Regulation Of Hyaluronan Metabolism Attenuates Organ Fibrosis
Xinyi Wang1, Swathi Balaji1, Alexander Blum1, Hui Li1, Emily Steen1, Natalie Templeman1, Paul Bollyky2, Sundeep Keswani1.
1BCM, Houston, TX, USA, 2Stanford University, Stanford, CA, USA.
Background: Organ fibrosis is a common disease endpoint that confers high morbidity. We and others have reported that skin injury alters hyaluronan (HA) metabolism, and that HA regulation by IL-10 or HA-synthase(HAS1) encoding for anti-inflammatory HA subtypes promotes regenerative wound healing. Here, we postulate that HA dysregulation is central to fibrosis and further hypothesis that the regulation of altered HA metabolism attenuates renal fibrosis. Methods: Dermal and renal fibroblasts (FB) were isolated from C57BL/6J mice to determine the effect of IL-10(100 ng/ml) with/without TGFb (to induce a fibrotic phenotype), and/or the HA inhibitor, 4-methylumbelliferone(4MU), on expression of HAS1-3 and hyaluronidases (HYAL1-2). We performed unilateral ureteral obstruction(UUO) as a renal fibrosis model with/without IL-10 or HAS1(1x1010IU) overexpression through the injection under the kidney capsule. UUO/sham kidneys were collected at d3, d7, d14 for RNA, ELISA, and immunohistochemistry. HA metabolism levels were assessed by qPCR, Western blot, and ELISA. HA molecular weight was assessed by gel electrophoresis. Data mean+/-SD; p-values by ANOVA. Results: In vitro: TGFb significantly upregulated gene expression of collagen-1(p<0.05) in both FB cell lines, as expected; expression of HAS1-3 in skin FB (3.23±0.37, 1.73±0.23, 5.25±1.10), and HAS1&3 and HYAL1 in renal FB (6.62±0.89, 1.83±0.54, 1.84±0.92) was also significantly dysregulated (p<0.05). 4MU treatment abrogated TGFb-induced fibrotic phenotypes in both FBs. IL-10 restored HAS gene levels in both FBs (1±0.42 fold). In vivo: UUO significantly altered HA synthase and hyaluronidases, resulting in dysregulated HA metabolism. HAS gene expression levels spiked at d7 in UUO and HYAL1 increased 6-fold at d3 (p<0.05), while HA expression peaked at d14 (4.72x103 ng/ml). IL-10, but not HAS1, regulated post-translational levels of HA, attenuated scarring, and reduced α-SMA relative to controls. Conclusions: Our findings demonstrate that regulation of HA metabolism can attenuate post-injury fibrosis in multi-organ fibrosis models, identifying a potential common biology. Moreover, our discovery of mechanisms behind the HA-attenuated fibrosis could inspire novel therapeutics.
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