CRISPR/Cas9-Driven Fndc5 Knockout Reveals Augmented Mitochondrial Structural and Dynamic Alterations in Diabetic Nephropathy.

FNDC5 has been implicated in glucose homeostasis and is associated with mitochondrial function. Its role in diabetes and diabetic nephropathy (DN) remains unclear. This study hypothesizes that FNDC5 deficiency predisposes the kidney to accelerated mitochondrial dysfunction in diabetes and DN. Systemic Fndc5 knockout (KO) C57BL/6 mice were generated using CRISPR/Cas9. DN were induced in six-week-old Fndc5 wild-type (WT) and KO mice using high-fat diet combined with streptozotocin injection. Weekly blood and urine analyses assessed glucose, cholesterol, triglycerides, blood urea nitrogen, creatinine, and proteinuria. At 15 weeks, kidneys and metabolic tissues including pancreas, muscle and adipose were collected for histological and molecular analyses. Results showed that while both Fndc5 WT and KO mice were successfully induced with hyperglycemia, the Fndc5 KO DN group exhibited a slightly lower cumulative glycemic burden compared with the WT DN group. Despite this milder metabolic stress, proteinuria remained comparable between the two groups. Furthermore, histological analysis revealed that Fndc5 KO DN mice displayed more severe mesangial expansion, glomerular basement membrane thickening, and podocyte effacement compared with WT DN mice. The elevated lipid peroxidation, reduced PGC-1α expression, and increased DNA fragmentation were also evident in Fndc5 KO DN. More swollen mitochondria with a significantly higher percentage of disrupted cristae were observed in Fndc5 KO DN mice compared with WT DN mice. This was accompanied by the upregulation of mitochondrial fission-related genes (Dnm1l and Fis1), downregulation of the fusion-related gene (Mfn1), and reduced expression of ATP synthase subunits (ATP5A1 and ATP5B). Systemic analysis of other metabolic tissues, including the pancreas and muscle and adipose tissues, revealed increased lipid peroxidation and decreased PGC-1α expression. These findings underscore FNDC5's role in maintaining mitochondrial integrity and cellular health under diabetic conditions, positioning FNDC5 as a potential therapeutic target.