Inhibition of PKM2 by shikonin impedes TGF-β1 expression by repressing histone lactylation to alleviate renal fibrosis.

Abstract

Background: Macrophage-myofibroblast transition (MMT) plays a significant role in the progression of renal fibrosis in chronic kidney disease (CKD), making inhibition of MMT a promising therapeutic strategy. Pyruvate kinase M2 (PKM2) and its metabolite lactate are implicated in the pathogenesis of renal fibrosis; however, the mechanisms through which they contribute to this process remain poorly understood.

Purpose: To investigate the effects of PKM2 inhibition by shikonin on renal fibrosis and the underly mechanisms.

Methods: Mice were subjected to unilateral ureteral obstruction (UUO) to establish a CKD model. Renal fibrosis was assessed using histochemistry and western blotting. The MMT and histone lactylation levels were evaluated by immunofluorescence and western blotting. The interaction between the Tgfb1 promoter and lactylated histone H3 (K18) was examined using chromatin Immunoprecipitation (ChIP).

Results: PKM2 expression was significantly elevated in the renal tubular cells of UUO mouse kidneys, resulting in increased pyruvate and lactate production. Similarly, lactate levels were elevated in TGF-β1-treated TCMK-1 cells and in the serum of CKD patients. In UUO mice, treatment with shikonin, a potent PKM2 inhibitor, effectively reduced lactate production, alleviated renal fibrosis, decreased TGF-β1 expression, and suppressed the MMT process. Mechanistic studies revealed that lactate treatment stimulates Tgfb1 expression in TCMK-1 cells. Consequently, TGF-β1 in conditioned media from lactate-treated TCMK-1 cells promoted M2 macrophage polarization and upregulated fibrotic gene expression in RAW264.7 cells. Pharmacological intervention demonstrated that TGF-β1 activates the Smad3 pathway to drive the MMT process. In TCMK-1 cells, both lactate treatment and PKM2 overexpression induced Tgfb1 expression by promoting histone H3K18 lactylation.

Conclusions: Our findings indicate that PKM2-induced excessive lactate production renal tubular cells contributes to renal fibrosis. Lactate promotes histone lactylation, leading to TGF-β1 expression in these cells, which subsequently activates the Smad3 pathway in macrophages, driving the MMT and fibrosis in the kidney. Therefore, targeting PKM2, as with shikonin treatment, may represent an effective therapeutic strategy for managing renal fibrosis in CKD.