MCL-1反义寡核苷酸对WM451细胞增殖和凋亡的调控作用

Effect of Mcl-1 Antisense Oligonucleotide on WM451 Cell Proliferation and Apoptosis

目的探讨脂质体转染M c l-1反义寡核苷酸(M c l-1 AS-ODN)对黑色素瘤细胞WM451增殖及凋亡的影响。方法用阳离子脂质体法转染反义M c l-1至转移性人黑色素瘤细胞株WM451。利用RT-PCR、W estern b lot及免疫组化法检测细胞转染前后M c l-1表达情况,MTT法检测M c l-1 AS-ODN对细胞增殖的影响,流式细胞仪和电镜观察转染后细胞的凋亡情况。结果M c l-1 AS-ODN显著降低M c l-1 mRNA和蛋白水平,而正义序列寡核苷酸(S-ODN)和无关序列寡核苷酸(NS-ODN)对基因表达无明显影响。MTT检测显示M c l-1 AS-ODN可以抑制WM451细胞增殖,观察M c l-1 AS-ODN转染后细胞呈现典型凋亡特征,流式细胞仪测定转染后细胞凋亡率为13.6%,明显高于对照组(6.3%)。结论M c l-1AS-ODN能够下调M c l-1基因表达,抑制细胞增生并诱导其凋亡,M c l-1可作为黑色素瘤基因治疗的一个新靶点。

Objective To explore the effect of liposomal transfection of Mcl-1 antisense oligonucleotide（ Mcl- 1 AS-ODN） on WM451cell proliferation and apoptosis. Methods Mcl-1 AS-ODN was transfected into a metastatic human melanoma cell line WM451 with lipofectamine 2000. Mcl-1 expression was analyzed by RT-PCR, Western blotting and immunohistochemistry. Cell viability was assessed by MTT assay, apoptosis was evaluated by flow eytometry and electron microscopy. Results The expression level of Mcl-1 mRNA and protein was decreased in AS-ODN group, whereas there was no significant difference in S-ODN group and NS-ODN group,compared with control group. MTT assay demonstrated that Mcl-1 AS-ODN could reduce the viability of WM451cells. Electron microscopy showed that WM451 cells exhibited characteristic morphologic changes of apoptosis after incubation with Mcl-1 AS-ODN. The apoptosis rate of WM451 cells treated with AS-ODN was 13.6%, which is higher than that of control（6.3% ） by flow cytometry. Conclusion Mcl-1 AS-ODN can downregulate Mcl-1 expression, inhibit WM451 cells proliferation and induce apoptosis,whieh is a promising target for antisense therapy of melanoma.