Department of Physiology, School of Basic Medical Sciences, Cheeloo College of Medicine, Shandong University, Jinan 250012, China.
Department of Medical Psychology and Ethics, School of Basic Medical Sciences, Cheeloo College of Medicine, Shandong University, Jinan 250012, China.
Cells. 2023 Mar 9;12(6):854. doi: 10.3390/cells12060854.
We previously found that osteopontin (OPN) played a role in hypoxia-ischemia (HI) brain damage. However, its underlying mechanism is still unknown. Bioinformatics analysis revealed that the OPN protein was linked to the lysosomal cathepsin B (CTSB) and galectin-3 (GAL-3) proteins after HI exposure. In the present study, we tested the hypothesis that OPN was able to play a critical role in the lysosomal damage of microglia/macrophages following HI insult in neonatal mice. The results showed that OPN expression was enhanced, especially in microglia/macrophages, and colocalized with lysosomal-associated membrane protein 1 (LAMP1) and GAL-3; this was accompanied by increased LAMP1 and GAL-3 expression, CTSB leakage, as well as impairment of autophagic flux in the early stage of the HI process. In addition, the knockdown of OPN expression markedly restored lysosomal function with significant improvements in the autophagic flux after HI insult. Interestingly, cleavage of OPN was observed in the ipsilateral cortex following HI. The wild-type OPN and C-terminal OPN (Leu152-Asn294), rather than N-terminal OPN (Met1-Gly151), interacted with GAL-3 to induce lysosomal damage. Furthermore, the secreted OPN stimulated lysosomal damage by binding to CD44 in microglia in vitro. Collectively, this study demonstrated that upregulated OPN in microglia/macrophages and its cleavage product was able to interact with GAL-3, and secreted OPN combined with CD44, leading to lysosomal damage and exacerbating autophagosome accumulation after HI exposure.
我们之前发现骨桥蛋白（OPN）在缺氧缺血（HI）脑损伤中发挥作用。然而，其潜在机制尚不清楚。生物信息学分析显示，OPN 蛋白在 HI 暴露后与溶酶体组织蛋白酶 B（CTSB）和半乳糖凝集素-3（GAL-3）蛋白有关。在本研究中，我们测试了以下假设：OPN 能够在新生小鼠 HI 损伤后发挥关键作用，导致小胶质细胞/巨噬细胞的溶酶体损伤。结果表明，OPN 表达增强，特别是在小胶质细胞/巨噬细胞中，并与溶酶体相关膜蛋白 1（LAMP1）和 GAL-3 共定位；这伴随着 LAMP1 和 GAL-3 表达增加、CTSB 漏出以及 HI 早期自噬流受损。此外，下调 OPN 表达可明显恢复溶酶体功能，HI 损伤后自噬流显著改善。有趣的是，HI 后同侧皮质中观察到 OPN 的切割。野生型 OPN 和 C 末端 OPN（Leu152-Asn294），而不是 N 末端 OPN（Met1-Gly151），与 GAL-3 相互作用诱导溶酶体损伤。此外，分泌型 OPN 通过与体外小胶质细胞上的 CD44 结合刺激溶酶体损伤。总之，本研究表明，小胶质细胞/巨噬细胞中上调的 OPN 及其切割产物能够与 GAL-3 相互作用，分泌型 OPN 与 CD44 结合，导致 HI 暴露后溶酶体损伤和自噬体积累加剧。
