Abstract
OBJECTIVES: To investigate the effects of Qingshen Granules (QSG) on folic acid (FA)-induced renal fibrosis in mice and TGF-β1-stimulated HK-2 cells and the underlying mechanism. METHODS: A mouse model of FA-induced renal fibrosis were treated daily with QSG at 4.0 g·kg(-)¹ via gavage for 4 weeks. Renal histopathological changes were evaluated using HE and Masson staining, and renal expressions of fibrosis markers, key glycolytic pathway proteins, and histone lactylation-related proteins (Pan Kla, and lactylation sites of H3/H4) were detected using Western blotting. Renal expressions of α-SMA, FN, E-cad, LDHA, and H3K18 lactylation (H3K18la) were also examined with immunofluorescence staining. Serum creatinine (SCR) and blood urea nitrogen (BUN) were measured by ELISA, and renal lactate content was determined with colorimetric assay. In HK-2 cells stimulated with TGF-β1, the effects of QSG-medicated serum from SD rats on cell viability, proliferation, protein expressions of α‑SMA, E-cad, LDHA, and H3K18la, extracellular oxygen consumption rate (OCR), extracellular acidification rate (ECAR), and intracellular lactate levels were analyzed. RESULTS: The mouse models of renal fibrosis showed increased renal expressions of α‑SMA, FN, HIF-1α, HK2, PFKM, PKM2, LDHA, Pan Kla, and H3K18la, elevated serum SCR and BUN and renal lactate level, and decreased E-cad expression. QSG treatment effectively reversed these changes and alleviated renal fibrosis in the mouse models. Immunofluorescence staining showed that QSG treatment significantly reversed the elevation of renal α‑SMA, FN and LDHA expressions and reduction of LDHA/H3K18la co-localization, and enhanced E-cad expression. In HK-2 cells, TGF‑β1 treatment significantly reduced cell viability, proliferation and OCR, and increased ECAR, lactate, α‑SMA, LDHA, and H3K18la, which were all reversed by treatment with QSG-medicated serum. Exogenous lactate obviously reversed the inhibitory effects of QSG on these proteins. CONCLUSIONS: QSG alleviates renal fibrosis in mice by modulating glycolytic reprogramming, reducing lactate accumulation, and inhibiting H3K18la.