Abstract
PURPOSE: Macrophages with an anti-inflammatory phenotype are critical for resolving inflammation and preventing chronic tissue injury. Estradiol is known to promote this favorable macrophage profile. This study evaluated the therapeutic potential of the secretome, delivered as conditioned medium, from estradiol-treated macrophages in experimental rheumatoid arthritis (RA) in Wistar rats. METHODS: Rheumatoid arthritis was induced in Wistar rats using complete Freund's adjuvant. Animals were assigned to five groups: healthy controls, arthritic rats receiving vehicle, arthritic rats treated with prednisolone, arthritic rats treated with conditioned medium from untreated macrophages, and arthritic rats treated with conditioned medium from estradiol-exposed macrophages. The lyophilized media were administered intraperitoneally on days 4, 12, and 20 post-induction; the study ended on day 24. RESULTS: Conditioned medium from estradiol-treated macrophages exhibited significantly higher levels of anti-inflammatory mediators such as interleukin-10 (IL-10), transforming growth factor-beta, and indoleamine 2,3-dioxygenase, along with increased messenger RNA expression of regulatory genes including early growth response 2 and mannose receptor. In vivo, this treatment notably reduced arthritis severity and improved weight gain compared to medium from untreated macrophages. These effects correlated with a marked decrease in antigen-specific proliferation and serum levels of inflammatory markers such as C-reactive protein (CRP), myeloperoxidase (MPO), nitric oxide (NO), IL-1, and tumor necrosis factor-alpha. Additionally, bone-destructive factors like receptor activator of nuclear factor kappa-B ligand (RANKL) and matrix metalloproteinase-9 (MMP-9) were significantly downregulated in treated rats. CONCLUSION: The conditioned medium derived from estradiol-treated macrophages, enriched with anti-inflammatory and regulatory components, presents a promising cell-free therapeutic strategy for immunotherapy in RA.