Abstract: TH-PO357

PolyIC Induces the Expression of Retinoic Acid-Inducible Gene I and Melanoma Differentiation-Associated Gene 5 in Podocytes and Modulates Inflammatory Responses and Podocyte Damage

Session Information

Category: Cell Biology

  • 201 Cell Signaling, Oxidative Stress

Authors

  • Narita, Ikuyo, Hirosaki University, Hirosaki, aomori, Japan
  • Shimada, Michiko, Hirosaki University, Hirosaki, aomori, Japan
  • Fujita, Takeshi, Hirosaki University, Hirosaki, aomori, Japan
  • Murakami, Reiichi, Hirosaki University, Hirosaki, aomori, Japan
  • Nakamura, Norio, Hirosaki University, Hirosaki, aomori, Japan
  • Saleem, Moin, University of Bristol, Bristol, United Kingdom
  • Mathieson, Peter W., University of Bristol, Bristol, United Kingdom
  • Tomita, Hirofumi, Hirosaki University, Hirosaki, aomori, Japan
Background

Viral infection often exacerbates proteinuria and activation of innate immunity in renal cells is suggested as pathogenesis. As well as some of the toll-like receptors, retinoic acid-inducible gene-I (RIG-I)-like helicase receptors (RLRs) recognize double-stranded RNA (dsRNA) produced during viral replication. RLRs are located in the cytoplasm, and RIG-I and melanoma differentiation-associated gene 5 (MDA5) are the members of RLRs. It is reported that dsRNA induces the expression of RIG-I and MDA5 in mesangial cells, however, the effect on podocytes is not well elucidated. In this study, we tested the effect of polyinosinic polycytidylic acid (polyIC) on the expressions of RIG-I and MDA5 and on the down-stream inflammatory responses and podocyte damages.

Methods

Conditionally immortalized human podocytes were grown in 33 degrees centigrade and differentiated in 37 degrees centigrade, then treated with 2 to 500 µg/ml of polyIC, synthesized dsRNA for 3 to 36h. The expression levels of RIG-I and MDA5 were assessed by quantitative RT-PCR and western blotting. The expression of IFN-β, TNFα and IL-6 were assessed by quantitative RT-PCR. We further tested the role of RIG-I and MDA5 by the temporal knockdown utilizing siRNA. F-actin staining was performed to assess actin re-organization as a feature of podocyte damages.

Results

PolyIC induced the expression of RIG-I and MDA5 in podocytes in dose and time dependent manners, as demonstrated by quantitative RT-PCR and western blotting. PolyIC also increased mRNA expression of IFN-β, TNFα and IL-6. PolyIC induced actin re-organization by F-actin staining. Temporal knockdown of RIG-I by siRNA resulted in decreased expression of IFN-β and TNFα induced by polyIC, while temporal knockdown of MDA5 by siRNA inhibited IFN-β, TNFα and IL-6.

Conclusion

PolyIC dramatically induced the expression of RIG-I and MDA5 in podocytes in dose and time dependent manners. Temporal silencing of RIG-I and MDA5 by siRNA significantly suppressed the expression of inflammatory cytokines induced by polyIC leading to podocyte damages. These results suggest that not only TLRs but also RLRs play an important role in podocyte damage during viral infection.