shRNA沉默卵巢癌细胞SKOV-3的IDO基因表达在体外对NK细胞杀伤作用敏感性的影响

The effect of silencing IDO gene expression using shRNA targeting IDO vector on the cytoxic activity of natural kill cells in vitro

目的通过shRNA沉默吲哚胺2,3-双加氧酶(indoleamine2,3-dioxygenase,IDO)基因表达,研究IDO表达在体外对NK细胞杀伤能力的作用。方法 shRNA沉默IDO基因表达质粒和空白质粒分别稳定转染至人卵巢癌细胞SKOV-3,应用Western blot检测IDO在SKOV-3、SKOV-3/Mock和SKOV-3/shIDO细胞中的表达情况,用MTT试剂盒检测3组肿瘤细胞体外生长速度和对NK细胞杀伤作用的敏感性。结果 IDO蛋白在SKOV-3/shIDO细胞中表达被抑制,在SKOV-3和SKOV-3/Mock细胞中有表达。3组肿瘤细胞体外生长曲线比较差异无统计学意义(P>0.05)。SKOV-3/shIDO细胞存活的百分比明显低于其他2组对照(SKOV-3和SKOV-3/Mock)细胞,差异有统计学意义(P<0.05)。结论本研究应用shRNA沉默IDO基因表达质粒稳定转染卵巢癌细胞SKOV-3,获得IDO无表达卵巢癌细胞SKOV-3/shIDO,结果显示抑制IDO表达对卵巢癌细胞体外生长速度无影响,但可增强卵巢癌细胞SKOV-3对NK细胞杀伤作用的敏感性。因此,IDO可以作为卵巢癌基因治疗的潜在新靶点,shRNA沉默IDO基因表达可以作为卵巢癌治疗的新方法。

Objective To investigate the effect of Indoleamine 2,3-dioxygenase（IDO） expression on the sensitivity of ovarian cancer cells against NK cells after silencing IDO gene expression.Method SKOV-3 cells were stably transfected with shIDO expression vector and Mock vector,respectively.Western blot analysis was used to detect IDO expression in SKOV-3,SKOV-3/Mock and SKOV-3/shIDO cells.Cell growth curves were performed using MTT kit.Sensitivity of SKOV-3,SKOV-3/Mock and SKOV-3/shIDO cells against NK cells was investigated using colorimetric assay with MTT.Result IDO expression was detected in the control（SKOV-3 and SKOV-3/Mock） cells but not in the SKOV-3/shIDO cells.There was no intergroup difference in doubling time in vitro（P0.05）.The percent survival ratio of SKOV-3/shIDO cells was significantly lower than that of the control（SKOV-3 and SKOV-3/Mock） cells（P0.05）.Conclusion IDO expression can be down-regulated by shRNA targeting IDO expression vector.IDO expression did not influence the cell growth of SKOV-3 cells in vitro.The cytotoxic activity of ovarian cancer cells against NK cells can be improved by inhibition of IDO expression.Therefore,IDO acts as a potential therapeutic gene target of ovarian cancer and shIDO can be used as a novel therapeutic strategy against ovarian cancer.