γ射线诱导人单核细胞系THP-1细胞结合珠蛋白的表达

Expression of haptoglobin induced by γ-ray irradiation in THP-1 cell line

目的探讨γ射线诱导THP-1细胞中结合珠蛋白的表达及调控的机制。方法RT-PCR和Westernblot检测结合珠蛋白mRNA和蛋白的变化。EMSA检测γ射线照射后THP-1细胞核提取物中与结合珠蛋白启动子区域寡聚核苷酸探针复合物的形成,抗体阻滞实验检测可能参与复合物形成的转录因子。结果γ射线能够诱导THP-1细胞中结合珠蛋白的表达,而且有时间和剂量依赖效应,分别包含结合珠蛋白启动子区域的3个不同顺式作用元件的寡聚核苷酸探针均能与核提取物形成复合物,其中B和C区域的复合物形成受γ射线影响明显,stat-3在γ射线照射后表达升高,C/EBPβ-LIP出现先升高而后下降的过程。抗体阻滞实验中Anti-C/EBPα和Anti-C/EBPβ抗体能够形成明显的超迁移条带,Anti-stat3能够影响细胞核提取物与寡聚核苷酸探针复合物的形成。结论γ射线能够诱导THP-1细胞中结合珠蛋白的表达,C/EBPβ-LIP和stat-3参与了其中的调控过程。

Objective To study the induction and regulation mechanisms for haptoglobin induced by γ- ray irradiation in THP-1 cell line. Methods The mRNA and protein of haptoglobin or C/EBPs were detected by RT-PCR and Western blot respectively. EMSA was used to determine the compounds of nuclear extracts and DNA probe which included the sequence of haptoglobin gene promoter. Super shift assay was also applied to analyze the transcription factors that may participate in the formation of compounds. Results γ-ray irradiation induced the synthesis of haptoglobin, which showed a dose-and time-dependence. Western blot analysis showed that γ-ray irradiation increased Stat3 expression, but reduced expression of C/EBPβ-LIP. EMSA and super shift assay demonstrated that Stat3, C/EBPα, and C/EBPβ3 were bound to probe of promoter. Conclusions γ-rays can induce haptoglobin expression in THP-1 cell line. Stat3 and C/EBPβ3-LIP are involved in the activation and regulation of haptoglobin gene expression.