Sources for anti‐inflammatory annexin A1 signals during acute anti‐Thy‐1.1 nephritis

Abstract

Leukocyte infiltration presents a hallmark feature of immune‐mediated acute kidney injury and defective resolution of inflammation may promote progression towards chronic kidney failure. Leukocytes exert distinctive effects during this process and may either aggravate kidney damage or foster renal repair. The glucocorticoid‐inducible protein Annexin A1 has been shown to induce resolution of inflammation and to shift macrophage polarization towards the anti‐inflammatory and repair‐promoting M2 phenotype. The cellular source of the putative Annexin A1 signal and its regulation during the course of renal inflammation remains to be elucidated.Adult Wistar rats were injected with an antibody directed against the Thy‐1.1 protein located on mesangial cells to induce mesangioproliferative glomerulonephritis. Animals were examined after 24h (initiation phase), 5d (proliferation phase), and 15d (resolution phase). Regulation of Annexin A1 was studied by qPCR and immunohistochemistry. Annexin A1+ cells were characterized by triple labelling immunofluorescence using antibodies for CD68 (monocytes and macrophages), CD206 (macrophages with M2 polarization), CD3, CD4, and CD8 (T‐cell subpopulations), CD20 (B‐lymphocytes), and myeloperoxidase (neutrophil granulocytes). Quantification of immunoreactive cells was performed by cell counting on confocal micrographs.Renal Annexin A1 mRNA levels increased rapidly after induction of anti‐Thy‐1.1 nephritis (24h: +92±16%; 5d: +128±19%; 15d: +78±33% relative to controls; p<.05). Immunofluorescence labelling showed an interstitial accumulation of total, and CD206+ M2 macrophages during the proliferation and resolution phase (5d: +150±40% and +260±29%; 15d: +378±43% and +600±100% relative to controls; p<.05). Elevated numbers of CD68+/CD206+/Annexin A1+ interstitial M2 macrophages were revealed by triple labelling studies at d5 and d15 (+240±27% and +450±80% relative to controls; p<.05). Abundant expression of Annexin A1 was further found in CD4+ and CD8+ T‐lymphocytes but their number remained unaffected by renal inflammation. CD20+ B‐lymphocytes were negative for Annexin A1. Granulocytes were found in subset of glomeruli at 24h and d5. All granulocytes stained positive for Annexin A1.In conclusion, we have shown a significant increase of Annexin A1 gene expression and protein abundance during the course of anti‐Thy‐1.1 nephritis. The increased amount of Annexin A1+ leukocytes in animals with anti‐Thy‐1.1 nephritis suggest that these cells present a major source for anti‐inflammatory and pro‐resolving signals during renal inflammation.Support or Funding InformationDeutsche Forschungsgemeinschaft; FOR1368