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目的探讨硒蛋白K(Selenoprotein K,Sel K)基因沉默(RNA silence,RNAi)对小鼠T淋巴细胞内钙流和白介素2受体(Interleukin-2 Receptor,IL-2R)分泌的影响。方法制作小鼠T淋巴细胞干扰载体p GPU-Sel K,用脂质体Lipofectin 2000将干扰质粒和对照质粒转染小鼠T淋巴细胞,Real time PCR和Western Blot分别检测核酸和蛋白水平Sel K的表达,流式细胞仪检测细胞内钙离子浓度,Real time PCR检测IL-2R的分泌。结果与转染的对照组质粒p GPU-NC比较,转染p GPU-Sel K载体的细胞Sel K在m RNA和蛋白水平表达分别显著降低了42.37%(P<0.01)和45.44%(P<0.01)。另外干扰组整体细胞内钙离子浓度显著低于对照组(P<0.05),细胞分泌IL-2R水平也显著降低(P<0.05)。结论干扰载体p GPU–Sel K可以下调小鼠T淋巴细胞Sel K的表达,抑制细胞的钙流和IL-2R的分泌,从而可能改变小鼠T淋巴细胞增殖分化。
Objective To investigate the effect of selenoprotein K (Sel K) gene silencing on the secretion of intracellular calcium and Interleukin-2 Receptor (IL-2R) in T lymphocytes of mice. Methods Mouse T lymphocyte interference vector p GPU-Sel K was made. Mouse interference plasmids and control plasmids were transfected into mouse T lymphocytes with Lipofectin 2000. Real-time PCR and Western Blot were used to detect the mRNA and protein levels of Sel K The intracellular calcium concentration was detected by flow cytometry. The secretion of IL-2R was detected by Real time PCR. Results Compared with p GPU-NC plasmid transfected control group, Sel K transfected with p GPU-Sel K vector significantly decreased mRNA and protein levels by 42.37% (P <0.01) and 45.44% (P < 0.01). In addition, the total intracellular calcium concentration in the interference group was significantly lower than that in the control group (P <0.05), and the level of IL-2R secreted by the cells was also significantly decreased (P <0.05). Conclusion The interfering vector p GPU-Sel K can down-regulate the expression of Sel K in mouse T lymphocytes and inhibit the secretion of calcium and IL-2R, which may change the proliferation and differentiation of mouse T lymphocytes.