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目的:通过检测阿霉素诱导人胆囊癌细胞多药耐药过程中糖基化神经酰胺合成酶(GCSmRNA)、蛋白及caspase3的变化情况,探讨神经酰胺(ceramide)代谢中起关键作用的酶GCS在人胆囊癌多药耐药产生过程中的作用。方法:阿霉素200μg/L连续作用人胆囊癌细胞株GBC-SD12周,采用MTT法、RT-PCR、Western blotting分别检测GBC-SD、GBC-SD(4周)、GBC-SD(12周)时的细胞毒性、GCSmRNA、蛋白的变化情况,荧光分光光度法检测此过程中caspase3的变化。结果:诱导12周时,细胞耐药倍数增加3·8倍,GCSmRNA、蛋白表达明显上调(P<0·01)。3种细胞的增殖同阿霉素浓度呈负相关,GBC-SD、GBC-SD(4周)增殖同作用时间呈负相关,而GBC-SD(12周)细胞同作用时间无关。GCS的表达同caspase3的活性呈负相关。结论:阿霉素通过诱导细胞凋亡抑制人胆囊癌细胞增殖,人胆囊癌细胞多药耐药的产生同GCS mRNA、蛋白表达的上调相关,GCS的表达上调抑制了caspase3的激活,GCS可能是诱导胆囊癌多药耐药产生的机制之一。
OBJECTIVE: To detect the changes of glycosylated ceramide synthetase (GCSmRNA), caspase3 and protein during the multidrug resistance of human gallbladder carcinoma cells induced by doxorubicin, and to explore the role of GCS (key enzyme) in ceramide metabolism The role of multidrug resistance in human gallbladder cancer. METHODS: Human gallbladder carcinoma cell line GBC-SD was treated with 200μg / L doxorubicin continuously for 12 weeks. The expressions of GBC-SD, GBC-SD and GBC-SD were detected by MTT, RT- ) Cytotoxicity, GCS mRNA, protein changes, fluorescence spectrophotometry to detect changes in the process of caspase3. Results: After 12 weeks of induction, the cell multiplicity of resistance increased by 3.8 times and the expression of GCS mRNA and protein were significantly increased (P <0.01). The proliferation of three kinds of cells was negatively correlated with the concentration of doxorubicin. The proliferation of GBC-SD and GBC-SD (4 weeks) was negatively correlated with the duration of action, while the cells with GBC-SD (12 weeks) had no correlation with the duration of action. The expression of GCS was negatively correlated with the activity of caspase3. CONCLUSION: Doxorubicin inhibits the proliferation of human gallbladder carcinoma cells through induction of apoptosis. The multidrug resistance of human gallbladder carcinoma cells is related to the up-regulation of GCS mRNA and protein expression. The up-regulation of GCS inhibits the activation of caspase 3. GCS may be Induction of gallbladder carcinoma multidrug resistance one of the mechanisms.