环腺苷酸对人白血病多药耐药K562/ADM细胞的诱导分化作用

Cyclic Adenosine Monophosphate Induced Differentiation of Human Leukemia K562/ADM Cells with Multidrug Resistance

目的:研究环腺苷酸(cyclicadenosinemonophosphate,cAMP)对人白血病多药耐药K562/ADM细胞的诱导分化作用。方法:K562/ADM细胞经0.25～2.0mmol/LcAMP处理后,MTT法检测细胞增殖活性,流式细胞术(flowcytometryFCM)检测细胞周期和P-糖蛋白(PglycoproteinPgp)的表达,免疫细胞化学法检测细胞周期蛋白cyclinD1和转录因子E2F的表达;同时观察细胞形态学变化和血红蛋白合成功能。结果:cAMP呈浓度和时间依赖性地抑制K562/ADM细胞的增殖(P<0.01),细胞形态学上出现红系细胞的形态特征;被诱导的细胞能够合成血红蛋白、cyclinD1和E2F的表达减低。cAMP(0.25mmol/L和1.0mmol/L)诱导24h,多数细胞被阻滞在G1期,S期和G2+M期细胞显著降低;诱导72h后K562/ADM细胞Pgp表达的阳性率无明显变化(99.8%～99.9%),但Pgp的表达量显著增加,平均荧光强度分别增高1.24倍和1.28倍。结论:K562/ADM细胞仍保留了分化成熟的潜能,可被cAMP诱导向正常血细胞分化,但在分化诱导过程中,化学诱导剂可导致耐药细胞发生Pgp的应激性表达增强而可能阻抑后续的化学治疗,或对诱导剂产生耐受性。

Objective: To investigate the differentiation of multidrug resistant human leukemia cell line K562/ADM induced by cyclic adenosine monophosphate (cyclic AMP, cAMP). Methods: After treatment of K562/ADM cells with cAMP(0.25-2.0mmol/L), the proliferating activity was determined with a MTT colorimetric assay, the cell cycle progression and expression of P glycoprotein (P gp) were analyzed by flow cytometry (FCM), the production of cyclin D1 and transcription factor E2F was examined with immunocytochemical methods, cytomorphology and synthesis of hemoglobin were also observed. Results: cAMP treatment of K562/ADM cells resulted in a concentration and time dependent growth inhibition (P< 0.01). The cells showed some morphological features of normal erythroid cells, appeared to synthesize hemoglobin and decrease the production of cyclin D1 and E2F. Treatment with cAMP (0.5 mmol/L and 1.0 mmol/L) for 24 hours, most of K562/ADM cells were arrested in G1 phase, and the prolongation of phase G1 caused a markedly decrease of the cells in phase S and phase G2+M. While the number of P gp positive cells were similar in either untreated or cAMP treated K562/ADM cells(99.8%-99.9%), but cAMP enhanced the P gp expressing capacity in K562/ADM cells, the mean fluorescence intensity (MFI) of the P gp+ cells was increased by 1.24 times and 1.28 times respectively. Conclusions: Drug resistant K562/ADM cells still retain terminal differentiating potential, and differentiation can be induced by cAMP. Exposure of the drug resistant cells to chemical inducers (cAMP, etc) results in stress activated expressing enhancement of P glycoprotein which may block the late chemotherapy or cause the tolerance of the cells to those inducers.