Mechanism of N-butanol extract of Pulsatilla decoction protected vaginal epithelial cells against Candida albicans infection by HIF-1α signaling and glucose metabolism revealed by transcriptomics and molecular docking.

ETHNOPHARMACOLOGICAL RELEVANCE: Infection of vaginal epithelial cells by Candida albicans (C. albicans) can cause vulvovaginal candidiasis (VVC). Pulsatilla decoction has been reported and used to treat VVC, but the specific mechanism of its protective effect on cells is still unclear.

AIM OF THE STUDY: In this study, transcriptomic profiling and molecular docking were used to predict the protective effects and regulatory mechanisms of the n-butanol extract of Pulsatilla Decoction (BEPD) in vaginal epithelial cells stimulated with C. albicans hyphae, and these findings were further validated experimentally.

MATERIALS AND METHODS: To prepare the n-butanol extract from Pulsatilla decoction, transcriptome analysis was performed to identify significantly enriched genes and signaling pathways in vaginal epithelial cells stimulated with C. albicans hyphae. RT-qPCR was used to detect the expression levels of genes related to the HIF-1α pathway. Molecular docking was employed to verify the interactions between the main active components of BEPD and HIF-1α, as well as glucose metabolism-related proteins within the pathway. Immunofluorescence was used to detect the expression levels of HIF-1α, PFK2, ENO1, PDK1, Glut1, and LDHA. The cellular contents of ATP, lactic acid, and LDH were measured to reflect changes in sugar metabolism-related products. Western blot and ELISA were used to determine the levels of inflammatory factors such as IL-1β and IL-2 in vaginal epithelial cells stimulated by C. albicans hyphae and treated with BEPD.

RESULTS: BEPD exhibits a significant protective effect on vaginal epithelial cells stimulated by C. albicans hyphae. Transcriptomic analysis, including GO enrichment and KEGG pathway analysis, indicates that BEPD can regulate HIF-1α and its associated glucose metabolism disturbances in vaginal epithelial cells exposed to C. albicans hyphae. RT-qPCR confirmed the upregulation of HIF-1α gene expression in cells following BEPD treatment, while promoting the downregulation of genes related to anaerobic metabolism and attenuating the downregulation of genes involved in aerobic metabolism. Furthermore, studies on the binding interactions between drug compounds and cellular proteins revealed that the primary active components of BEPD can specifically bind to HIF-1α and proteins related to glucose metabolism. Immunofluorescence and Western blot analyses of HIF-1α and glucose metabolism-related proteins demonstrated that BEPD treatment results in significant upregulation of HIF-1α protein expression, which, in turn, markedly reduces the expression of genes associated with anaerobic metabolism and proteins involved in aerobic metabolism. ELISA assays further showed that BEPD can protect vaginal epithelial cells stimulated by C. albicans hyphae and decrease the expression levels of inflammatory factors in these cells.

CONCLUSIONS: This study demonstrates that BEPD exerts a protective effect on vaginal epithelial cells stimulated by C. albicans hyphae. The underlying mechanism may involve the regulation of HIF-1α signaling and glucose metabolism homeostasis, which represents a novel mechanism against C. albicans infection. These findings provide a new theoretical basis for the prevention and treatment of vulvovaginal candidiasis, and further studies in animal models and clinical settings are warranted in the future.