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Kidney Week

Abstract: TH-PO471

Macrophages Mediate Renal Fibrosis via C5AR1 Signaling

Session Information

  • CKD: Mechanisms - I
    November 07, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
    Abstract Time: 10:00 AM - 12:00 PM

Category: CKD (Non-Dialysis)

  • 2103 CKD (Non-Dialysis): Mechanisms

Authors

  • Sahu, Ranjit K., University of Virginia, Charlottesville, Alabama, United States
  • Xavier, Sandhya, University of Virginia, Charlottesville, Alabama, United States
  • Chauss, Daniel, NIH/NIDDK, Bethesda, Maryland, United States
  • Afzali, Behdad, King's College London, London, United Kingdom
  • Portilla, Didier, University of Virginia Health System, Charlottesville, Virginia, United States
Background

Macrophages regulate innate immunity, inflammation, metabolism and tissue repair. We reported increased expression of intracellular complosome components, C1q, C3, C5 and anaphylatoxin receptors C3aR1 and C5aR1 in macrophages isolated from kidney tissue of mice receiving folic acid (FA) or unilateral ureteral obstruction (UUO). Recently, single cell transcriptomic study performed in UUO mice identified new proximal tubule sub clusters, but also increased expression of C5aR1 in UUO macrophages. We hypothesized that deletion of C5aR1 in macrophages reduces kidney fibrosis

Methods

C5aR1GFPfl/fl mice were generated by inserting LoxP sites flanking exon 2 as previously reported. C5aR1GFPfl/fl mice were crossed with mice expressing Cre-recombinase under control of the LysM promoter to generate LysM-Cre-C5aR1GFPfl/fl mice. LysM-Cre-C5aR1GFPfl/fl mice and C5aR1GFPfl/fl (control mice) received intraperitoneal injections of either vehicle (sodium bicarbonate) or FA in sodium bicarbonate. Two weeks later kidney tissue was harvested for analysis. Macrophages expressing C5aR1 and C5aR1 negative control macrophages were isolated from C5aR1GFPfl/fl mice by flow cytometry following UUO-injury and we performed RNA seq analysis of their trancriptome

Results

Flow studies and confocal microscopy using C5aR1GFPfl/fl reporter mice confirmed that macrophages are the dominant expressors of C5aR1 in both models of fibrosis. RNAseq analysis of GFP+ macrophages isolated from C5aR1GFPfl/fl mice subjected to UUO confirmed increased expression of C5aR1 mRNA, as well as increased transcripts encoding markers of inflammation, including Tlr4 and Cxcl13. GFP+ macrophages also exhibit increased expression of iron homeostatic genes, iron transporters and iron-recycling transcription factors. Immunohistochemistry, flow cytometry, qRT-PCR and western blots demonstrated reduced inflammation and fibrosis in whole kidney tissues from mice with selective deletion of C5aR1 in macrophages (LysMCreC5aR1GFPfl/fl) compared to control mice treated with FA.

Conclusion

Increased expression of C5aR1 in macrophages represents an important pathophysiologic mechanism leading to increased tissue inflammation and organ fibrosis. We propose that increased iron-recycling and overload in C5aR1+ macrophages represents an important cellular mechanism leading to kidney fibrosis.

Funding

  • NIDDK Support