Mir199a-3p and 5p Influence Fibrotic Disease States, And Increase In Expression After Radiation

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The mir199a family, composed of 3p and 5p splice variants, are associated with fibrotic cardiac, lung and renal diseases of different etiologies. Mir199a-3p and 5p target vital negative regulators of fibrosis-inducing pathways including JAK/STAT via targeting of SOCS7, and caveolin-dependent internalization pathway through targeting of CAV1. The objective of this study was to investigate if mir199a 3p and 5p expression increase after irradiation (RT) and regulate radiation-induce fibrosis via the similar pathways. C57BL/6 were treated with focal 14Gy renal irradiation and kidneys and overlaying skin harvested after 24hrs, 10wks, or 20wks. Untreated mouse kidneys and skin served as age-matched controls. RNA was isolated from the skin and kidney tissue using a trizol extraction method, and quantitative PCR analysis was performed to assess gene expression changes in mir199a-3p and 5p and critical components of the JAK/STAT and caveolin pathways. Both mir199a-3p and 5p expression increased in kidneys at the 20wks, with increases over untreated controls of 2.1-fold (p<0.001) and 2.4-fold (p<0.001) respectively. A 10-fold increase in expression of mir199a precursor RNA was seen for skin at 20 wks, and this translated into a 3.1-fold increase of mir199a-3p (p = 0.046) but without a measurable increase in the mir199a-5p variant. SOCS7 expression levels decreased at 20wk consistent with mir199a targeting, although not significantly when comparing to untreated controls. Levels of CAV1 increased significantly over untreated controls at the 10wk (1.8-fold, p = 0.04). At 20 weeks, transcript levels of CAV1 decreased to a level no longer significantly different than in age-matched controls, consistent with mir199a targeting. We show that mir199a family members are elevated in the kidneys and overlaying skin after 14Gy of radiation and mechanistically-associated with fibrosis. On-going studies are evaluating mir199a-3p and 5p after more clinically-relevant doses of RT.