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目的:探讨蝎毒多肽(PESV)对多药耐药白血病干细胞MDR1 mRNA和P-gp表达的影响。方法:以K562/A02细胞株为例,免疫磁珠法分选对数生长期细胞后经CCK-8法确认其耐药性,将耐药的K562/A02干细胞接种于BABL/c裸鼠右腋下形成瘤块,再将瘤块皮下包埋建立耐药性稳定的白血病模型。成模裸鼠随机分为6组:模型对照组、ADM组、PESV组、ADM+高剂量PESV组(高剂量组)、ADM+中剂量PESV组(中剂量组)和ADM+低剂量PESV组(低剂量组)。模型对照组给予等体积0.9%氯化钠溶液腹腔注射,其余组予相应剂量ADM和(或)PESV腹腔注射,连续给药14天。第21天观察各组裸鼠移植瘤生长情况,流式细胞术检测各组瘤组织P-gp表达,RT-PCR法检测各组瘤组织MDR-1 mRNA表达水平。结果:K562/A02细胞经免疫磁珠分选前后的CD34+CD38-细胞比率和耐药率分别为31.5%、(60.33±10.68)μg/m L,92.8%、(58.33±9.72)μg/m L,分选后细胞干性显著提高,而耐药性无差异性损失。各组造模小鼠成瘤率100%,P-gp表达结果显示模型对照组89.8%、ADM组91.9%、PESV组88.4%、高剂量组53.9%、中剂量组78.0%、低剂量组78.7%,PESV+ADM下调P-gp表达,并与PESV呈现浓度依赖关系。MDR1mRNA的表达量PESV组>低剂量组>高剂量组>中剂量组>ADM组,其中PESV组、中剂量组、低剂量组高表达,P-gp表达与MDR1 mRNA水平具有一定的一致性。结论:PESV降低ADM作用K562/A02干细胞耐药性的强度,并与PESV剂量呈现正相关,其机制可能为PESV通过调控P-gp和MDR-1 mRNA的表达,增强了K562/A02干细胞对ADM的敏感度。
Objective: To investigate the effects of scorpion venom peptide (PESV) on MDR1 mRNA and P-gp expression in multidrug-resistant leukemia stem cells. Methods: The K562 / A02 cell line was used as an example. The cells in logarithmic growth phase were sorted by immunomagnetic beads and their drug resistance was confirmed by CCK-8 assay. The resistant K562 / A02 stem cells were inoculated on the right side of BABL / c nude mice Underarm tumor blocks formed, and then embedding the tumor subcutaneously to establish a stable model of drug-resistant leukemia. The model nude mice were randomly divided into 6 groups: model control group, ADM group, PESV group, ADM + high dose PESV group (high dose group), ADM + medium dose PESV group (middle dose group) and ADM + low dose PESV group group). The model control group was given 0.9% sodium chloride solution by intraperitoneal injection, and the remaining groups were given intraperitoneal injection of ADM and / or PESV for 14 consecutive days. On day 21, the growth of xenografts in nude mice in each group was observed. The expression of P-gp in each group was detected by flow cytometry. The expression of MDR-1 mRNA in each group was detected by RT-PCR. Results: The percentage of CD34 + CD38-cells and the drug resistance of K562 / A02 cells before and after immunomagnetic bead sorting were 31.5%, 60.33 ± 10.68 μg / m L, 92.8%, 58.33 ± 9.72 μg / m L, cells were significantly increased after sorting, and no difference in drug resistance loss. The tumor formation rate of mice in each group was 100%. The expression of P-gp was 89.8% in the model control group, 91.9% in the ADM group, 88.4% in the PESV group, 53.9% in the high dose group, 78.0% in the middle dose group and 78.7% in the low dose group %, PESV + ADM down-regulated the expression of P-gp and showed a concentration-dependent relationship with PESV. The expression of MDR1 mRNA in PESV group> low dose group> high dose group> middle dose group> ADM group. The expression of MDR1 mRNA in PESV group was higher than that in low dose group. The expression of P-gp was consistent with MDR1 mRNA level. CONCLUSION: PESV can decrease the drug resistance of K562 / A02 stem cells induced by ADM and positively correlated with the dose of PESV. The possible mechanism is that PESV enhances the resistance of K562 / A02 stem cells to ADM by regulating the expression of P-gp and MDR- Sensitivity.