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目的研究环孢素A(CsA)和苯基棕榈酰胺吗啡丙醇(PPMP)联合应用对人白血病多药耐药细胞株K562/AO2的葡萄糖神经酰胺合酶(GCS)基因表达的影响及对白血病多药耐药的逆转作用,探索逆转白血病细胞耐药的策略。方法应用Alamar BlueTM多功能细胞染色法测定阿霉素(ADR)对K562和K562/AO2细胞的半数抑制浓度(IC50)及判断CsA的非细胞毒性剂量;采用RT-PCR法检测GCS基因和mdr1基因的mRNA表达。结果CsA浓度低于4.5μg/ml对K562/AO2细胞无细胞毒性。ADR对K562和K562/AO2细胞的IC50分别为(0.84±0.04)μg/ml和(89.24±1.27)μg/ml;当1μg/ml CsA和25μmol/L PPMP单用或联合作用于K562/AO2细胞时,ADR的IC50分别为(7.81±0.74)(、17.49±0.53)(、2.82±0.07)μg/ml;而低浓度CsA(0.01μg/ml)和PPMP(10μmol/l)联合作用时IC50为(16.08±1.25)μg/ml。CsA联合PPMP可明显降低GCS基因的mRNA表达,低浓度CsA和PPMP联用也有此作用。结论CsA联合PPMP可通过抑制K562/AO2细胞的GCS基因mRNA表达有效逆转其耐药,低浓度CsA联合PPMP在逆转其耐药的同时降低了细胞毒性。
Objective To investigate the effects of cyclosporin A (CsA) and phenyl palmitamide (PPMP) on the gene expression of glucosylceramide synthase (GCS) in human leukemia multidrug-resistant cell line K562 / AO2 and the effect on leukemia Multidrug resistance reversal effect, explore the reversal of leukemia cell resistance strategy. Methods The half inhibitory concentration (IC50) of adriamycin (ADR) on K562 and K562 / AO2 cells and the non-cytotoxic dose of CsA were determined by Alamar BlueTM multi-function cell staining. The GCS and mdr1 genes were detected by RT- MRNA expression. Results CsA concentration of less than 4.5μg / ml of K562 / AO2 cells without cytotoxicity. The IC50 of ADR to K562 and K562 / AO2 cells were (0.84 ± 0.04) μg / ml and (89.24 ± 1.27) μg / ml respectively. When K562 / AO2 cells were treated with 1μg / ml CsA and 25μmol / , The IC50 of ADR was (7.81 ± 0.74) (, 17.49 ± 0.53) (2.82 ± 0.07) μg / ml, while the IC50 of ADR was (0.01μg / ml) and PPMP (16.08 ± 1.25) μg / ml. CsA combined with PPMP can significantly reduce the mRNA expression of GCS gene, low concentrations of CsA and PPMP combination also has this effect. Conclusion CsA combined with PPMP can effectively reverse the drug resistance by inhibiting the expression of GCS mRNA in K562 / AO2 cells. Low concentration of CsA combined with PPMP can reverse the drug resistance and reduce the cytotoxicity.