Mitigation of Ferroptosis in Diabetic Kidney Disease Through Mesenchymal Stem Cell Intervention via the Smad2/3/ <scp>METTL3</scp> / <scp>S1PR1</scp> Axis

Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease (ESRD) and is associated with heightened cardiovascular risk and increased overall mortality. Although mesenchymal stem cells (MSCs) have demonstrated therapeutic potential in DKD, their precise mechanisms of action have not been fully elucidated. This study aimed to investigate the involvement of Smad signaling, N6-methyladenosine (m6A) modifications, and ferroptosis in MSC-mediated treatment of DKD. Cellular and animal models of DKD were used to evaluate MSC intervention effects, supported by gene knockdown and overexpression experiments. Protein expression and phosphorylation levels of Smad2/3, m6A-associated enzymes, and markers of ferroptosis were assessed. Additionally, transcriptional targets associated with ferroptosis were identified through integrated transcriptome and m6A methylation sequencing analyses, followed by subsequent validation. Elevated Smad2/3 phosphorylation and ferroptosis were observed in DKD, while MSC interventions effectively alleviated these processes, resulting in improved renal lesions. Furthermore, MSC treatment reduced the heightened levels of m6A modification observed in DKD. Mechanistic investigations identified methyltransferase-like 3 (METTL3) as a key regulator of m6A modification in DKD. Suppression of METTL3 reversed the upregulated m6A modification and ferroptosis induced by Smad2 overexpression. Importantly, sphingosine-1-phosphate receptor 1 (S1PR1) was identified as a protective target gene against ferroptosis in DKD. In DKD models, Smad2 facilitated the m6A modification of the S1PR1 gene by interacting with METTL3 following its nuclear translocation, thereby influencing S1PR1 expression and promoting cellular ferroptosis. Intervention with MSCs mitigated this process, providing further insights into the regulatory mechanisms through which MSCs modulate ferroptosis in podocytes affected by DKD.