Abstract: TH-PO0290
S1PR4 Overexpression Activates SPHK2 and Drives Intracellular S1P-Mediated Cytokine Dysregulation in Podocytes
Session Information
- Glomerular Diseases: Cell Biology
October 22, 2026 | Location: Exhibit Hall A, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Glomerular Diseases
- 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology
Authors
- Insenga, Arianna, University of Miami Department of Medicine, Miami, Florida, United States
- Njeim, Rachel, University of Miami Department of Medicine, Miami, Florida, United States
- Molina David, Judith T., University of Miami Department of Medicine, Miami, Florida, United States
- Tolerico, Matthew, University of Miami Department of Medicine, Miami, Florida, United States
- Fontanella, Antonio Miguel, University of Miami Department of Medicine, Miami, Florida, United States
- Zevola, Mario, University of Miami Department of Medicine, Miami, Florida, United States
- Fornoni, Alessia, University of Miami Department of Medicine, Miami, Florida, United States
- Merscher, Sandra, University of Miami Department of Medicine, Miami, Florida, United States
Background
Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid that signals through five cell surface G-protein-coupled receptors (S1PR1–5). Intracellular S1P has receptor-independent functions, including inhibition of HDAC1/2. Mutations in S1P lyase, the enzyme that degrades S1P in the cell, cause nephrotic syndrome, suggesting intracellular S1P accumulation contributes to podocyte injury. We recently showed that S1PR4 expression is upregulated in Alport Syndrome, and that S1PR4 antagonism protects from podocyte injury in vitro and in vivo. However, the exact mechanism by which S1PR4 contributes to podocyte injury remains elusive. We hypothesized that elevated S1PR4 activates intracellular S1P generation by sphingosine kinase 2 (SPHK2), thereby driving podocyte injury in a receptor-independent mechanism.
Methods
Human podocytes with S1PR4 expression (S1PR4 OE) or empty vector (EV) were generated by lentiviral infection. Bulk RNA-seq, RT-qPCR, and Western blotting were employed to assess S1PR4 signaling pathways. Intracellular S1P was quantified by LC-MS/MS. SPHK2 expression and activation were assessed by Western blot and the phospho-SPHK2/total SPHK2 ratio respectively. Pharmacological rescue used the S1PR4 antagonist CYM50358 and the SPHK2 inhibitor Opaganib.
Results
S1PR4 OE podocytes showed elevated phospho-SPHK2 and increased intracellular S1P. Bulk RNA-seq revealed a significant inflammatory signature, with induction of IL1B, IL6, CCL2, and CXCL chemokines, upregulation of galectin-3 and galectin-9 indicative of lysosomal stress, and engagement of necroptosis-associated transcripts (MLKL, CASP8) without classical apoptotic activation in OE podocytes. p62/SQSTM1 was elevated, consistent with impaired autophagic flux. Interestingly, there was a dissociation of two sister cytokines: IL-1β was induced at mRNA level, while IL-18 mRNA and protein were nearly abolished. Treatment with either opaganib or CYM50358 fully restored IL-18 expression, implicating S1PR4-driven SPHK2 activation and intracellular S1P accumulation as the upstream driver of IL-18 silencing.
Conclusion
S1PR4 overexpression activates SPHK2 to elevate intracellular S1P in association to significant changes in the inflammatory signature of podocytes. We identify a novel S1PR4, SPHK2, intracellular S1P axis linking surface receptor signaling to receptor-independent intracellular S1P toxicity.
Funding
- NIDDK Support