大黄素对K562/ADM细胞阿霉素耐药的抑制作用

探讨大黄素抑制人慢性粒细胞白血病细胞耐阿霉素变异株K562/ADM的药效学及作用机制。以维拉帕米为阳性药,阿霉素为工具药,采用MTT法测定大黄素与阿霉素合用对细胞增殖的抑制作用;使用流式细胞仪考察了大黄素对于阿霉素诱导K562/ADM细胞周期阻滞和凋亡的促进作用;并采用Western blot法检测了大黄素对P-gp、Bcr-Abl和STAT5蛋白表达的影响;进一步通过P-gp单克隆藻红蛋白键合UIC2抗体结合实验探讨了大黄素是否是P-gp的底物。实验结果显示,大黄素与阿霉素合用可显著抑制K562/ADM的增殖,并呈量效关系;可促进阿霉素诱导的K562/ADM细胞G₂/M期周期阻滞和细胞凋亡;不同剂量的大黄素可有效抑制P-gp、Bcr-Abl、STAT5蛋白表达及磷酸化;UIC2抗体结合实验结果则表明大黄素可能不是P-gp的底物。因此,大黄素能够在体外有效抑制K562/ADM阿霉素耐药,其机制可能不是通过底物竞争性抑制作用,而是与大黄素直接降低P-gp、Bcr-Abl及STAT5蛋白表达和磷酸化有关。     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

目的 观察白血病药物敏感和耐受细胞中白血病干细胞(ISC)、耐药蛋白表达和耐药性的关系.方法 以白血病多药耐药株K562/ADM细胞及其亲本K562细胞为模型,MTT比色法测定细胞的药物耐受性;流式细胞术检测细胞免疫标志、P-糖蛋白(P-gp)和乳腺癌耐药蛋白(BCRP)的表达;甲基纤维素集落形成法检测细胞的自我更新和增殖潜能.结果 K562/ADM细胞对阿霉素、柔红霉素和鬼臼乙叉苷高度耐受.K562/ADM细胞中CD34+、CD123+和CD34+CD38-细胞含量均显著高于K562细胞,LSC细胞相对含量是K562细胞的4.12倍;共表达P-gP和BCRP的K562/ADM细胞比K562细胞高11.25倍,其中CD34+ CD38- CD123+ BCRP+和CD34+ CD38- P-gp+ BCRP+细胞数量分别为K562细胞的3.66倍和11.37倍.K562/ADM细胞集落形成率是K562细胞的4.17倍,与LSC含量相当.结论 白血病K562/ADM耐药细胞中存在的高表达耐药蛋白的耐药性LSC是其多药物耐受性的根源.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

白血病K562/ADM细胞耐药性与白血病干细胞及耐药蛋白表达的关系

Objective To investigate the drug resistance, proportion of leukemia stem cells (LSC) and expression of P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) in drug-sensitive and multidrug-resistant leukemia cell population. Methods The multidrug-resistant leukemia K562/ADM cell and its parental K562 cell were used as the model cells. The drug sensitivity was tested with a MTI" assay. Flow cytometry was employed to detect the immunophenotype of stem celia and the expression of P-gp and BCRP. The serf-renewal and proliferating potential were examined with methylcellulose colony-formlng unit assay. Results K562/ADM cells were highly resistant to adriamycin, daunorubincin and etoposide. The amount of CD34+, CD123+ and CD34+ CD38- cells in K562/ADM cells was much higher than that in K562 cells, and the proportion of CD34+ CD38-CD123+ cells(LSC) in K562/ADM cells was (5.23 ± 0. 21) % versus (1.27 ± 0. 17) % in K562 cells, which was 4. 12-fold higher than that in K562 cells. Both P-gp and BCRP were overexpressed in K562/ADM cells relative to K562 cells, and the K562/ADM cells co-expressing P-gp and BCRP were 11.25-fold higher over K562 cells. The proportion of CD34+ CD38- CD123+BCRP+ and CD34+ CD38- P-gp+ BCRP+cells in K562/ADM cells were (4. 13±0.40)% and (5. 80 ± 1.19)% respectively, which were 3.66- and 11. 37-fold higher than the same cells in K562 cells [(1.13 ± 0. 15) % and (0. 51 ± 0. 01) %]. Furthermore, drug-resistant K562/ADM cells displayed 4. 17-time greater colony-forming ability over the parent K562 cells, corresponding to the proportion of LSCs in K562/ADM cells. Conclusions The ABC transporters-overexpressing LSC population exists in drug-resistant leukemic K562/ADM cells relative to drug-sensitive K562 cells, and the drug-resistant LSCs may be the source of chemotherapeutic resistance of leukemia.     

