冬凌草甲素通过下调Brg1表达抑制Jurkat细胞生长

Oridonin inhibits proliferation of Jurkat cells via the down-regulation of Brg1

摘要:

目的 探讨冬凌草甲素对人急性T淋巴细胞白血病Jurkat细胞的作用及其机制。方法 体外培养Jurkat细胞株，用不同浓度冬凌草甲素（0、1.25、2.5、5和10 μmol/L）作用Jurkat细胞不同时间（24、48、72 h），MTT实验观察细胞增殖情况，荧光显微镜观察不同浓度冬凌草甲素处理Jurkat细胞12 h后的细胞核形态变化。采用Western blot半定量法检测不同浓度冬凌草甲素作用Jurkat细胞24 h，以及5 μmol/L冬凌草甲素处理Jurkat细胞不同时间（0、2、6、12和24 h）后的Brg1、P53和C-myc蛋白表达。用siRNA沉默Jurkat细胞的Brg1，观察其对P53、C-myc蛋白表达以及对增殖的影响。结果 与无处理组相比，冬凌草甲素对Jurkat细胞增殖有抑制作用（P < 0.05），且呈浓度和时间依赖性；荧光显微镜下发现，冬凌草甲素处理后的Jurkat细胞出现细胞核浓集、固缩等典型凋亡形态变化。与无处理组相比，5 μmol/L冬凌草甲素作用下，Jurkat细胞的Brg1和C-myc表达下降，而P53表达上升。siRNA沉默Jurkat细胞的Brg1后，P53表达升高，C-myc蛋白表达下降，细胞生长明显受抑（P < 0.05）。结论 冬凌草甲素对Jurkat细胞生长具有抑制作用，其机制可能通过影响Brg1信号通路起作用。

Abstract:

Objective To investigate the effect of oridonin on the human acute lymphocytic leukemia cell line Jurkat and its mechanism. Methods Jurkat cells were cultured in vitro and treated with various concentrations (0, 1.25, 2.5, 5, and 10 μmol/L) of oridonin for different lengths of time (24, 48, and 72 hours). The proliferation of Jurkat cells was analyzed by MTT assay. The changes in nuclear morphology were evaluated by fluorescence microscopy at 12 hours after treatment with various concentrations of oridonin. The expression levels of Brg1, P53, and C-myc were determined by semi-quantitative Western blot in Jurkat cells treated with various concentrations of oridonin for 24 hours or 5 μmol/L oridonin for various lengths of time (0, 2, 6, 12, and 24 hours). The expression levels of P53 and C-myc and proliferation of Jurkat cells were evaluated after Brg1 expression was knocked down by Brg1-specific siRNA. Results Compared with the control group, the proliferation of oridonin-treated Jurkat cells was significantly inhibited in a concentration-and time-dependent manner (P < 0.05). According to the florescence microscopic analysis, oridonin treatment led to nuclear pyknosis in Jurkat cells. Compared with the control group, Jurkat cells treated with 5 μmol/L oridonin had reduced expression of Brg1 and C-myc but elevated expression of P53. Brg1 knock-down led to a significant reduction in proliferation of Jurkat cells (P < 0.05), up-regulated expression of P53, and down-regulated expression of C-myc. Conclusions Oridonin can inhibit the proliferation of Jurkat cells, probably via the Brg1 signaling pathway.