# Targeting the lactylation of ENO1 alleviates endothelial dysfunction in sepsis

## TL;DR

This study shows that inhibiting ENO1 lactylation can reduce endothelial damage and improve survival in sepsis by preventing TRIM21 from disrupting blood vessel integrity.

## Contribution

The study identifies ENO1 lactylation as a novel mechanism in sepsis-induced endothelial dysfunction and proposes a targeted therapeutic strategy.

## Key findings

- ENO1 lactylation at K71 prevents TRIM21 mRNA degradation, increasing TRIM21 levels.
- TRIM21 binds to VE-Cadherin, causing its degradation and increasing endothelial permeability.
- Inhibiting ENO1 lactylation with a peptide reduces endothelial injury and improves survival in septic mice.

## Abstract

Elevated lactate is associated with vascular endothelial dysfunction, a factor that can contribute to organ failure in sepsis. However, the specific mechanisms involved have yet to be fully elucidated. Here, we investigated the role of enolase 1 (ENO1) lactylation in modulating the functions of endothelial cells (ECs) in sepsis pathogenesis.

The septic mouse model was established using two methods: cecal ligation and puncture (CLP) and intraperitoneal injection of LPS. AAV‐ENO1 shRNA was administered to ablate ENO1 in vascular endothelial cells of mice. Tail vein injection of .5% Evans Blue Dye (EBD) was utilised to assess microvascular permeability in septic mice. Post‐translational modification (PTM) mass spectrometry was employed to detect key proteins undergoing lactylation in endothelial cells. Additionally, CCK‐8 assay, Transwell assay, and scratch wound healing assay were performed to evaluate the fundamental functions of ECs. Further investigations were conducted through Western blotting, Co‐immunoprecipitation (CO‐IP), RT‐qPCR, RNA immunoprecipitation (RIP) and RNA sequencing to examine genes/proteins involved in vascular endothelial injury and their interactions.

We found that elevated lactate in sepsis promoted the lactylation of ENO1 at the K71 residue, facilitated by the increased activity of the lactyltransferase P300. This modification reduced the binding of TRIM21 mRNA to ENO1, thereby preventing its degradation by limiting the recruitment of CNOT6. Consequently, the stability and expression of TRIM21 mRNA were enhanced. Elevated TRIM21 subsequently binds to vascular endothelial‐cadherin (VE‐Cadherin), promoting its ubiquitination and degradation, disrupting endothelial adherens junctions (AJs) and increasing endothelial permeability. Targeting the lactylation of ENO1 at K71 with a specific inhibitory peptide alleviated endothelial injury and improved survival rates in septic mice.

These findings suggest that ENO1 lactylation plays a pivotal role in vascular endothelial dysfunction during sepsis. Inhibiting lactylation may offer a therapeutic strategy for sepsis treatment.

Lactylation of ENO1 reduced the binding of TRIM21 mRNA to ENO1, thereby preventing mRNA degradation. Elevated TRIM21 subsequently binds to VE‐Cadherin, promoting its ubiquitination. Targeting the lactylation of ENO1 with a specific inhibitory peptide alleviated endothelial dysfunction.