川芎嗪对转基因多药耐药细胞K562/MDR耐药性的逆转作用  

Effect of tetramethylpyrazine on reversing multidrug resistance of K562/MDR cells with gene transfer

作　　者：马海英[1] 赵瑾瑶[1] 金伟[1] 孔力[1] 

机构地区：[1]大连医科大学组织胚胎学教研室,辽宁大连116044

出　　处：《吉林大学学报（医学版）》2009年第4期599-603,共5页Journal of Jilin University：Medicine Edition

基　　金：国家自然科学基金资助课题(39170853)

摘　　要：目的：研究川芎嗪（TMP）与环孢菌素A（CsA）单独及联合逆转人红白血病多药耐药细胞K562/MDR的多药耐药性（MDR）,并进一步探讨其耐药逆转机制。方法：分别以倍比稀释后不同浓度的TMP（506 400 mg·L^-1）和CsA（0.5256 mg·L^-1）作用于体外培养的K562/MDR细胞72 h,采用MTT法检测细胞生长率,确定TMP和CsA的非细胞毒性剂量;采用非细胞毒性剂量,实验分为5组：G1组（TMP＋ADM＋K562/MDR）、G2组（CsA＋ADM＋K562/MDR）、G3组（TMP＋CsA＋ADM＋K562/MDR）、阴性对照组（以K562/S代替K562/MDR）、空白对照组（以1640培养基代替药物）,检测各组细胞生长抑制50%的ADM浓度,即IC50,计算耐药倍数和逆转倍数;利用荧光分光光度法检测各组细胞内ADM浓度;流式细胞术检测跨膜糖蛋白P-gp的表达情况。结果：TMP及CsA的非细胞毒性剂量分别为320和2.0 mg·L^-1,K562/MDR细胞的耐药倍数为19.2倍;TMP及CsA均能降低K562/MDR细胞的耐药性,逆转倍数分别为5.2及9.6倍,并且两者联合应用,逆转倍数为15.6倍;TMP和CsA单独及联合应用均能明显增加K562/MDR细胞内ADM浓度;TMP及CsA单独应用均能降低K562/MDR细胞P-gp的表达,并且两者联合应用使K562/MDR细胞P-gp的表达率明显降低,与阴性对照组比较差异有显著性（P〈0.01）。结论：TMP和CsA单独及联合应用可逆转K562/MDR细胞对化疗药物ADM的MDR,且两者具有协同作用。其逆转机制可能为降低P-gp水平,升高细胞内ADM浓度,增强对ADM的敏感性,从而发挥治疗肿瘤的作用。Objective To investigate the reversal effects of tetramethylpyrazine（TMP） and cyclosporin A（CsA） on multidrug resistance（MDR） of human erythroleukemia cells with gene transfer K562/MDR and to discuss the possible correlative mechanism.Methods K562/MDR cells in culture medium were treated with TMP（50—6 400 mg·L^-1） and CsA（0.5—256 mg·L^-1） respectively.The growth rates of these cells were measured by MTT assay.Non-cytotoxic doses of TMP and CsA were determined.There were 5 growps in the experiment：G1（TMP＋ADM＋K562/MDR）,G2（CsA＋ADM＋K562/MDR）,G3（TMP＋CsA＋ADM＋K562/MDR）,negative control（K562/MDR subsitituted by K562/S）,control（drugs subsitituted by 1640 culture medium）,the inhibitory effects of adriamycin（ADM） used alone or in combination with TMP or/and CsA on the proliferation of K562/S and K562/MDR cells were observed by resisting fold and reversal folds.The effects of TMP and CsA on ADM accumulation in K562/S and K562/MDR cells were analyzed by fluorospectrophotometry.The protein level of P-glycoprotein（P-gp） was detected by flow cytometry（FCM）.Results The non-cytotoxic doses of TMP and CsA were 320 and 2.0 mg·L^-1,respectively.The resistance of K562/MDR cells to ADM was 19.2 folds of that of K562/S cells.When TMP and CsA were added along with ADM to the K562/MDR culture,the reversal folds were 5.2 and 9.6.When TMP in combination with CsA was added along with ADM to the K562/MDR culture,the reversal folds were 15.6.When various concentrations ADM were added to the K562/MDR culture,TMP and CsA could increase the intracellular accumulation of ADM.The effect of TMP in combination with CsA was more evident.Compared with control,TMP and CsA could inhibit the overexpression of P-gp protein（P〈0.01）.Conclusion The single or combinated application of TMP and CsA could increase the drug concentration of K562/MDR cells and reverse the MDR of tumor cells.Its activity may be related to the inhibition of P-gp protein overexpression.

关 键 词：川芎嗪 环孢菌素A 多药耐药逆转 K562/MDR细胞 

分 类 号：R737.9[医药卫生—肿瘤]