miR-155对含SH2区域的肌醇5′磷酸酶1转录后调控在急性髓系白血病发病机制中作用的初步研究

Preliminary study of role of post-transcription regulation on SH2 domain-containing inositol 5′-phosphatase 1 gene expression by miR-155 in the pathogenesis of acute myeloid leukemia

目的探讨miR-155对人类含SH2区域的肌醇5′磷酸酶1（SHIPl）的转录后调控在急性髓系白血病（AML）发病机制中的作用。方法应用反转录聚合酶链反应（RT-PCR）法检测30例AML患者miR-155、SHIPl的mRNA表达水平，选取同年龄健康人骨髓为对照组。人白血病U937细胞转染miR-155类似物后，RT—PCR法检测转染细胞中miR-155、SHIPl的mRNA表达水平。Western blot法检测转染后细胞SHIP1、AKT、pAKT蛋白水平。流式细胞术检测转染后细胞凋亡的变化。结果30例AML患者中，15例AML—M4及AML—M5患者SHIP1蛋白水平较非AML—M4及AML—M5患者明显降低，而miR-155表达水平相应升高（均P〈0．05）。U937细胞转染miR-155后，SHIP1蛋白水平较转染阴性对照组降低（P〈0．05），而p-AKT水平较转染阴性对照组明显升高，转染后细胞凋亡明显受抑（P〈0．05）。结论miR-155可对SHIP1进行转录后调控，miR-155可能通过降低SHIPl活性而激活PI3K—AKT途径，抑制白血病细胞的凋亡，从而促进AML的发生。

Objective To investigate the role of microRNA-155 （miR-155） on post-transcription regulation of SH2 domain-containing inositol 5＇-phosphatase 1 （SHIP1） gene expression in the pathogenesis of acute myeloid leukemia （AML）. Methods Quantitative real-time polymerase chain reaction （RT-PCR） was performed to detect the expression of miR-155 and SHIP1 mRNA in the AML patients and controls, miR-155 mimics was transfected into U937cells （U937m） by using X-treme GENE siRNA transfection reagent. Cells without transfection （U937c） and cells with negative transfection （U937mc） were used as controls. RT-PCR was performed to detect the expression of miR-155 and SHIP1 mRNA in these cells. The expression of SHIP1, TAKT and pAKT were detected by Western blot in U937 cells. Apoptosis was studied by flow cytometry （FCM）. Results The average level of SHIP1 protein content in 15 samples of patients with AML-M4 or AML-M5 from 30 AML patients was significantly lower compared with that of patients with the other types of AML, and the levels of miR-155 were significantly higher in the same group of patients （P 〈 0.05）. Significantly decreased levels of SHIP1 protein were found in U937m cells compared that of U937c and U937mc （P 〈 0.05）. Significantly decreased rate of apoptosis was observed in U937m cells compared with that of U937mc and U937c. U937m cells also exhibited no alteration in total AKT content, while increased p-AKT levels were found （P 〈 0.05）. Conclusion SHIP1 was also a primary target of miR-155 in AML. Overexpression of miR-155 could activate PI3K-AKT pathway to depressing SHIP1 and decrease the apoptosis rate of AML cells.