Modulation of the Apolipoprotein M/S1PR4 Pathway Reduces Podocyte Lipid Overload in Alport Syndrome via Distinct Autophagy and Efflux Mechanisms.

Abstract

We identified dysregulation of the apolipoprotein M/sphingosine-1-phosphate/sphingosine-1-phosphate receptor 4 (S1PR4) axis in the glomeruli and podocytes of a mouse model of Alport syndrome. Exogenous apolipoprotein M or antagonism of S1PR4 was sufficient to prevent kidney failure, podocyte injury, and lipid accumulation. Apolipoprotein M reduced lipid accumulation in podocytes via cholesterol efflux, while S1PR4 antagonism promoted autophagy of lipid droplets. Renal lipid dysmetabolism contributes to glomerular disease progression, including Alport syndrome. We recently identified alterations in the apolipoprotein M (APOM)/sphingosine-1-phosphate (S1P)/S1P receptor 4 signaling axis in glomeruli from patients with glomerular disease. We used Col4a3 knockout mice and immortalized podocytes derived from these mice as a mouse model of Alport syndrome. Mice and podocytes were treated with recombinant APOM or the S1P receptor 4-specific antagonist, CYM50358. Col4a3-/- glomeruli and podocytes exhibited reduced APOM and increased S1P receptor 4 expression and increased sphingosoine-1-phosphate levels, mirroring findings in patients with glomerular disease. Treatment with APOM or CYM50358 reduced albuminuria, BUN, and plasma creatinine and ameliorated glomerulosclerosis, tubulointerstitial fibrosis, podocyte loss, and foot process effacement. Both treatments reduced triglyceride and cholesterol accumulation in glomeruli and podocytes. RNA-seq analysis of Col4a3-/- revealed that S1P receptor 4 antagonism upregulated lysosomal and autophagy-related genes. Western blot analysis confirmed increased LC3-II/LC3-I ratios and decreased p62, indicating enhanced autophagic flux. Treated podocytes showed increased lysosome numbers and colocalization with lipid droplets. By contrast, APOM had no effect on autophagy but promoted cholesterol efflux. Furthermore, knockdown of APOM or overexpression of sphingosine-1-phosphate receptor 4 was sufficient to cause podocyte cell death. We found that the APOM/S1P axis was dysregulated in Col4a3-/- podocytes. Targeting this pathway through APOM supplementation or S1P receptor 4 antagonism improved kidney function and reduced lipid accumulation by enhancing either cholesterol efflux or autophagy, respectively.