Wnt1-Ngn2-En1/2在西玛津损伤小鼠生命早期多巴胺能神经元功能中的作用

doi:10.3969/j.issn.1004-616x.2024.06.004

摘要/Abstract

摘要： 目的： 观察除草剂西玛津对小鼠子代胚胎期及出生后第21天(PND21)多巴胺能神经元发育的影响，并探讨Wnt1-Ngn2-En1/2信号通路在神经元损伤中的作用。方法： 健康SPF级昆明小鼠雌性30只，雄性15只，按照2∶1进行合笼。将孕鼠随机分为对照组(0.3%甲基纤维素溶液)和西玛津低剂量(50 μg/kg)、高剂量组(200 μg/kg)进行灌胃染毒，收集胚胎第8.5天(E8.5)、第12.5天(E12.5)和PND21仔鼠的中脑组织，采用免疫荧光杂交技术(FISH)、实时荧光定量PCR(qPCR)和Western blot分别检测Wnt1-Ngn2-En1/2信号通路相关基因(Wnt1、Ngn2、En1、En2、Nr4a2与Th) mRNA和蛋白的表达；通过旷场实验检测孕鼠染毒西玛津后对PND21雌、雄仔鼠自主行为能力的影响。结果： qPCR与Western blot检测结果显示，与对照组相比，在E8.5胚胎组织中，西玛津低剂量染毒组Wnt1和Ngn2以及高剂量组Wnt1、Ngn2、En1和En2的mRNA与蛋白表达均明显下降(P<0.05)；在E12.5胚胎组织中，低剂量染毒组Nr4a2和Th以及高剂量组Nr4a2、Th、Wnt1、Ngn2、En1、En2的mRNA和蛋白表达均显著下降(P<0.05)。FISH检测结果显示，与对照组相比，西玛津低、高剂量染毒组E8.5与E12.5胚胎组织中Wnt1的mRNA表达均明显下降(P<0.05)；西玛津高剂量组PND21雄性仔鼠的自主探索能力与运动能力显著减弱(P<0.05)，且PND21仔鼠组织中Wnt1与Th的蛋白表达也明显下调(P<0.05)。结论： 西玛津能够通过Wnt1-Ngn2-En1/2信号通路影响子代小鼠多巴胺能神经元发育，并损伤雄性仔鼠的自主行为能力。

关键词: 西玛津, 生命早期, 神经毒性, 多巴胺能神经元

Abstract: OBJECTIVE： To investigate the role of the Wnt1-Ngn2-En1/2 signaling pathway in the herbicide simazine-induced damage to the dopaminergic neurons in embryonic and weaned mice. METHODS： Fifteen female and fifteen male healthy SPF Kunming mice were kept in containers in a 2∶1 ratio. During pregnancy，three groups of maternal mice were randomized∶control group (0.3% methyl cellulose solution)，low-dose group (50 μg/kg) and high-dose group (200 μg/kg). The tissues of the offspring on the day 8.5 (E8.5)，day 12.5 (E12.5)，and postnatal day 21 (PND21) of development were obtained. mRNA and protein expression of the Wnt1-Ngn2-En1/2 signaling pathway-related genes (Wnt1，Ngn2，En1，En2，Nr4a2 and Th) were detected using immunofluorescence hybridization (FISH)，real-time fluorescence quantitative PCR (qPCR) and Western blot. Through an open field experiment，effect of the treated pregnant mice on the autonomic behavior of PND21 female and male mice were identified. RESULTS： Western blot and qPCR data in E8.5 demonstrated that in comparison to the control group，low simazine-exposed groups had a decrease in the mRNA and protein expression of Wnt1 and Ngn2 (P<0.05)，and the high-dose group had a substantial decline in the expressions of Wnt1，Ngn2，En1 and En2 (P<0.05). The results of E12.5 embryo show the mRNA and protein expression of Nr4a2 and Th in the low-dose group declined as compared with the control group，(P<0.05)，and the expression of Nr4a2，Th，Wnt1，Ngn2，En1 and En2 in high-dose group decreased significantly compared with that in control group (P<0.05). The FISH data verified that Wnt1 mRNA expression was substantially reduced in the low-dose and high-dose sections of E8.5 and E12.5 as opposed to that of the control group (P<0.05). The high-dose group of PND21 offspring exhibited significantly reduced motor and self-exploration abilities in comparison to the male offspring of the control group (P<0.05)，and there was a notable down-regulation of Wnt1 and Th protein expressions (P<0.05). CONCLUSION： Simazine affected the development of dopaminergic neurons in male offspring through the Wnt1-Ngn2-En1/2 signaling pathway，and damaged the ability of independent behavior of male mice.

Key words: simazine, early life, neurotoxicity, dopaminergic neuron

中图分类号:

R994.6

刘茄琦, 闻杰, 张佳钰, 李百祥, 李雪婷. Wnt1-Ngn2-En1/2在西玛津损伤小鼠生命早期多巴胺能神经元功能中的作用[J]. 癌变·畸变·突变, 2024, 36(6): 444-451.

LIU Jiaqi, WEN Jie, ZHANG Jiayu, LI Baixiang, LI Xueting. The role of Wnt1-Ngn2-En1/2 in the simazine-induced injury to the dopaminergic neurons in early life of mice[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2024, 36(6): 444-451.

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