Podocyte-Proximal Tubule Interactions in Diabetic Kidney Disease

While glycemic control, RAAS blockade, and SGLT2i are important therapies, a significant proportion of these individuals with diabetic kidney disease (DKD) still progress to end-stage kidney disease. Along with endothelial injury and mesangial expansion, podocyte loss compromises the glomerular filtration barrier, leading to initial kidney injury in DKD. Although podocyte loss is the inciting event, recent studies suggest that the severity of proximal tubule (PT) injury is the primary determinant of DKD progression. In this proposal, the hypothesis is that the signaling between podocytes and PT cells is critical to preventing the progression of DKD. To ascertain how podocytes regulate PT health in DKD, mice with podocyte-specific expression of human Krüppel-Like Factor 6 (KLF6), a zinc finger transcription factor critical for podocyte homeostasis, was generated. These mice exhibited significantly less podocyte loss, glomerulosclerosis, PT injury, and interstitial fibrosis than the control mice under diabetic conditions. While it was initially suspected this improvement in PT injury was due to a reduction in podocyte loss, single nuclei RNA-sequencing demonstrated that podocyte KLF6 primes Ca2+/calmodulin-dependent protein kinase ID (CaMK1D) signaling in the first segment of the PT, proximal to the podocytes. The podocyte secretome from the KLF6 mice mitigated mitochondrial injury in PT cells under high glucose conditions, but the genetic and pharmacological inhibition of Camk1d in PT cells negated these protective effects by exacerbating mitochondrial injury and reducing cell survival. Utilizing a combination of tandem mass spectrometry from the podocyte secretome and single nuclei ATAC-sequencing, it was observed that KLF6 triggers the release of the N-glycosylated apolipoprotein J (ApoJ) from podocytes by inducing the expression of genes critical to N-glycosylation. Interestingly, previous studies report increased glomerular ApoJ expression and urinary ApoJ levels in individuals with early DKD. ApoJ also binds to low- density lipoprotein-related protein 2 (LRP2) and undergoes cellular uptake in PT cells. Subsequently, ApoJ binds to calmodulin to activate CaMKK-CaMK1D signaling to attenuate mitochondrial injury under diabetic conditions. Based on these data, the hypothesis is that the podocyte preconditions the PT against injury through ApoJ-CaMK1D signaling to mitigate the progression of DKD. This hypothesis will be tested through the following specific aims: (1) Determine the salutary role of N-glycosylated ApoJ signaling in DKD, (2) Ascertain how ApoJ primes CaMK1D signaling in PT cells in DKD, and (3) Establish how the activation of CaMK1D signaling preconditions the PT cells against injury in DKD. This proposal will address a current gap in the field by elucidating novel mechanism(s) by which the podocyte regulates PT health in DKD. The long-term goal of our project is to identify the mechanism(s) that mitigate PT injury in DKD so it can be leveraged as a therapeutic strategy for individuals with rapidly progressing DKD.