nc886 Regulates Actin Dynamics and Maintains Podocyte Integrity in Idiopathic Nephrotic Syndrome and Viral Infection Models

Background: Idiopathic nephrotic syndrome (INS) is a major cause of nephrotic syndrome in children, characterized by massive proteinuria. The involvement of T cell-derived humoral factors has long been suggested in INS. We previously reported that changes in DNA methylation in naive helper T cells (Th0s) closely correlated with the pathophysiological status in INS. Using a genome-wide approach, we further investigated the changes of DNA methylation profiles in Th0s and identified a differentially methylated region that includes a noncoding RNA gene, nc886. Building on these findings, we conducted experiments using a human podocyte cell line to elucidate the role of nc886 in INS and viral infection models. Methods: We employed InfiniumR HumanMethylation450K for genome-wide DNA methylation analysis in Th0s isolated from participants. nc886 expression was measured using quantitative real-time PCR. We stimulated ciPods with polyinosinic-polycytidylic acid (poly(I:C)) as viral infection model, and with patient serum at relapse and remission as INS model. F-actin was quantitated using an immunofluorescence antibody technique to assess actin reorganization in ciPods. We produced nc886 knockdown (KD) ciPods using antisense oligos. Protein kinase RNA-activated (PKR) phosphorylation was analyzed via western blotting utilizing an anti-phospho PKR antibody, to assess the functional effect of nc886 KD in ciPods. Results: The individuals who had a high DNA methylation ratio in nc886 region, known as a polymorphic imprinting region, in peripheral Th0s, were significantly increased in the patient group. Although the methylation ratio was not significantly different between Th0s derived at relapse and at remission, the nc886 expression level significantly lower in the relapse sample. In ciPods, the nc886 expression levels decreased following stimulation with INS relapse serum, and with poly(I:C), respectively, inducing actin depolymerization. In nc886 KD ciPods, the PKR activation significantly increased and the fluorescence intensity of F-actin was significantly decreased. Conclusion: Decreased nc886 expression in podocytes results in actin depolymerization, inducing podocyte dysfunction and proteinuria in INS and viral infection model. This reserch provides new insights into podocyte dysfunction. Funding: Government Support – Non-U.S.