;
2. 南京医科大学附属淮安第一人民医院肿瘤内科,江苏淮安 223300

分类号：R-33; R73.3

出版年·卷·期（页码）：2012·31·第一期（31-35）

摘要：

目的:建立耐伊立替康(CPT-11)的人结直肠癌细胞株(HT-29/CPT-11),并研究其生物学特性。方法:采用药物浓度梯度递增间歇诱导法建立人结直肠癌耐药株HT-29/CPT-11;CCK-8法检测CPT-11对亲本细胞HT-29和耐药细胞株HT-29/CPT-11细胞的半数抑制率(IC₅₀);绘制细胞生长曲线,并计算细胞倍增时间;流式细胞术分析细胞周期分布;RT-PCR检测HT-29/CPT-11和HT-29细胞MDR-1 mRNA的表达。结果:(1) 成功建立人结直肠癌耐药细胞株HT-29/CPT-11,耐药指数为6.51。(2) HT-29/CPT-11耐药细胞株倍增时间较原代细胞HT-29延长。(3) 流式细胞术检测细胞周期结果显示,耐药细胞株HT-29/CPT-11 G₁期细胞数目增加,S期细胞数目减少(P<0.05)。(4) 耐药细胞HT-29/CPT-11的MDR-1 mRNA表达水平明显高于亲本细胞株HT-29,分别为1.086±0.054和0.416±0.02(P<0.05)。结论:成功构建了人结直肠癌耐药细胞株HT-29/CPT-11,为下一步研究耐药机制奠定实验基础。

Objective: To establish a irinotecan(CPT-11) resistance human colorectal cancer cell line and study its biological characteristics,Methods: HT-29/CPT-11 cell line was established by stepwise selection in increasing dose of CPT-11. IC₅₀ of CPT-11 in HT-29 and HT-29/CPT-11 cell lines was detected by CCK-8. Cell growth curve was drawed and the doubling time was accounted. The cell cycle distribution was determined by flow cytometry. The expression levels of MDR-1 mRNA in HT-29 and HT-29/CPT-11 cell lines were examined by RT-PCR. Results: (1) The CPT-11 resistance human colorectal cancer cell line HT-29/CPT-11 was successfully established, the resistance index was 6.51. (2) The doubling time of HT-29/CPT-11 obviously longer than that of HT-29 cell[(41.29±1.69)h vs (27.12±2.73)h, P<0.05]. (3) Flow cytometry demonstrated that HT-29/CPT-11 cells of phase G1 were increased and S phase were reduced. (4) The expression level of MDR-1 mRNA in HT-29/CPT-11 cell line was higher than that of HT-29(1.086±0.054 vs 0.416±0.02, P<0.05). Conclusion: We have successfully established CPT-11 resistance cell line HT-29/CPT-11, it lays a foundation for the study of resistance mechanism.

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