Wuling San regulates AVPR2-cAMP-PKA-CREB pathway to delay cellular senescence and ameliorate acute kidney injury

ETHNOPHARMACOLOGICAL RELEVANCE: Cellular senescence in renal resident cells plays a pivotal role in the progression of acute kidney injury (AKI), necessitating the expansion of effective drug targets. Traditional Chinese medicine (TCM) formulations, characterized by their multi-target effects, offer a promising perspective for advancing research on AKI. Wuling San (WLS), a well-established compound used in treating urological disorders, has yet to elucidate its potential pharmacological targets and mechanisms in ameliorating AKI and delaying cellular senescence. AIM OF THE STUDY: This study sought to elucidate the mechanisms by which WLS modulates the AVPR2-cAMP-PKA-CREB pathway to mitigate cellular senescence and promote recovery from AKI. METHODS: We first prepared WLS-containing serum and performed RT-qPCR experiments to screen for GPCRs that were differentially expressed in response to WLS. Next, we established an in vitro AKI mouse model to assess the renal protective effects of the WLS by measuring renal function, renal pathology, and oxidative stress levels. After this, we performed RNA sequencing (RNA-Seq) profiling to identify differentially expressed genes (DEGs) affected by WLS treatment. We also conducted Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses to identify potential signaling pathways involved. We then utilized the Gene Expression Omnibus (GEO) data to screen for cellular senescence related differentially expressed genes (CSRDEGs) in AKI patients and performed enrichment analysis, as well as a joint analysis of specific genes in relation to the RNA-Seq profiling results. We also examined how WLS affects the expression of proteins linked to cellular senescence in the AKI mouse model by targeting the AVPR2-cAMP-PKA-CREB pathway. RESULTS: WLS markedly enhanced the expression of Arginine Vasopressin Receptor 2 (AVPR2) and ameliorated renal function indicators, as well as pathological changes and oxidative stress levels in the mouse model of AKI. RNA-Seq profiling revealed significant enrichment of the cAMP signaling pathway following WLS intervention. Bioinformatics analysis indicated that genes associated with cellular senescence in AKI patients were notably enriched in the p53 signaling pathway. Data mining from the GEO database, in conjunction with RNA-Seq profiling, demonstrated a substantial reduction in key genes after WLS treatment. Additionally, WLS elevated both the expression and phosphorylation of pivotal proteins within the AVPR2-cAMP-PKA-CREB pathway, while concurrently decreasing proteins associated with cellular senescence. CONCLUSION: The results demonstrated that WLS significantly elevated the expression of AVPR2, which may underlie its nephroprotective effects and facilitate the mitigation of AKI by modulating the AVPR2-cAMP-PKA-CREB pathway, ultimately contributing to a delay in cellular senescence.