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目的探讨核转录因子NF-κB及上皮特异性黏蛋白MUC1基因及蛋白参与膀胱癌细胞耐药形成的机理。方法免疫组织化学SP法检测NF-κB p65和MUC1蛋白在B IU-87、T-24及耐药株B IU-87/A细胞中的表达。原位杂交技术检测NF-κB p65的mRNA在3种细胞质中的表达。流式细胞仪检测3种细胞在阿霉素、长春新碱、顺铂3种化疗药物作用48 h后的凋亡率。结果NF-κB p65蛋白在B IU-87、T-24细胞质中均有阳性表达,细胞核几乎无表达,在B IU-87/A耐药株的胞质、胞核中均有阳性表达。3组细胞平均吸光度值差异无统计学意义(P>0.05)。MUC1在B IU-87、T-24细胞膜、胞质中均有弱阳性表达,在B IU-87/A的胞膜、胞质中均有强阳性表达,两者平均吸光度值差异有统计学意义(P<0.01)。3种细胞中NF-κB p65 mRNA表达均在胞质,核无表达,且3组细胞平均吸光度值差异无统计学意义(P>0.05)。3种细胞自发凋亡率分别为1.15%、1.40%、0.90%,3种化疗药物作用48 h后,B IU-87细胞凋亡率分别为45.69%、47.70%、44.50%,T-24细胞凋亡率分别为43.79%、46.17%、44.50%,用药前后差异有统计学意义(P<0.01)。两种细胞之间及3种化疗药之间差异无统计学意义(P>0.05)。B IU-87/A在阿霉素、长春新碱作用48 h后的凋亡率分别为19.88%、21.41%,用药前后差异有统计学意义(P<0.05)。结论NF-κB是膀胱癌细胞B IU-87/A耐药形成的中心环节,B IU-87/A耐药形成中NF-κB过度活化导致MUC1高表达可能是另一途径。
Objective To investigate the mechanism of nuclear transcription factor NF-κB and epithelial-specific mucin MUC1 gene and protein involved in the drug resistance of bladder cancer cells. Methods The expression of NF-κB p65 and MUC1 protein in B IU-87, T-24 and drug resistant B IU-87 / A cells were detected by immunohistochemical SP method. In situ hybridization was used to detect the mRNA expression of NF-κB p65 in three kinds of cytoplasm. Flow cytometry was used to detect the apoptotic rates of three kinds of cells after 48 h treatment with doxorubicin, vincristine and cisplatin. Results The expression of NF-κB p65 protein was positive in both B IU-87 and T-24 cytoplasm. The expression of NF-κB p65 protein in the cytoplasm and nucleus of B IU-87 / A was almost undetectable. There was no significant difference in mean absorbance between the three groups (P> 0.05). MUC1 was weakly positive in both B IU-87 and T-24 cell membranes and in the cytoplasm, strongly positive in the membrane and cytoplasm of B IU-87 / A, and the difference between the two was statistically significant Significance (P <0.01). The expressions of NF-κB p65 mRNA in the three kinds of cells were all expressed in the cytoplasm and nucleus, and there was no significant difference between the three groups (P> 0.05). The apoptotic rates of B IU-87 cells were 45.69%, 47.70%, 44.50%, respectively. The apoptosis rate of T-24 cells was 1.15%, 1.40% and 0.90% The apoptotic rates were 43.79%, 46.17% and 44.50%, respectively, with significant difference before and after treatment (P <0.01). There was no significant difference between the two kinds of cells and the three chemotherapeutic drugs (P> 0.05). The apoptosis rate of B IU-87 / A treated with doxorubicin and vincristine for 48 h was 19.88% and 21.41%, respectively. The difference was statistically significant (P <0.05) before and after treatment. Conclusions NF-κB is the central part of B IU-87 / A resistance in bladder cancer cells. Over-activation of NF-κB in B IU-87 / A may lead to high MUC1 expression.