Mechanism study of Tacrolimus inhibiting FXR mediated toxicity of renal tubular epithelial cells

Objective: Tacrolimus-induced nephrotoxicity represents a significant limitation to the long-term success of renal transplantation, with currently insufficient preventive and therapeutic strategies in clinical settings. The hallmark pathological manifestation of this nephrotoxicity is vacuolization within renal tubular epithelial cells; however, the precise mechanisms underlying this effect remain inadequately elucidated. The farnesoid X receptor (FXR), a nuclear transcription factor, has been demonstrated to confer protection against nephrotoxicity induced by agents such as vancomycin and cisplatin. This study aims to delineate the specific mechanisms by which FXR modulates tacrolimus-induced toxicity in renal tubular epithelial cells and to provide clinical insights for mitigating tacrolimus-associated nephrotoxicity.
Methods: Renal biopsy samples were obtained from five matched pairs comprising healthy donors and renal transplant recipients who developed tacrolimus-induced nephrotoxicity. Histopathological evaluation was conducted using hematoxylin and eosin (HE) staining, periodic acid–schiff (PAS) staining, and electron microscopy. Renal function was assessed via serum creatinine (SCr), blood urea nitrogen (BUN), and urinary N-acetyl-β-D-glucosaminidase (NAG) measurements. FXR expression within renal tissue was quantified. To investigate the role of FXR, a renal tubular epithelial cell-specific FXR knockout mouse model and a model with renal overexpression of FXR mediated by AAV9 vectors were established. These models were subjected to tacrolimus-induced nephrotoxicity to evaluate functional impact of FXR. Transcriptomic sequencing was performed to identify signaling pathways altered by tacrolimus exposure and FXR knockout, with subsequent validation through quantitative real-time PCR (qRT-PCR), western blotting, immunofluorescence and co-immunoprecipitation (co-IP). Additionally, the FXR agonist GW4064 was treated to mice with tacrolimus-induced nephrotoxicity to assess the therapeutic potential of FXR activation.
Results: Both clinical specimens and mouse models of tacrolimus-induced nephrotoxicity demonstrated vacuolization of renal tubular epithelial cells, concomitant with reduced FXR protein and mRNA expression in renal tissue. Targeted knockout of FXR in renal tubular epithelial cells exacerbated tacrolimus-induced vacuolization, whereas renal-specific FXR overexpression ameliorated this phenotype. Transcriptomic analysis revealed that tacrolimus-indued nephrotoxicity was associated with disruptions in cell adhesion and focal adhesions (FAs) disassembly. FXR knockout suppressed the Apelin signaling pathway, enhanced plasma membrane translocation of aquaporin-2 (AQP2), and upregulated integrin alpha 8 (ITGA8), a marker indicative of focal adhesions disassembly. Treatment with the FXR agonist GW4064 effectively reversed the nephrotoxicity induced by tacrolimus.
Conclusion: Tacrolimus impairs the FXR-Apelin signaling axis, leading to upregulation of AQP2-mediated pathways, modulation of focal adhesions molecule expression, and subsequent disassembly of focal adhesions in renal tubular epithelial cells. These molecular alterations promote epithelial cell shedding and vacuolization, thereby contributing to tacrolimus-induced nephrotoxicity.