镉处理对小鼠睾丸间质细胞PI3K/AKT信号通路的影响

Effect of cadmium on PI3K/AKT signal pathway in mouse leydig cells

目的探讨镉对小鼠睾丸间质细胞3-磷酸肌醇激酶/蛋白激酶B(PI3K/AKT)信号通路的影响。方法 TM3细胞经20μmol/L Cd Cl2处理,分别在加入Cd Cl24 h和8 h后收集细胞;对照组给予相应体积的PBS,8 h后收集细胞。利用实时定量RT-PCR法测定炎症相关细胞因子白细胞介素(IL)-10、肿瘤坏死因子(TNF)-α、IL-6、单核细胞趋化蛋白(MCP)-1、巨噬细胞炎性蛋白(MIP)-2、MIP-1和环氧合酶(COX)-2 mRNA的表达水平以及用Western blot法检测细胞COX-2、AKT和p-AKT蛋白的表达水平。结果与对照组相比,镉能够显著诱导TNF-α和IL-6 mRNA的表达(P<0.01);镉处理4 h组MCP-1 mRNA表达水平明显高于对照组(P<0.01),而镉处理8 h组MCP-1 mRNA表达水平与对照组相比差异无统计学意义;镉对TM3细胞MIP-1和MIP-2mRNA表达差异无统计学意义;镉能够明显诱导TM3细胞COX-2蛋白和mRNA的表达(P<0.01)。此外,镉处理组pAKT蛋白表达水平也明显高于对照组(P<0.01)。结论镉可能通过激活TM3细胞中PI3K/AKT信号通路进而选择性调节部分炎症相关细胞因子的表达。

Objective To study the effects of cadmium on phosphatidylinositol 3-kinase/protein kinase B （PI3K/ AKT） signal pathway in mouse leydig ceils. Methods TM3 cells （ mouse leydig cell line） were incubated with 20 μmol/L CdCI： and collected cells at 4 h and 8 h after cadmium exposure. The control group was incubated with corresponding volume PBS and collected cells after 8 h. The mRNA expression levels of interleukin（IL）-10, tumor necrosis factor （TNF） -ct, IL-6, monocyte chemotactic protein （MCP） -1, macrophage inflammatory protein （MIP） -2, MIP-1 and cyclooxygenase（COX）-2 were detected by RT-PCR; the protein expression levels of COX-2, AKT and p-AKT were measured by Western blot. Results Compared with the control group, cadmium significantly in- creased the mRNA expression levels of IL-6 and TNF-a（P 〈 0. 01 ） ; the mRNA expression level of MCP-1 was al- so increased at 4 h group after cadmium exposure （P 〈0. 01 ） , however, cadmium did not affects the mRNA ex- pression level of MCP-1 at 8 h group after cadmium exposure. No significant differences on the mRNA expression levels of MIP-1 and MIP-2 were observed between the control group and cadmium groups. Cadmium markedly up- regulated the mRNA and protein levels of COX-2 in TM3 cells （P 〈0.01 ）. In addition, compared with the control group, the protein expression of p-AKT also was significantly increased in cadmium groups （ P 〈 0. 01 ）. Conclu- sion These results suggest that cadmium could activate PI3 K/AKT signal pathway which might partially contribute to cadmium-induced the secretion of some inflammatory cytokines in mouse testicular leydig cells.