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目的:探讨自噬相关蛋白Atg5及P62在邻苯二甲酸二乙基己酯(diethylhexyl phthalate,DEHP)引起新生鼠卵巢卵泡发育异常这一过程中的作用。方法:将健康6~8周龄清洁级成年雌性昆明鼠随机分为对照组(玉米油)和DEHP组,每组30只,均不做任何处理。成年雌鼠生产后的新生鼠分为对照组(玉米油)和DEHP组,采用腹腔注射方式,DEHP组按0.62 mg/kg浓度对新生鼠进行注射,对照组注射相同体积玉米油,从新生鼠出生第1天开始注射,每天1次,连续5 d,注射结束后收集新生鼠卵巢组织,HE染色观察卵巢各级卵泡的发育情况,q RT-PCR及Western blot检测卵泡发育标志分子GDF9及自噬相关蛋白Atg5、P62的表达情况。结果:与对照组比较,HE染色结果显示DEHP组原始卵泡数量明显减少(P=0.005),而初级卵泡数量无明显改变(P=0.555);q RT-PCR结果显示DEHP组新生鼠卵巢组织中卵泡发育标志分子GDF9 m RNA表达明显降低(P=0.034),而自噬标志分子Atg5、P62 m RNA表达明显增加(P值分别为0.001、0.024)。Western blot结果显示DEHP组新生鼠卵巢中自噬相关蛋白Atg5及P62均明显增加,差异有统计学意义(P值分别为0.008、0.001)。结论:在DEHP暴露引起新生鼠卵巢卵泡发育异常这一过程中,自噬标记蛋白Atg5及P62可能发挥着重要作用。
AIM: To investigate the role of autophagy-related proteins Atg5 and P62 in the development of ovarian follicles in neonatal rats induced by diethylhexyl phthalate (DEHP). Methods: Healthy female Kunming mice, aged 6-8 weeks, were randomly divided into control group (corn oil) and DEHP group, 30 rats in each group, without any treatment. The newborn rats were divided into control group (corn oil) and DEHP group. The neonatal mice were injected with 0.62 mg / kg DEHP group and the same volume of corn oil was injected into control group. On the first day of life, the mice were injected once a day for 5 consecutive days. After the injection, newborn rat ovarian tissues were harvested and the development of ovarian follicles was observed by HE staining. The follicle development marker GDF9 was detected by q RT-PCR and Western blotting Atg5, P62 protein expression. Results: Compared with the control group, HE staining showed that the number of primordial follicles in DEHP group decreased significantly (P = 0.005), while the number of primary follicles did not change significantly (P = 0.555). Q RT-PCR results showed that in DEHP group, The expression of GDF9mRNA in follicle development significantly decreased (P = 0.034), while the expression of Atg5 and P62mRNA in ovarian follicles significantly increased (P = 0.001 and 0.024, respectively). Western blot results showed that the expression of Atg5 and P62 in DEHP group were significantly increased (P = 0.008, 0.001 respectively). CONCLUSION: Autophagy marker proteins Atg5 and P62 may play an important role in the abnormal ovarian follicle development induced by DEHP exposure.