KBV200细胞多药耐药与蛋白激酶C表达上调的关系

目的 研究KBV200多药耐药细胞蛋白激酶C(PKC)的表达,探讨PKC的表达上调与肿瘤多药耐药的关系。方法 KBV200细胞分对照组和佛波酯(PMA)组,PMA组应用200nmol/L的PMA预孵育细胞,而对照组不用。³²P掺入法测定多药耐药KBV200细胞株的PKC活性,通过Westernblotting检测PKC各亚型表达和亚细胞分布。用MTT法检测细胞耐药性。结果 PMA预孵育可提高KBV200细胞的PKC总活性和膜组分PKC活性,降低浆组分PKC活性(P<0.01)。PMA预孵育使膜组分PKCα表达增加,浆组分PKCα表达降低,膜组分PKCβ无明显变化,浆组分PKCβ的表达稍增强。PMA可升高长春新碱、阿霉素对KBV200细胞的IC₅₀值(P<0.01)。结论 PMA使KBV200细胞耐药性增加,可能与PKC表达上调有关。

Correlation of multidrug resistance with up-regulation of protein kinase C expression in KBV200 cells

Objective To investigate protein kinase C (PKC) expression and its association with multidrug resistance (MDR) in KBV200 cells. Methods KBV200 cells were preincubated with PKC activator phorbol-12-myristate-13-acetate (PMA, 200nmol/L) and PKC activity was assayed by measurement of peptide substrate 32p incorporation from γ-32p]ATP, with the cells without PMA preincubation serving as the control. Western blotting was performed for assessing the expression of PKC isoform, and the cell inhibition rate was evaluated by MTT assay. Results PMA preincubation of the cells significantly enhanced the activity of the total PKC and the membrane fraction, but lowered the PKC activity of the cytosol fraction, as compared with the cells without PMA treatment (P＜0.01). PKCα expression was upregulated in the membrane fraction and down-regulated in the cytosol fraction in KBV200 cells after PMA preincubation. PKCβ expression was slightly elevated in the cytosol fraction but exhibited no obvious changes in the membrane fraction after PMA pretreatment of the cells. The values of IC50 of vincristine and adriamycin in PMA-treated cells were increased to 2275.5 nmol/L and 233.25 nmol/L, respectively (P＜0.01).Conclusion PMA can increase the multidrug resistance of KBV200 cells, which suggests the possible involvement of PKC in the mechanism ofmultidrug resistance of tumor cells.