School of Pharmacy, Xianning Medical College, Hubei University of Science and Technology, 437100, China; School of Stomatology and Ophthalmology, Xianning Medical College, Hubei University of Science and Technology, 437100, China; Hubei Key Laboratory of Diabetes and Angiopathy, Hubei University of Science and Technology, Xianning 437100, China.
Basic Medical School, Xianning Medical College, Hubei University of Science and Technology, Xianning 437100, China.
Toxicology. 2023 Mar 1;486:153432. doi: 10.1016/j.tox.2023.153432. Epub 2023 Jan 22.
Neuroinflammation is one of the important mechanisms of trimethyltin chloride (TMT) central neurotoxicity. Artemisinin (ARS) is a well-known antimalarial drug that also has significant anti-inflammatory effects. Prokineticin 2 (PK2) is a small molecule secreted protein that is widely expressed in the nervous system and plays a key role in the development of neuroinflammation. However, it remains unclear whether ARS can ameliorate neuroinflammation caused by TMT and whether PK2/PKRs signaling pathway plays a part in it. In this research, male Balb/c mice were administered TMT (2.8 mg/kg, i.p.) followed by immunohistochemistry to assess the expression of PK2, PKR1, and PKR2 proteins in the hippocampus. Network pharmacology was used to predict the intersection targets of ARS, central nervous system(CNS) injury and TMT. The neurobehavior of mice was evaluated by behavioral scores. Histopathological damage of the hippocampus was evaluated by HE, Nissl and Electron microscopy. Western blotting was used to identify the expression of synapse-related proteins (PSD95, SYN1, Synaptophysin), PK system-related proteins (PK2, PKR1, PKR2), and inflammation-related proteins (TNF-α, NF-κB p65). Immunohistochemistry showed that TMT resulted in elevated PK2 and PKR2 protein expression in the CA2 and CA3 regions of the hippocampus in mice, while PKR1 protein was not significantly altered. Network pharmacology showed that PK2 could interact with the intersectional targets of ARS, CNS injury, and TMT. ARS remarkably attenuated TMT-induced seizures and hippocampal histological damage. Further studies demonstrated that ARS treatment attenuated TMT-induced hippocampal ultrastructural damage, possibly by increasing the number of rough endoplasmic reticulum and mitochondria as well as upregulating the levels of synapse-associated proteins (PSD95, SYN1, Synaptophysin). Western blotting results revealed that ARS downregulated TMT-induced TNF-α and NF-κB p65 protein levels. In addition, ARS also decreased TMT-induced protein expression of PK2 and PKR2 in the mouse hippocampus, but had no significant effect on PKR1 protein expression. Our results suggested that ARS ameliorated TMT-induced abnormal neural behavior and hippocampal injury, which may be achieved by regulating PK2/PKRs inflammatory pathway and ameliorating synaptic injury. Therefore, we suggest that PK2/PKRs pathway may be involved in TMT neurotoxicity and ARS may be a promising drug that can relieve TMT neurotoxicity.
神经炎症是三甲基锡（TMT）中枢神经毒性的重要机制之一。青蒿素（ARS）是一种著名的抗疟药物，具有显著的抗炎作用。促动力蛋白 2（PK2）是一种小分子分泌蛋白，广泛表达于神经系统，在神经炎症的发生发展中起着关键作用。然而，青蒿素是否能改善 TMT 引起的神经炎症，以及 PK2/PKRs 信号通路是否参与其中，目前尚不清楚。本研究中，雄性 Balb/c 小鼠腹腔注射 TMT（2.8mg/kg）后，通过免疫组织化学评估海马 CA2 和 CA3 区 PK2、PKR1 和 PKR2 蛋白的表达。网络药理学用于预测 ARS、中枢神经系统（CNS）损伤和 TMT 的交集靶点。通过行为评分评估小鼠的神经行为。通过 HE、尼氏和电子显微镜评估海马的组织病理学损伤。Western blot 用于鉴定突触相关蛋白（PSD95、SYN1、Synaptophysin）、PK 系统相关蛋白（PK2、PKR1、PKR2）和炎症相关蛋白（TNF-α、NF-κB p65）的表达。免疫组织化学显示，TMT 导致小鼠海马 CA2 和 CA3 区 PK2 和 PKR2 蛋白表达升高，而 PKR1 蛋白无明显变化。网络药理学显示，PK2 可与 ARS、CNS 损伤和 TMT 的交集靶点相互作用。青蒿素显著减轻 TMT 诱导的惊厥和海马组织学损伤。进一步研究表明，青蒿素治疗可减轻 TMT 诱导的海马超微结构损伤，可能通过增加粗面内质网和线粒体的数量以及上调突触相关蛋白（PSD95、SYN1、Synaptophysin）的水平来实现。Western blot 结果显示，青蒿素下调 TMT 诱导的 TNF-α和 NF-κB p65 蛋白水平。此外，青蒿素还降低了 TMT 诱导的小鼠海马 PK2 和 PKR2 蛋白表达，但对 PKR1 蛋白表达无明显影响。我们的结果表明，青蒿素改善了 TMT 诱导的异常神经行为和海马损伤，这可能是通过调节 PK2/PKRs 炎症通路和改善突触损伤来实现的。因此，我们认为 PK2/PKRs 通路可能参与 TMT 神经毒性，青蒿素可能是一种缓解 TMT 神经毒性的有前途的药物。
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