H3K18 lactylation is involved in the PANoptosis of brain microvascular endothelial cells induced by hypoxia
ZHENG Mingze
LI Ming
GUAN Ruili
LIU Xinqin
Abstract：Objective To explore the mechanism of lactylation in the occurrence of PANoptosis in brain microvascular endothelial cells induced by hypoxia.Methods A total of 20 male C57BL/6 mice aged 8 weeks were selected to establish an animal model simulating a hypoxic environment at an altitude of 6 000 m for 28 d.The 20 mice were randomly divided into normoxic group and hypoxic group(10 mice in each group).An in vitro model of bEnd.3 cell line exposure to normoxia(210 mL/L O2)and hypoxia(10 mL/L O2)for 24 h was constructed.Immunofluorescence labeling of CD31 and GSDMD was performed on the hippocampal tissue of the mice.Transmission electron microscopy was used to observe the ultrastructural changes of brain microvascular endothelial cells in the hippocampal tissue.qRT-PCR was utilized to detect the expression of PANoptosis-related molecules such as ZBP1,NLRP3,NLRP12,ASC,CASP4,and GSDMD in hippocampal tissue and bEnd.3 cells.Western blotting was used to detect the expression of GSDMD,Pan-Kla,and H3K18 lactylation(H3K18la)in hippocampal tissue and bEnd.3 cells.A lactate assay kit was used to measure the L-lactic acid(L-LA)content in hippocampal tissue and bEnd.3 cells,and a lactate dehydrogenase(LDH)assay kit was used to determine the release of LDH in peripheral blood and bEnd.3 cells.Further,sodium oxamate(OXA)was used to inhibit lactate metabolism in bEnd.3 cells,followed by detection of L-LA,LDH,and the expression levels of the aforementioned PANoptosis molecules.Results Immunofluorescence staining results indicated that the number of CD31+GSDMD+cells in the hippocampal tissue of the hypoxic group was increased compared to the normoxic group.Transmission electron microscopy revealed characteristic pyroptotic endothelial cells in the hippocampal microvessels of the hypoxic group.The levels of lactate(P＜0.01)and LDH(P＜0.01)in the peripheral blood and hippocampal tissue of the hypoxic group were higher than those in the normoxic group.qRT-PCR results showed that the expression of ZBP1(P＜0.01),NLRP3(P＜0.01),NLRP12(P＜0.01),ASC(P＜0.01),CASP4(P＜0.01),and GSDMD(P＜0.01)in the hippocampal tissue of the hypoxic group was significantly higher than that in the normoxic group.Western blotting showed that the levels of Pan-Kla(P＜0.01)and H3K18la(P＜0.01)in the hippocampus of the hypoxic group were higher than those of the normoxic group.Compared to the normoxic group,the expression of GSDMD protein in bEnd.3 cells in hypoxic group for 24 h increased,and the levels of Pan-Kla(P＜0.01)and H3K18la(P＜0.01)were elevated.The content of lactate(P＜0.01)and LDH(P＜0.01)was higher than that of the normoxic group.Compared to bEnd.3 cells in the hypoxic group,the lactate content(P＜0.05)and LDH release(P＜0.05)were reduced,and the damage was relatively alleviated in the hypoxia+OXA group,with corresponding decrease in the expression of PANoptosis molecules(P＜0.05).Conclusion Hypoxic exposure can induce the occurrence of PANoptosis in brain microvascular endothelial cells,and H3K18la may serve as an important target involved in hypoxia-induced PANoptosis of brain microvascular endothelial cells.
Keywords：hypoxiacognitive disordersneuroinflammationbrain microvascular endothelial cellsPANoptosisprotein lactylation
Publication Date：2025-05-31
Online Publishing Date：2026-08-26(First online date of this platform, not the publication date of the document)
Pages：8( 583-590 )