Effects of ENO1 knockdown and LPS+Lactate stimulation on EA.HY926 cells
Elevated lactate was associated with vascular endothelial cells (ECs) dysfunction in sepsis. However, the underlying mechanisms remain to be defined. In this study, we found that elevated lactate in sepsis promotes lactylation of enolase 1 (ENO1) at K71 site through the action of lactyltransferase P300. The lactylation at ENO1 K71 site reduces the binding of TRIM21(an E3 ubiquitin ligase) mRNA to ENO1 (an RNA-binding protein), resulting in enhanced TRIM21 mRNA stability and expression. Elevated TRIM21 can bind to VE-Cadherin and exert its E3 ubiquitin ligase function, causing the ubiquitination and degradation of VE-Cadherin, which disrupts endothelial adhesions junctions (AJs) and increases endothelial permeability. Targeting lactylation of ENO1 K71 via a specific peptide alleviated endothelial injury and enhanced survival rates among septic mice. These findings suggest that lactyaltion of ENO1 plays a pivotal role in vascular endothelial dysfuncyion during sepsis, while inhibiting lactylation may offer a novel strategy for sepsis treatment. Overall design: To investigate the function of ENO1 lactylation in endothelial cells after LPS and lactate stimulation, we conduct four groups: 1.Negtive control(NC), without stimulation; 2.LPS+lactate stimulation (NC-L-L); 3.ENO1 knockdown with siRNA (siENO1); 4.LPS+lactate stimulation after ENO1 knockdown with siRNA (siENO1-L-L).