中药功劳木逆转K562/ADM,MCF7/ADM细胞的作用

目的 :研究中药功劳木 (gonglaomu ,GLM)提取物在体外对白血病耐药细胞K5 6 2 /ADM和乳腺癌耐药细胞MCF7/ADM逆转MDR1的作用 ,以便寻找和确定逆转肿瘤细胞多药耐药有效的中药新药 .方法 :应用MTT法、台盼蓝拒染试验法测定应用GLM提取物后ADM对K5 6 2 /ADM ,MCF7/ADM细胞的细胞毒作用 ,应用Rh1 2 3荧光技术检测应用GLM提取物后对耐药细胞内药物浓度的影响 .结果 :应用GLM提取物后MTT法显示ADM对K5 6 2 /ADM ,MCF7/ADM细胞的抑制率高于未用GLM组 ,且有显著性差异 (P <0 .0 1 ) .台盼蓝拒染试验测定显示应用GLM提取物后ADM对K5 6 2 /ADM ,MCF7/ADM活细胞率为 34.5 %和 38.5 % ,低于未用GLM组的 6 8.2 %和77.4 % ,且有显著性差异 (P <0 .0 1 ) .逆转倍数K5 6 2 /ADM ,MCF7/ADM细胞分别为 6 .2 8和 5 .78.Rh1 2 3荧光技术显示随着GLM提取物浓度的提高K5 6 2 /ADM细胞内药物浓度随之增高 .结论 :GLM提取物可提高了ADM对耐药细胞的细胞毒作用 ,增加了耐药细胞内药物浓度 ,且呈剂量依赖性     

多药耐药相关蛋白-2在白血病耐药细胞株K562/A02的表达

目的:研究多药耐药相关蛋白-2（MRP-2）与白血病细胞耐药的关系,并探讨其在白血病多药耐药中的意义。方法：应用RT-PCR方法检测白血病细胞株K562与耐药细胞株K562/A02中MRP-2 mRNA的差异表达,然后将MRP-2反义及正义寡核苷酸片段分别用脂质体转染至耐药细胞株K562/A02,采用MTT法、流式细胞术及RT-PCR法检测转染后细胞内阿霉素的荧光强度、耐药指数及MRP-2 mRNA的表达情况。结果：耐药细胞株K562/A02中MRP-2 mRNA表达水平明显高于白血病细胞株K562（P<0.05）。MRP-2反义寡核苷酸片段转染后耐药细胞株K562/A02细胞内的阿霉素浓度升高,耐药指数较未转染细胞明显降低（P<0.05）;转染后耐药细胞株K562/A02细胞MRP-2 mRNA表达水平较未转染细胞下降了67.5%,两者比较差异具有显著性（P<0.05）。结论：MRP-2的高表达导致白血病耐药细胞内化疗药物浓度降低,可能是其导致白血病多药耐药的机制之一。     

汉防己甲素逆转白血病细胞株K562/A02耐药的机制

目的:研究汉防己甲素（TTD）对白血病细胞株K562/A02多药耐药(MDR)逆转的机理。方法：以白血病细胞系K562及其耐药细胞系K562/A02为TTD作用的靶细胞。细胞水平检测实验分5组（K562组、K562/A02组、K562+ADM组、K562/A02+ADM组和K562/A02+TTD+ADM组）,采用MTT法检测TTD对K562和K562/A02细胞的非细胞毒性剂量,流式细胞术检测细胞内阿霉素(ADM)的浓度,基因、酶学、蛋白水平检测实验分3组（K562组、K562/A02组和K562/A02+TTD组）,采用RT-PCR法检测mdr1 mRNA的表达,免疫细胞化学方法检测谷胱甘肽S转移酶π（GST-π）和拓扑异构酶Ⅱ（Topo Ⅱ）的表达水平,Western-blotting法检测P-糖蛋白（P-gp）和bcl-2表达。结果：1.562 5 mg•L^-1的TTD处理K562/A02细胞后,细胞内ADM的浓度较单用ADM组明显提高（P<0.01）;与空白对照组比较,K562/A02细胞内mdr1 mRNA/P-gp的表达量减少（P<0.01）;GST-π和TopoⅡ表达无明显变化;凋亡抑制基因bcl-2的表达量减少（P<0.01）。结论:TTD主要通过增加细胞内ADM浓度,下调mdr1/P-gp和bcl-2表达逆转耐药。     

