硫酸镍对小鼠睾丸间质细胞增殖及睾酮合成影响

Effects of nickel sulfate on cell proliferation and testosterone synthesis in mice testicular Leydig cells

目的研究硫酸镍对小鼠睾丸间质细胞(以下简称"TM3细胞")增殖及睾酮合成的影响。方法分别以质量浓度为13. 14、26. 28、78. 84、131. 40 mg/L硫酸镍染毒TM3细胞,对照组不予硫酸镍染毒处理,分别于处理后0、6、12、24、48 h采用CCK-8法检测TM3细胞的细胞存活率;采用酶联免疫吸附法测定细胞上清液中睾酮水平,并通过蛋白免疫印迹法检测细胞内类固醇激素合成急性调控蛋白(St AR)、细胞色素P450胆固醇侧链裂解酶(P450SCC)、细胞色素P45017α-羟化酶(P45017α)、17β-羟类固醇脱氢酶(17β-HSD)蛋白的表达水平。结果经硫酸镍染毒后,TM3细胞存活率在染毒剂量和染毒时间主效应以及两者的交互效应上均有统计学意义(P <0. 01)。以剂量为78. 84、131. 40 mg/L硫酸镍染毒12 h后,与对照组比较,TM3细胞睾酮水平及St AR、P450SCC、P45017α、17β-HSD相对表达水平均降低(P <0. 01),且131. 40 mg/L组上述指标低于78. 84 mg/L组(P <0. 01)。结论硫酸镍可能通过影响细胞内St AR及部分睾酮合成关键酶的合成,从而抑制TM3细胞的增殖及睾酮的分泌。

Objective To study the effects of nickel sulfate（ Ni SO4） on the cell proliferation and testosterone biosynthesis in mice testicular Leydig cells（ Tm^3 cells） in vitro. Methods The Tm^3 cells were treated with 13. 14,26. 28,78. 84 and 131. 40 mg/L Ni SO4. The control group was treated with no Ni SO4. The CCK-8 method was used to detect the cell survival rate of Tm^3 cells at time points of 0,6,12,24 and 48 hours after treatment. Determination of testosterone levels in cell supernatants was performed by enzyme-linked immunosorbent assay. Detection of intracellular steroidogenic acute regulatory proteins（ St AR）,P450 side chain cleavage enzyme（ P450 SCC）,cytochrome P450 17α-hydroxylase enzyme（ P45017α） and 17β-hydroxysteroid dehydrogenase（ 17β-HSD） proteins was conducted by Western blotting.Results After exposure to Ni SO4,the cell survival rate of Tm^3 cells was statistically decreased in groups with different Ni SO4 dose and different time（ P〈0. 01）. The level of testosterone,and the expression of St AR,P450 SCC,P45017αand17β-HSD decreased in groups treated with 78. 84 and 131. 40 mg/L Ni SO4 for 12 hours compared with the control group（ P〈0. 01）. The above indexes level of the 131. 40 mg/L group was lower than that of the 78. 84 mg/L group（ P〈0. 01）. Conclusion Ni SO4 inhibits the proliferation of Tm^3 cells and the secretion of testosterone by affecting the key enzymes in St AR and testosterone synthesis.