SMPDL3b Overexpression Is Associated with STING Activation in Podocytes in CKD

Background: Chronic kidney disease (CKD) is a global health challenge marked by rising incidence, prevalence, and poorly understood pathogenesis. Our prior work suggests that sphingolipids, especially sphingomyelin phosphodiesterase acid-like 3b (SMPDL3b), regulate podocyte function and survival. Sphingolipids also modulate inflammation, a key driver of CKD progression. Our recent findings link STING activation to podocyte foot process disruption, proteinuria, and CKD progression. Here, we aim to explore if increased SMPDL3b expression leads to podocyte injury through chronic STING activation. Methods: Illumina sequencing RNA data analysis, qRT-PCR and Western blot analysis were used to characterize the immortalized control (CTRL) or overexpression (SMP OE) human podocytes. Cells were treated with c-diAMP (10μM), a STING specific agonist, for 24h. 8-week-old mice with podocyte specific Smpdl3b deficiency (pSMPfl/fl) or overexpression (pSMPTg) were used to evaluate glomerular STING activation. pSMPfl/fl and pSMPTg mice were intraperitoneally injected with a single dose of c-diAMP, 50mg/kg or 5% DMSO and sacrificed 72h after injection, followed by urinary albumin-to-creatinine ratio (ACR), histological and serum analyses. Two-tailed t-test or One-Way ANOVA followed by Tukey’s post-test were used to detect statistical changes. Results: We observed significant changes in the expression of genes related to cytosolic DNA sensing pathways in SMP OE podocytes. Both in vitro and in vivo experiments revealed elevated levels of STING phosphorylation (pSTING). Treatment with c-diAMP increased pSTING levels in control podocytes but not in SMP OE podocytes. While baseline proteinuria was absent in both pSMPfl/fl and pSMPTg mice, administration of c-diAMP led to a notable increase in ACR specifically in pSMPTg mice. No significant changes in body weight or levels of serum BUN and creatinine in either pSMPfl/fl or pSMPTg mice following c-diAMP treatment. TEM analysis revealed FPE in control and pSMPTg mice treated with c-diAMP, but not in pSMPfl/fl mice. Conclusion: Our data indicate that SMPDL3b overexpression is associated with STING activation in podocytes in vitro and STING-dependent proteinuria in vivo. Funding: Private Foundation Support