目的: 探讨血红素氧合酶-1(HO-1)在骨肉瘤U2OS细胞多柔比星(DOX)耐药中的作用及相关分子机制。方法: 体外培养U2OS细胞, 建立U2OS-DOX耐药株, 分为U2OS-WT组和U2OS-DOX组。采用siRNA HO-1转染U2OS-DOX细胞, CCK-8法检测细胞活性; RT-PCR法检测缺氧诱导因子1 (HIF-1α)及HO-1的mRNA表达; WB法检测HIF-1α及HO-1的蛋白表达水平; 流式细胞仪检测罗丹明Rh123在细胞内的蓄积。结果: DOX可降低U2OS细胞活性并随剂量的增加愈加明显, 这种诱导作用可以被抗氧化剂(NAC)所逆转(P＜0.01)。U2OS-DOX组HIF-1α及HO-1的mRNA和蛋白表达以及P糖蛋白(P-gp)表达水平均显著增加(P＜0.05)。转染可恢复U2OS-DOX细胞对DOX化疗敏感性并增加其对Rh123的蓄积(P＜0.001)。结论: HO-1可能通过抗氧化应激、增加化疗药物的蓄积等机制发挥U2OS细胞对DOX的耐药性。

Objective: Explore the role and the molecular mechanisms of HO-1 on U2OS cells resistance to DOXorubicin (DOX). Methods: U2OS cells were cultured in vitro and established the U2OS-DOX-resistant strains, experiment were divided into U2OS-WT group and U2OS-DOX group. The DOX-resistant cell line U2OS-DOX were transfected with specific small interfering RNAs (siRNA) of HO-1 gene. Cell viability of U2OS cell line was analyzed by Cell Counting Kit-8; Fluorescence quantitative real-time polymerase chain reaction was used to examine the mRNA expression levels of hypoxia-inducible factor-1α (HIF-1α) and heme oxygenase-1 (HO-1); Western blot was used to determine the protein expression levels of HIF-1 and HO-1; Rhodamine (Rh123) accumulation in cells were analyzed by flow cytometry. Results: DOX can reduce U2OS cell activity in a dose dependent manner, this induction can be rescued by an antioxidant (NAC) (P＜0.05). Compared with the control group, the mRNA and protein expression level of HIF-1α and HO-1 were significantly increased in U2OS-DOX group, as well as P-Glycoprotein (P-gp) expressing (P＜0.05). siRNA HO-1 transfected cells restored U2OS-DOX chemosensitivity and increased accumulation of Rh123 (P＜0.05). Conclusions: HO-1 may play an anti-oxidative stress role and increase the accumulation of chemotherapy drugs, this can recover the resistance of osteosarcoma U2OS cells to DOX chemotherapy.