Yao Chunhong, Wang Jinfeng
Department of Geriatrics, People's Hospital of Dongxihu District, 48 Jinbei 1st Road, Jinghe Street, Wuhan, 430040, Hubei, China.
Sci Rep. 2025 Jul 13;15(1):25312. doi: 10.1038/s41598-025-07811-y.
The aim of this study was to investigate the effects and mechanisms of ANGPTL4 on cognitive impairment in vascular dementia rats. 36 SD rats were randomly divided into Sham(n= 9), VaD(n= 9), VaD + ANGPTL4 OE(n= 9), and VaD + ANGPTL4 KD(n= 9). A bilateral carotid artery ligation (2-VO) rat VaD was established to study the effects of ANGPTL4. Spatial memory was tested in rats using the Morris water maze. Morphological changes of neurons were detected in the CA1 region of the hippocampus by hematoxylin-eosin staining. The expression of ANGPTL4, p-Syk in the cells of hippocampal CA1 area was also detected by immunohistochemistry. Afterwards, protein expression of ANGPTL4, p-Syk, p-JNK, BNIP3 was detected by Western blot (WB). Afterwards, the mechanism of ANGPTL4 effect on cognitive impairment in vascular dementia rats was further explored by ANGPTL4 OE hippocampal cells 1%O low-serum low-glucose stimulation with Syk inhibitor, JNK inhibitor. ANGPTL4 OE aggravated cognitive dysfunction in 2VO rats. ANGPTL4 KD treatment improved the memory performance of 2VO rats.Hippocampal tissue damage was obvious in the VaD group.Hippocampal tissue damage was aggravated in the ANGPTL4 OE group (P < 0.001).ANGPTL4 KD group Pathological features were significantly improved(P < 0.001). WB assay showed that the expression of ANGPTL4, p-Syk, p-JNK, and BNIP3 proteins was increased in 2VO rats(P < 0.05), which was further up-regulated by ANGPTL4 OE treatment, and significantly inhibited by ANGPTL4 KD treatment in 2VO rats(P < 0.05). In vitro cellular experiments revealed that ANGPTL4 OE treatment again up-regulated ANGPTL4, integrin, p-Syk, and p-JNK protein expression consistent with the in vivo results(P < 0.05), where the Syk inhibitor suppressed both p-Syk, and p-JNK protein expression(P < 0.05). It is also worth noting that the mitochondrial autophagy-related proteins BNIP3, PINK1, Parkin and JC-1 mitochondrial membrane potential assayed by wb revealed that ANGPTL4 OE treatment exacerbated mitochondrial stress(P < 0.05) and apoptosis(P < 0.05) in hippocampal cells, and that Syk inhibitor, and JNK inhibitor significantly inhibited the modulation of ANGPTL4 OE(P < 0.05). ANGPTL4 promotes mitochondrial autophagy and apoptosis in the hippocampal CA1 region by activating the integrin/p-Syk signalling pathway, thus aggravating cognitive impairment in vascular dementia rats.
本研究旨在探讨血管生成素样蛋白4（ANGPTL4）对血管性痴呆大鼠认知功能障碍的影响及机制。将36只SD大鼠随机分为假手术组（n = 9）、血管性痴呆组（VaD，n = 9）、血管性痴呆+ANGPTL4过表达组（VaD + ANGPTL4 OE，n = 9）和血管性痴呆+ANGPTL4基因敲低组（VaD + ANGPTL4 KD，n = 9）。通过双侧颈总动脉结扎（2-VO）建立大鼠血管性痴呆模型，以研究ANGPTL4的作用。采用Morris水迷宫测试大鼠的空间记忆。通过苏木精-伊红染色检测海马CA1区神经元的形态学变化。采用免疫组织化学法检测海马CA1区细胞中ANGPTL4、磷酸化脾酪氨酸激酶（p-Syk）的表达。随后，通过蛋白质免疫印迹法（WB）检测ANGPTL4、p-Syk、磷酸化应激活化蛋白激酶（p-JNK）、BNIP3的蛋白表达。之后，用Syk抑制剂、JNK抑制剂对ANGPTL4过表达的海马细胞进行1%低血清低糖刺激，进一步探讨ANGPTL4对血管性痴呆大鼠认知功能障碍的作用机制。ANGPTL4过表达加重了2-VO大鼠的认知功能障碍。ANGPTL4基因敲低治疗改善了2-VO大鼠的记忆表现。VaD组海马组织损伤明显。ANGPTL过表达组（ANGPTL4 OE）海马组织损伤加重（P < 0.001）。ANGPTL4基因敲低组（ANGPTL4 KD）病理特征明显改善（P < 0.001）。WB检测显示，2-VO大鼠中ANGPTL4、p-Syk、p-JNK和BNIP3蛋白表达增加（P < 0.05），ANGPTL4过表达处理使其进一步上调，而ANGPTL4基因敲低处理在2-VO大鼠中显著抑制其表达（P < 0.05）。体外细胞实验表明，ANGPTL4过表达处理再次上调了ANGPTL4、整合素、p-Syk和p-JNK蛋白表达，与体内结果一致（P < 0.05），其中Syk抑制剂抑制了p-Syk和p-JNK蛋白表达（P < 0.05）。还值得注意的是，通过WB检测线粒体自噬相关蛋白BNIP3、PTEN诱导激酶1（PINK1）、帕金蛋白（Parkin）和JC-1线粒体膜电位发现，ANGPTL4过表达处理加剧了海马细胞的线粒体应激（P < 0.05）和细胞凋亡（P < 0.05），而Syk抑制剂和JNK抑制剂显著抑制了ANGPTL4过表达的调节作用（P < 0.05）。ANGPTL4通过激活整合素/p-Syk信号通路促进海马CA1区线粒体自噬和细胞凋亡，从而加重血管性痴呆大鼠的认知功能障碍。