RNA 干扰沉默同源异型基因A5 逆转K562/ADM 细胞耐药性的研究

目的 研究RNA 干扰（RNAi）技术沉默同源异型基因A5（HOXA 5）对白血病耐阿霉素细胞株K562/ADM 细胞耐药性的影响并探讨其机制。方法 Western blot 检测各组细胞中HOXA5、p38 和p-p38 的蛋白表达,qRT-PCR 检测HOXA5、p38 的mRNA 表达;CCK8 检测各组细胞对阿霉素的敏感性变化;流式细胞术（FCM）检测各组细胞的凋亡及周期变化。结果 K562/ADM 细胞中HOXA 5 基因的表达高于K562 细胞,且其耐药性是K562 细胞的4.94 倍。转染后实验组K562/ADM 细胞中HOXA 5 基因的表达受抑制,而实验组细胞的IC50 较对照组降低2.55 倍。同时,实验组细胞中p38 的mRNA 及p-p38的蛋白表达高于对照组。与对照组比较,实验组的细胞周期G0/G1 期增高,S 期降低。经ADM（2.5μg/ml）诱导后,实验组细胞凋亡率高于对照组。结论 RNAi 沉默HOXA5 能在一定程度上逆转白血病耐药,其机制可能与p38MAPK 信号转导通路的激活有关。     

SiRNA逆转人红白血病细胞株(K562/ADM)阿霉素耐药性的实验研究

目的:研究小分子干扰RNA片段(small inteffering RNA,siRNA)对人红白血病细胞株K562/ADM细胞mdr1基因表达及药物敏感性的影响,探索新的耐药逆转途径.方法:siRNA根椐GeneBank已知序列设计,在脂质体介导下转染K562/ADM细胞;用流式细胞仪检测K562/ADM细胞Pgp的表达;用MTT检测其对阿霉素ADM的敏感性;用PCR-ELISA法检测K562/ADM细胞的端粒酶活性.结果:siRNA转染后K562/ADM细胞Pgp的表达明显下降,对阿霉素ADM的IC50从8.7μg/ml降到5 μg/ml,端粒酶活性明显下调.结论:siRNA有效逆转了mdr1介导的耐药性,不失为一种新型、有效的肿瘤耐药逆转途径.     

CSA逆转白血病细胞株K562/ADM耐药的实验研究

目的研究不同浓度环孢霉素A(2、0.3、0μg/mL)和粒细胞集落刺激因子以及小剂量化疗药物(米托蒽醌 替尼泊甙 阿糖胞苷)组合后,不同组合对耐药白血病细胞株K562/ADM的作用情况,为环孢霉素A逆转耐药的临床应用提供实验依据。方法采用四唑氮蓝(MTT)药敏法测定K562/ADM细胞株的增殖抑制率,观察细胞形态学改变,在不同培养时间进行细胞活力测定,并采用流式细胞仪检测K562/ADM细胞株的P-糖蛋白表达和细胞周期分析。结果在每个时间点,大剂量环孢霉素A(2μg/mL) 粒细胞集落刺激因子 化疗组对K562/ADM细胞株的抑制率最大;培养96 h后,P-糖蛋白平均荧光强度下降最明显(P<0.01)。粒细胞集落刺激因子 小剂量化疗药与不同浓度环孢霉素A结合的三组组合,在培养72 h后,大量细胞出现S期阻滞。结论大剂量环孢霉素A既降低了细胞耐药性,粒细胞集落刺激因子预激又促使细胞进入细胞周期,有利化疗对于白血病细胞进行最大限度的杀伤,二者有明显协同作用。     

siRNA对人红白血病细胞株（K562／ADM）阿霉素耐药性的影响

目的：研究小分子干扰RNA片段（small interfering RNA，siRNA）对人红白血病细胞株（K562／ADM）细胞mdr1基因表达及功能的影响，探索一新的耐药的逆转途径。方法：siRNA根椐GeneBank已知序列设计，在脂质体介导下转染K562／ADM细胞：用流式细胞仪检测K562／ADM细胞Pgp的表达及细胞周期的改变；用TUNEL法检测其凋亡；用MTTT检测其对阿霉素ADM的敏感性。结果：siRNA转染后K562／ADM细胞Pgp的表达明显下降，凋亡率明显提高，对阿霉素ADM的敏感性明显提高。结论：siRNA有效逆转了mdr1介导的耐药性，不失为一种新型、有效的肿瘤耐药逆转途径。     

