LAUSR

The downregulation of the insulin receptor (INSR) in podocytes, leading to insulin resistance, is an important cause of diabetic kidney disease (DKD) podocyte damage. However, the mechanism is unclear. Our previous studies have shown that DNA methylation levels in the promoter region of the renal INSR gene are elevated in db/db mice. DNA methylation can inhibit gene expression. Therefore, we aim to explore: (1) whether DNA methylation participates in the downregulation of INSR in the podocytes of DKD, and (2) whether the DNA methyltransferase inhibitor can mitigate podocyte injury by upregulating INSR expression, ultimately improving DKD. In this study, 8-week-old diabetic db/db mice and age-matched wild-type mice were utilized. The db/db mice received intraperitoneal injections of DNA methyltransferase inhibitor 5-azacytidine (1 or 2 mg/kg) or PBS as control every other day for 12 weeks. Samples of mice were collected post-treatment for analysis. DNA methyltransferase 1 (DNMT1) is the key enzyme responsible for mediating DNA methylation. Treatment with 5-azacytidine significantly reduced the expression of DNA methyltransferase 1 (DNMT1) in the kidneys and podocytes of db/db mice. Furthermore, the expression of INSR in the podocytes of db/db mice was significantly decreased, accompanied by an elevation in DNA methylation levels within its promoter region. After treatment with 5-azacytidine, the DNA methylation level of INSR in the kidneys of db/db mice decreased, while the expression of INSR in podocytes increased accordingly. These findings suggest that DNA methylation plays a crucial role in the downregulation of INSR in podocytes in diabetic kidney disease (DKD). We discovered for the first time that DNA methyltransferase inhibitor could effectively decrease blood glucose levels and HOMA-IR in a dose-dependent manner in db/db mice. The decrease in AUC during IPITT and IPGTT represents an improvement in insulin sensitivity and glucose tolerance. 5-azacytidine treatment significantly suppressed the increase in kidney weight and alleviated podocyte injury in db/db mice. Concurrently, urinary albumin excretion and serum creatinine levels were significantly reduced. The expression of transcription factors NF-κB p65 and AP-1 is increased in podocytes of DKD patients and db/db mice. The inhibition of transcription factors NF-κBp65 and AP-1 can reduce inflammation-induced overexpression of DNMT1. This suggests that AP-1/NF-κBp65 may mediate the upregulation of DNMT1, leading to the downregulation of INSR in podocytes of DKD, and subsequently causing podocyte damage. Our study elucidates a novel mechanism of podocyte injury in DKD. For the first time, we have discovered that the downregulation of INSR in podocytes is associated with hypermethylation in its promoter region, leading to podocyte insulin resistance and podocyte injury. The DNA methylation inhibitor, a commonly used clinical anticancer medication, alleviated podocyte injury by reversing DNA hypermethylation-mediated inhibition of INSR in podocytes of diabetic db/db mice and dose-dependently reduced the elevated blood glucose levels. This study provides a new perspective on the pathogenesis and treatment of diabetic kidney disease from the epigenetic point of view.