Department of Pathogen Biology, Anhui Province Key Laboratory of Microbiology & Parasitology, Anhui Provincial Laboratory of Zoonoses of High Institutions, School of Basic Medicine, Anhui Medical University, Hefei, China.
Department of Orthopaedics, the First Affiliated Hospital of Anhui Medical University, Anhui, China.
Parasit Vectors. 2023 Jan 25;16(1):30. doi: 10.1186/s13071-022-05618-8.
Alzheimer's disease presents an abnormal cognitive behavior. TgCtwh6 is one of the predominant T. gondii strains prevalent in China. Although T. gondii type II strain infection can cause host cognitive behavioral abnormalities, we do not know whether TgCtwh6 could also cause host cognitive behavioral changes. So, in this study, we will focus on the effect of TgCtwh6 on mouse cognitive behavior and try in vivo and in vitro to explore the underlying mechanism by which TgCtwh6 give rise to mice cognitive behavior changes at the cellular and molecular level.
C57BL/6 mice were infected orally with TgCtwh6 cysts. From day 90 post-infection on, all mice were conducted through the open field test and then Morris water maze test to evaluate cognitive behavior. The morphology and number of cells in hippocampus were examined with hematoxylin-eosin (H&E) and Nissl staining; moreover, Aβ protein in hippocampus was determined with immunohistochemistry and thioflavin S plaque staining. Synaptotagmin 1, apoptosis-related proteins, BACE1 and APP proteins and genes from hippocampus were assessed by western blotting or qRT-PCR. Hippocampal neuronal cell line or mouse microglial cell line was challenged with TgCtwh6 tachyzoites and then separately cultured in a well or co-cultured in a transwell device. The target proteins and genes were analyzed by immunofluorescence staining, western blotting and qRT-PCR. In addition, mouse microglial cell line polarization state and hippocampal neuronal cell line apoptosis were estimated using flow cytometry assay.
The OFT and MWMT indicated that infected mice had cognitive behavioral impairments. The hippocampal tissue assay showed abnormal neuron morphology and a decreased number in infected mice. Moreover, pro-apoptotic proteins, as well as BACE1, APP and Aβ proteins, increased in the infected mouse hippocampus. The experiments in vitro showed that pro-apoptotic proteins and p-NF-κBp65, NF-κBp65, BACE1, APP and Aβ proteins or genes were significantly increased in the infected HT22. In addition, CD80, pro-inflammatory factors, notch, hes1 proteins and genes were enhanced in the infected BV2. Interestingly, not only the APP and pro-apoptotic proteins in HT22, but also the apoptosis rate of HT22 increased after the infected BV2 were co-cultured with the HT22 in a transwell device.
Neuron apoptosis, Aβ deposition and neuroinflammatory response involved with microglia polarization are the molecular and cellular mechanisms by which TgCtwh6 causes mouse cognitive behavioral abnormalities.
阿尔茨海默病表现出异常的认知行为。TgCtwh6 是中国流行的主要弓形虫株之一。尽管弓形虫 II 型菌株感染可导致宿主认知行为异常，但我们不知道 TgCtwh6 是否也会导致宿主认知行为改变。因此，在这项研究中，我们将重点研究 TgCtwh6 对小鼠认知行为的影响，并试图在体内和体外从细胞和分子水平探讨 TgCtwh6 引起小鼠认知行为改变的潜在机制。
C57BL/6 小鼠经口感染 TgCtwh6 包囊。从感染后第 90 天开始，所有小鼠均进行旷场试验和 Morris 水迷宫试验，以评估认知行为。用苏木精-伊红（H&E）和尼氏染色观察海马的形态和细胞数量；此外，用免疫组织化学和硫黄素 S 斑块染色检测海马中的 Aβ 蛋白。用 Western blot 或 qRT-PCR 检测海马中的突触结合蛋白 1、凋亡相关蛋白、BACE1 和 APP 蛋白和基因。用 TgCtwh6 速殖子分别攻击海马神经元细胞系或小鼠小胶质细胞系，然后分别在孔中培养或在 Transwell 装置中共培养。用免疫荧光染色、Western blot 和 qRT-PCR 分析靶蛋白和基因。此外，用流式细胞术检测小鼠小胶质细胞系的极化状态和海马神经元细胞系的凋亡。
旷场试验和水迷宫试验表明，感染小鼠的认知行为受损。海马组织学检测显示，感染小鼠的神经元形态异常，数量减少。此外，感染小鼠海马中的促凋亡蛋白以及 BACE1、APP 和 Aβ 蛋白增加。体外实验显示，感染 HT22 的细胞中促凋亡蛋白和 p-NF-κBp65、NF-κBp65、BACE1、APP 和 Aβ 蛋白或基因显著增加。此外，感染 BV2 的细胞中 CD80、促炎因子、Notch、hes1 蛋白和基因也增强。有趣的是，不仅是 HT22 中的 APP 和促凋亡蛋白，而且是在 Transwell 装置中与 HT22 共培养的感染 BV2 后 HT22 的凋亡率也增加了。
神经元凋亡、Aβ 沉积和小胶质细胞极化相关的神经炎症反应是 TgCtwh6 引起小鼠认知行为异常的分子和细胞机制。
