Abstract: TH-PO010

Increased Expression of Complement Receptor C5aR1 in Macrophages Contributes to Kidney Fibrosis

Session Information

Category: Cell Biology

  • 204 Extracellular Matrix Biology, Fibrosis, Cell Adhesion

Authors

  • Sahu, Ranjit K., University of Virginia Health System , Charlottesville, Virginia, United States
  • Xavier, Sandhya, University of Virginia Health System , Charlottesville, Virginia, United States
  • Landes, Susan G., University of Virginia Health System , Charlottesville, Virginia, United States
  • Taylor, Ronald P., University of Virginia Health System , Charlottesville, Virginia, United States
  • Koehl, Joerg, University of Luebeck, Luebeck, Germany
  • Portilla, Didier, University of Virginia Health System , Charlottesville, Virginia, United States
Background

We recently demonstrated increased expression and activation of the complement system with increased expression of C5 and C5aR1 in whole kidney tissue homogenates of mice subjected to folic acid and Unilateral ureteral obstruction (UUO) injury. Given the central role of complement activation product C5a in the development of the inflammatory response through interaction with C5aR1, in the present studies we examined the cellular localization of C5aR1 during injury, and the potential mechanism(s) by which its activation contributes to kidney fibrosis.

Methods

C5aR1 expression was studied by immunohistochemistry, flow cytometry, qPCR, and by western blot analysis of wild type, GFP-C5aR1-floxed, and C3-/- mice subjected to the UUO model. RAW264.7 cells were used as an in vitro model to study how increased C5aR1 expression on macrophages was linked to the process of fibrosis.

Results

Immunohistochemistry studies localized expression of C5aR1 to proximal tubules in sham mice but increased expression was seen mostly in interstitial cells after UUO. C5aR1 mRNA levels were increased in CD45+ cells isolated from UUO mice. Flow cytometry analysis using GFP/C5aR1-Floxed mice demonstrated a 75-fold increase of GFP+C5aR1+CD11b+, F4/80+, LyC6- macrophages after UUO. Reduced staining of GFP+-C5aR1+ Prominin-1+ cells by flow suggests loss of renal tubular epithelial cells during UUO. C5aR1 expression as well as kidney fibrosis were significantly reduced in C3 null mice when compared to wild type mice subjected to UUO. Our studies suggest that increased C5aR1 expression on macrophages during UUO is associated with increased kidney fibrosis. In vitro studies using RAW 264.7 cells treated with LPS showed increased C5aR1 protein expression associated with increased production of IL1β and IL6, as well as inflammasome activation, all indicative of an increased inflammatory response.

Conclusion

Our results are the first to report that increased expression of C5aR1 on CD11b+ F4/80+ LyC6- macrophages correlates with the advent of kidney fibrosis. Increased cytokine production and inflammasome activation represent cellular mechanisms by which increased C5aR1 expression contributes to kidney fibrosis.

Funding

  • NIDDK Support