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[目的]研究微囊藻毒素-LR对睾丸支持细胞活性的影响及其对细胞凋亡相关基因表达的影响。[方法]分离纯化大鼠睾丸支持细胞,用低剂量(0~500×10-3μg/ml)和高剂量(1~20μg/ml)微囊藻度素-LR染毒细胞,细胞培养24h、48h和72h后,用四甲基偶氮唑蓝(MTT)实验和中性红吸收(NR)实验检测微囊藻毒素-LR对支持细胞活性的影响;将纯化后的部分睾丸支持细胞接种于6孔板中,给予不同剂量的微囊藻毒素-LR(0,1,10μg/ml),于24h和48h后提取细胞总RNA,RT-PCR方法检测细胞中凋亡相关基因P53、bax和bcl-2表达水平。[结果]低剂量(0~500×10-3μg/ml)微囊藻毒素-LR作用于支持细胞24、48、72h,细胞活性增强。高剂量微囊藻毒素-LR(10和20μg/ml)作用24、48h,细胞活性显著降低。时间效应实验结果显示随着暴露时间的延长细胞活性降低。细胞暴露于1μg/ml微囊藻毒素-LR后与对照组细胞相比较P53和bcl-2mRNA水平相对表达量增加,bax表达量降低。暴露于10μg/ml微囊藻毒素-LR后P53和bax mRNA水平相对表达增加,bcl-2表达降低。[结论]低剂量的微囊藻毒素-LR对细胞具有兴奋效应,随着剂量的增高及暴露时间的延长微囊藻毒素-LR能够明显抑制细胞活性。同时微囊藻毒素-LR可通过P53、bax和bcl-2基因的表达影响睾丸支持细胞的凋亡。
[Objective] To investigate the effect of microcystin-LR on the activity of testicular sertoli cells and its effect on the expression of apoptosis-related genes. [Method] Sertoli cells were isolated and purified from rat testis. The cells were exposed to microcystin-LR at low dose (0 ~ 500 × 10-3μg / ml) and high dose (1 ~ 20μg / ml) 48h and 72h later, the effect of microcystin-LR on the activity of supporting cells was detected by MTT assay and neutral red absorption (NR) assay. Part of the purified testis-supporting cells were seeded on 6 microtiter plates were given different doses of microcystin-LR (0, 1, 10μg / ml). Total RNA was extracted after 24h and 48h. The expression of apoptosis-related genes P53, bax bcl-2 expression level. [Result] The microcystin-LR at low dose (0-500 × 10-3 μg / ml) increased the cell viability at 24, 48, and 72 hours. High-dose microcystin-LR (10 and 20μg / ml) for 24,48h, cell activity was significantly reduced. The results of the time effect experiment showed that the cell activity decreased as the exposure time prolonged. After exposure to 1 μg / ml microcystin-LR, the relative expression of P53 and bcl-2 mRNA increased and the expression of bax decreased compared with the control cells. The relative expression of P53 and bax mRNA increased and the expression of bcl-2 decreased after exposure to 10 μg / ml microcystin-LR. [Conclusion] The low dose of microcystin-LR has an exciting effect on the cells. With the increase of dose and prolonging of exposure time, microcystin-LR can obviously inhibit the cell activity. At the same time microcystin-LR can affect the apoptosis of testicular support cells through the expression of P53, bax and bcl-2 genes.