Global-Scale Profiling of Differential Expressed Lysine-Lactylated Proteins in the Cerebral Endothelium of Cerebral Ischemia–Reperfusion Injury Rats

Acute ischemic stroke (AIS) is a serious threat to human health. Following AIS, cerebral ischemia–reperfusion injury (CIRI) must be treated to improve prognosis. By combining 4D label-free quantitative proteomics with lactylation modification-specific proteomics analysis, we assessed lysine lactylation (Kla) in cortical proteins of a CIRI rat model. We identified a total of 1003 lactylation sites on 469 proteins in this study, gathering quantitative information (PXD034232) on 660 of 310 proteins, which were further classified by cell composition, molecular function, and biological processes. In addition, we analyzed the metabolic pathways, domains, and protein–protein interaction networks. Lastly, we evaluated differentially expressed lysine lactylation sites, determining 49 upregulated proteins and 99 downregulated proteins with 54 upregulated sites and 54 downregulated sites in the experimental group in comparison with the healthy control group. Moreover, we identified the Kla of Scl25a4 and Slc25a5 in the Ca2+ signaling pathway, but the Kla of Vdac1 was eliminated, as confirmed in vivo. Overall, these results provide new insights into lactylation involved in the underlying mechanism of CIRI because this post-translational modification affects the mitochondrial apoptosis pathway and mediates neuronal death. Therefore, this study may enable us to develop new molecules with therapeutic properties, which have both theoretical significance and broad clinical application prospects.Graphical AbstractA new model of cerebral ischemia–reperfusion injury (CIRI) induced by lactylation through the regulation of key proteins of the Ca2+ signaling pathway.