Lactate controls glomerular endothelial barrier integrity in lupus nephritis
Jiaxin Lei, Xingyu Zhai, Yixin Wang, Ying Li, Lei Li, Mengdi Liu, Jing Guo, Lingyi Li, Zhezhuyun Chen, Qinghua Cao, Zhichun Liu, Ting Liu, Lin Xu, Zhenke Wen
Jiaxin Lei, Xingyu Zhai, Yixin Wang, Ying Li, Lei Li, Mengdi Liu, Jing Guo, Lingyi Li, Zhezhuyun Chen, Qinghua Cao, Zhichun Liu, Ting Liu, Lin Xu, Zhenke Wen
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Research Article Nephrology

Lactate controls glomerular endothelial barrier integrity in lupus nephritis

Abstract

Systemic lupus erythematosus (SLE) is a progressive autoimmune disease that affects multiple organs and tissues, with lupus nephritis (LN) as one of its most severe complications. Although LN progression is associated with compromised permeability of human renal glomerular endothelial cells (HRGECs), the underlying mechanisms are not fully defined. Herein, we demonstrate that aberrant glycolysis drives this glomerular endothelial barrier defect by suppressing the transcription of tight junction (TJ) genes. Mechanistically, circulating self-DNA in SLE plasma acts as a ligand that activates the cGAS/STING pathway in HRGECs, driving aberrant glycolytic adaption. The resulting glycolytic product, lactate, serves as a substrate for protein lactylation, leading to extensive lactylation and subsequent ubiquitination of enhancer of zeste homolog 2 (EZH2). In consequence, EZH2 deficiency results in reduced H3K27me3 levels, thereby suppressing the transcription of TJ genes. In a self-DNA–induced SLE model, inhibition of cGAS/STING signaling or lactate production effectively restored the integrity of TJs of HRGECs and concurrently alleviated key LN symptoms. Together, lactate programs lactylation and ubiquitination of EZH2 to impair the glomerular endothelial barrier in human SLE.

Authors

Jiaxin Lei, Xingyu Zhai, Yixin Wang, Ying Li, Lei Li, Mengdi Liu, Jing Guo, Lingyi Li, Zhezhuyun Chen, Qinghua Cao, Zhichun Liu, Ting Liu, Lin Xu, Zhenke Wen

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Figure 2

SLE plasma impairs the TJ barrier in HRGECs through glycolytic shift–driven lactate accumulation.

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SLE plasma impairs the TJ barrier in HRGECs through glycolytic shift–dri...
(A and B) Gene Set Enrichment Analysis (GSEA) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were performed on RNA-seq data derived from HRGECs treated with 10% plasma from HCs or patients with SLE. NES, normalized enrichment score. (C–E) ECs were treated with 10% plasma from HCs or patients with SLE for 24 hours. qPCR was performed to detect the mRNA expression of glycolysis-related genes (C), and Seahorse assay was used to measure ECAR (D and E). Data are mean ± SEM (3 individuals/group for E). (F) ECs were treated with 10% plasma derived from HCs or patients with SLE for 24 hours, and 2-NBDG levels were determined by flow cytometry. Mean ± SEM (6 individuals/group). (G–I) After 24 hours of treatment with SLE- or HC-derived plasma (10%), ECs were analyzed for pyruvate (G), lactate (H), and acetyl-CoA (I) using corresponding assay kits. Mean ± SEM (5–6 individuals/group). (J) After treatment with HC or SLE plasma (10%), gene expression of pyruvate dehydrogenase complex components in ECs was detected by qPCR. Mean ± SEM (5 individuals/group). (K and L) mRNA and protein levels of TJ-related proteins after SLE plasma treatment (10%) with or without 2-DG (1 mM) for 24 hours analyzed by qPCR (K) and Western blot (L). Mean ± SEM (5–6 individuals/group). (M) ECs were treated with 40 mM lactate or 10 mM lactic acid for 24 hours. Fluorescence intensity of 4 kDa FITC-dextran in the lower chamber was quantified via a microplate reader. Mean ± SEM (6 independent experiments/group). (N and O) After treatment with lactate (40 mM) or lactic acid (10 mM), mRNA and protein levels of TJ-related proteins in ECs were assessed by qPCR (N) and Western blot (O), respectively. Mean ± SEM (5 independent experiments/group). (P and Q) ECs were transfected with siRNA targeting LDHA and treated with 10% SLE plasma for 24 hours. mRNA and protein levels of TJ-related proteins were analyzed by qPCR (P) and Western blot (Q). Mean ± SEM (5 individuals/group). (R and S) ECs were treated with 10% SLE plasma with GSK2837808A (40 μM), and corresponding mRNA and protein levels were determined by qPCR (R) and Western blot (S), respectively. Mean ± SEM (5 individuals/group). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 with unpaired 2-tailed t test (E–I), 1-way (M) or 2-way ANOVA plus Tukey’s method (J–L, N, P, and R).