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Induction of A_2AR in nephritic glomeruli. (A) RNase protection analysis was performed to determine A_2AR mRNA expression in the glomeruli of anti-GBM glomerulonephritis in WKY rats. A_2AR was induced from day 3 onwards during anti-GBM glomerulonephritis. The expression of A_2AR peaked at day 5 and started to decrease by day 7. Rat ribosomal L-32 gene was used as a housekeeping gene. Probe contains polylinker regions and is longer than the protected bands. The expression levels of A_2AR mRNA are expressed relative to the mRNA levels of the housekeeping gene L-32. Data shown is a representative of three separate experiments. (B) Double immunohistochemical staining of ED1⁺ cells (brown color) overlap with A_2AR (red color), thus A_2AR was mainly in Mϕ. (C) Interestingly, A_2AR (red color) was not expressed in CD8+ cells (brown color) or (D) monocytes (red color) located in extraglomerular vessels. (B) and (C), right panel, show ED1⁺ or CD8⁺ cells staining after removal of red color (A_2AR staining) from (B) and (C) slides at left panel to demonstrate the overlap of A_2AR expression in ED1+ but not in CD8+ cells. Sections for staining were sampled on day 7 after anti-GBM antibody injection.