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目的探讨 p38丝裂原活化蛋白激酶(p38MAPK)对地塞米松(DEX)诱导急性淋巴细胞性白血病 CEM 细胞株凋亡过程中糖皮质激素受体α(GR_α)功能的影响。方法台盼蓝拒染法绘制增殖曲线;流式细胞仪解析细胞凋亡;Western blot 检测糖皮质激素受体蛋白及磷酸化 p38MAPK 蛋白表达。结果 (1)p38MAPK 特异阻滞剂 SB203580预处理后继续培养24～72 h,细胞的增殖率分别由单用 DEX 时的62.3%、35.5%、11.6%升至82.8%、54.7%和48.1%(P<0.01);(2)5 μmol/LDEX 处理 CEM 细胞36 h 时,亚二倍体细胞为26.2%,显著高于对照组(P<0.01)。SB203580预处理后,亚二倍体细胞由26.2%降至7.1%(P<0.01);(3)5 μmol/L 的 DEX 处理 CEM 细胞后,GR_α总蛋白的表达水平从12 h 开始分别上调至117%(12 h)、121%(24 h)、1225(36 h)、125%(48 h)。未与DEX 结合的 GR_α主要存在于细胞浆中,细胞核 GR_α与细胞浆 GR_α之比为0.27。从6～48 h,GR_α的胞核与胞浆之比分别为0.48、0.59、0.95、2.16、4.08;(4)5 μmol/L DEX 处理 CEM 细胞后,p-p38MAPK 蛋白在15 min 时即隐约可见,1 h 开始表达增强并持续增强至6 h,之后减弱至48 h 仍可见。SB203580预处理后,未检测到 p-p38MAPK 蛋白的表达;(5)SB203580预处理后,GR_α的核浆比例由4.08降至0.43(P<0.05),GR_α总蛋白无明显变化。结论 DEX 在诱导 CEM 细胞凋亡时上调GR_α的蛋白表达水平并促进转位入核。p38MAPK 信号转导通路的活化促进了 GR_α转位入核。
Objective To investigate the effect of p38 mitogen-activated protein kinase (p38MAPK) on the function of glucocorticoid receptor α (GR_α) during dexamethasone (DEX) -induced apoptosis in CEM cell lines of acute lymphoblastic leukemia. Methods The proliferation curve was drawn by trypan blue exclusion method. The apoptosis was analyzed by flow cytometry. The expressions of glucocorticoid receptor protein and phosphorylated p38MAPK protein were detected by Western blot. Results (1) The cell proliferation rate increased from 62.3%, 35.5%, 11.6% to 82.8%, 54.7% and 48.1% (P <0.01) when the p38MAPK specific antagonist SB203580 was pretreated for 24-72 h P <0.01). (2) When treated with 5 μmol / L LDEX for 36 h, the diploid cells were 26.2%, which was significantly higher than that of the control group (P <0.01). After pretreatment with SB203580, the sub-diploid cells decreased from 26.2% to 7.1% (P <0.01). (3) After treated with 5 μmol / L DEX, the expression level of total protein of GR_α increased from 12 h to 117% (12 h), 121% (24 h), 1225 (36 h), 125% (48 h). GR_α not bound to DEX mainly exists in cytoplasm, the ratio of GR_α to cytoplasm GR_α is 0.27. From 6 to 48 h, the ratio of nucleus to cytoplasm of GR_α were 0.48,0.59,0.95,2.16,4.08, respectively. (4) After treatment with 5 μmol / L DEX for CEM cells, p-p38MAPK protein was vague at 15 min Visible, 1 h began to enhance expression and continued to enhance to 6 h, then weakened to 48 h still visible. After pretreatment with SB203580, the expression of p-p38MAPK protein was not detected. (5) After pretreatment with SB203580, the ratio of GR_α nucleoplasm decreased from 4.08 to 0.43 (P <0.05), and the total protein of GR_α did not change significantly. Conclusion DEX can upregulate the expression of GR_α protein and induce translocation into the nucleus when CEM cells induce apoptosis. The activation of p38MAPK signal transduction pathway promotes the translocation of GR_α into the nucleus.