Figure 1.

Arg1^fl/fl;LysMCre/⁺ mice have reduced arginase-1 in the kidney after ischemic injury. Eight-week-old male Arg1^mko (Arg1^fl/fl;LysM^Cre/+) mice underwent IRI, and kidney lysates were collected for RNA and protein on day 0 and day 7 after surgery. (A) Arg1 mRNA expression level (relative to Hprt1) was significantly decreased in Arg1^mko mice at day 7 within the whole kidney (*P<0.05, t test). (B and C) Western blot analysis (B) demonstrates significantly decreased arginase-1 protein levels on day 7 after IRI in Arg1^mko mice, quantified in (C). Each lane represents one mouse kidney (*P<0.05, t test). (D) CD45⁺F4/80⁺ macrophages were isolated from kidneys at the indicated time by FACS and Arg1 mRNA expression quantified by quantitative rtPCR. Arg1 mRNA is significantly lower in day 7 macrophages from Arg1^mko compared with control mice (**P<0.01, t test). (E) Immunofluorescence staining of injured kidneys from Arg1^con mice (top panels) and Arg1^mko mice (bottom panels) on day 7 after IRI (arginase-1 [red], F4/80⁺ [green], DAPI [blue]) shows significantly reduced arginase-1 staining in F4/80⁺ macrophages from the Arg1^mko mouse. The boxed area in the left hand panels is magnified in the right panels. Asterisks indicate arginase-1⁺ and F4/80⁺ macrophage; arrows indicate arginase-1⁻ and F4/80⁺ macrophage. (F) Quantification of total F4/80⁺ cells demonstrates increased medullary macrophages on day 7 after IRI Arg1^con mice with a modest reduction seen in Arg1^mko mice. (**P<0.01, ANOVA). (G) Quantification of arginase-1⁺ cells shows an 84% reduction in the Arg1^mko mice. (****P<0.0001, ANOVA). n=5 mice per group. Casts exhibit green autofluorescence.