苦参碱逆转人白血病K562/ADM细胞对阿霉素耐药性的研究

[目的] 探讨苦参碱对人白血病K562／ADM细胞对阿霉素耐药性的逆转作用。[方法] MTT法测定苦参碱的细胞毒性及其对K562／ADM细胞药敏性的影响,荧光分光光度法检测细胞内药物浓度的改变,流式细胞术检测耐药细胞凋亡百分率的变化。[结果] 苦参碱的非细胞毒性剂量为50μg／mL,低细胞毒性剂量为125μg／mL。50μg／mL苦参碱可增加K562／ADM细胞内阿霉素(ADM)浓度和K562／ADM细胞凋亡百分率,使K562／ADM细胞的IC50由原来的35．2μg／mL降低至15．8μg／mL,其逆转倍数为2．2倍。[结论] 苦参碱可部分逆转人白血病K562／ADM细胞对阿霉素的耐药性,其逆转机制与增加细胞内药物积累有关。     

人参皂苷 Rh2抗白血病多药耐药细胞 K562/VCR作用研究

目的 通过观察人参皂苷 Rh2对人白血病多药耐药 (MDR) 细胞 K562/VCR 生长、凋亡的作用和逆转耐药的情况,探索该药在抗人白血病 MDR 方面的应用价值。方法 将人参皂苷Rh2与 K562、K562/VCR 细胞共培养 48 h 后采用 MTT法分析其对细胞生长的抑制率;将其与 K562/VCR 细胞于 37 ℃ 孵育 30 min 后,采用 Annexin V 和 PI 双染法在流式细胞仪上检测细胞凋亡情况;并观察其对 K562/VCR 细胞摄取柔红霉素 (DNR) 能力及细胞表面 P-糖蛋白 (P-gp) 表达的影响;在 DNR 与 K562/VCR 培养体系中加入不同质量浓度人参皂苷Rh2,孵育 48 h 后观察人参皂苷Rh2对 K562/VCR细胞耐药的逆转情况。结果 人参皂苷Rh2 可明显抑制 K562 和 K562/VCR 细胞的生长,并呈量效关系,人参皂苷Rh2对 K562 和 K562/VCR 的半数抑制浓度 (IC50) 分别为44.5、59.4 μg/mL。K562/VCR 经人参皂苷Rh2 在 37 ℃ 作用 30 min 后,细胞的凋亡明显增加,随人参皂苷Rh2质量浓度的增加,凋亡细胞的比例明显增加［人参皂苷 Rh2 300 μg/mL,Annexin V+ 细胞 (51.5±6.9)%］。K562 细胞经长春新碱 (VCR) 诱导耐药后,P-gp 表达率明显提高 (从 4.28% 到 93.80%),DNR 摄取率减少,25 μg/mL 以上质量浓度的人参皂苷Rh2即可明显提高 K562/VCR 对 DNR 的摄取,但 P-gp 的表达无明显改变。同时,它可明显提高 DNR 对 K562/VCR 的杀伤率,50 μg/mL 人参皂苷 Rh2 可使 K562/VCR 对 DNR 的敏感性提高到原来的 6.30 倍。 结论 人参皂苷 Rh2 能抑制 K562/VCR 细胞生长,诱导其凋亡,还可以逆转 K562/VCR 的耐药,是一种具有广阔开发前景的抗白血病药物。     

Reversal effect of bufalin on multidrug resistance in K562/VCR vincristine-resistant leukemia cell line

ObjectiveTo probe insights into the reversal effect of bufalin on vincristine-acquired multidrug resistance (MDR) in human leukemia cell line K562/VCR.MethodsProliferative inhibition rate and the reversal index (RI) of bufalin were determined by Methyl thiazolyl tetrazolium assay. The uptake of Adriamycin (ADM) in K562/VCR cells, cell cycle and apoptosis rate were determined by flow cytometry (FCM). Cell morphologic changes were observed with Wright-Giemsa staining. The expression of P-glycoprotein (P-gp), multidrug-associated protein-1 (MRP1), Bcl-xL and Bax protein were measured by immunocytochemistry.ResultsThe human leukemia multidrug resistant K562/VCR cells showed no cross-resistance to bufalin. The RIs of bufalin at concentrations of 0.0002, 0.001 and 0.005 μmol/L were 4.85, 6.94 and 14.77, respectively. Preincubation of 0.001 μmol/L bufalin for 2 h could increase intracellular ADM fluorescence intensity to 28.07% (P<0.05) and down-regulate MRP1 expression simultaneously, but no remarkable effect was found on P-gp protein. Cell cycle analysis indicated increased apoptosis rate and apparent decreased G2/M phase proportion after treatment with bufalin. When exposed to 0.01 μmol/L bufalin, typical morphological changes of apoptosis could be observed. Down-regulation of Bcl-xL and up-regulation of Bax expression in K562/VCR cells could be detected by immunocytochemistry.ConclusionBufalin could partly reverse the MDR of K562/VCR cells, with a possible mechanism of down-regulating MRP1 expression and activating apoptosis pathway by altering Bcl-xL/Bax ratio.