Objective: To express matrix remodeling associated 7 (MXRA7) in the human acute myeloid leukemia SHI-1 cell line and to assess the role of MXRA7 in the biological function of SHI-1 cells.
Methods: The full-length cDNA sequence of human MXRA7 was synthesized and subcloned into the lentivirus shuttle vector pRRL-Venus. SHI-1 cells were transfected with the lentivirus which was packaged with 293T cells. The YFP-positive cells were sorted by flow cytometry and the stable cell lines were obtained by expanded culture. The expression and distribution of MXRA7 in SHI-1 cells were verified by real-time qPCR, Western blot and laser confocal techniques. Cell proliferation and cell cycle were measured by flow cytometry, and apoptosis was determined by Annexin V and 7-AAD staining. The expression of apoptosis related proteins were detected by Western blot.
Results: The stable SHI-1 cell line overexpressing MXRA7 was established successfully. Laser confocal analysis confirmed that MXRA7 was expressed in the cytoplasm of SHI-1 cells. Compared with the control cell line, the overexpression of MXRA7 showed no effect on the cell proliferation and cell cycle, but reduced the percentage of apoptosis cells induced by methotrexate. Moreover, the expression of BCL-2 protein was increased by overexpression of MXRA7, which can inhibit cell apoptosis.
Conclusion: The SHI-1 stable cell line overexpressing MXRA7 was established successfully, and MXRA7 could inhibit drug-induced apoptosis through increasing the expression of BCL-2 protein.
题目: 基质重塑相关7（MXRA7）对SHI-1白血病细胞系生物学功能的影响.
目的: 构建过表达人全长基质重塑相关7（MXRA7）的SHI-1急性髓系白血病稳转细胞系，检测MXRA7对SHI-1细胞生物学功能的影响.
方法: 合成人MXRA7全长的cDNA序列，并连接到慢病毒穿梭载体pRRL-Venus中，使用293T细胞包装产生慢病毒，感染人白血病细胞系SHI-1，应用流式细胞术分选YFP + 细胞，扩大培养获得稳转细胞系。通过Real-time qPCR、Western blot及激光共聚焦技术验证MXRA7在SHI-1细胞中的表达及分布。采用流式细胞术检测细胞增殖和细胞周期，采用Annexin V和7-AAD染色流式检测细胞凋亡，采用Western blot技术检测与细胞凋亡相关蛋白的表达情况.
结果: 成功构建了过表达MXRA7的SHI-1稳转细胞系，激光共聚焦证明MXRA7表达于SHI-1细胞的胞浆中；与对照细胞相比，过表达MXRA7不会影响细胞增殖和细胞周期，但会降低甲氨蝶呤诱导的凋亡细胞的百分比，促进凋亡抑制蛋白BCL-2的表达.
结论: 成功构建了过表达MXRA7的SHI-1稳转细胞系， MXRA7通过促进BCL-2的表达来抑制药物诱导的细胞凋亡.
Keywords: BCL-2; MXRA7; acute myeloid leukemia; cell apoptosis.