Background/Purpose: G-protein coupled receptor kinase 5, GRK5, is a central regulator of G protein-coupled receptors (GPCRs), a vast family of cell surface receptors involved in a variety of physiological activities. GRK5 acts by phosphorylating GPCRs, causing desensitization and internalization. In addition, GRK5 exerts GPCR-independent effects and can enter the nucleus to modulate the expression of target genes. GRK5 has previously been implicated in the pathophysiology of inflammation and immunological responses, migration and invasion of cancer cells, neurodegenerative diseases and post-ischemic remodeling of the heart. However, its role in rheumatologic diseases and in particular in fibrotic tissue remodeling has not been investigated so far. Here, we tested the hypothesis that GRK5 might modulate the transcription of profibrotic target genes in fibroblasts to promote fibrotic tissue remodeling.

Methods: GRK5 expression was examined in SSc patients and experimental models of SSc using real-time PCR, Western Blot and immunofluorescence staining. GRK5 expression was modulated in vitro and in vivo with knockdown and knockout techniques and adenoviral overexpression. The antifibrotic efficacy of GRK5 inactivation was investigated in three SSc animal models: bleomycin-induced dermal and bleomycin-induced pulmonary fibrosis and fibrosis produced by overexpression of a constitutively active TGF receptor type I (TBRI^act). Target genes of GRK5 in fibroblasts were identified by RNA-Seq.

Results: Increased mRNA levels and nuclear accumulation of GRK5 protein were observed in fibroblasts in fibrotic skin and lung samples compared to non-fibrotic control samples. GRK5 expression was also upregulated in murine models of SSc. TGFβ, but not the other profibrotic or proinflammatory mediators such as EGF, FGF9 or IL-1, induced GRK5 overexpression. Knockdown of GRK5 in fibroblasts reduced their sensitivity to the profibrotic effects of TGFβ with impaired fibroblast-to-myofibroblast transition and decreased collagen synthesis. Similarly, fibroblast-specific knockout of GRK5 demonstrated potent antifibrotic effects in the murine models of bleomycin-induced dermal and bleomycin-induced pulmonary fibrosis. In addition, GRK5 knockout mice were also protected from TBRI^act-induced fibrosis. Overexpression of GRK5 in fibroblasts, on the other hand, caused fibroblast-to-myofibroblast transition and collagen release. In a mouse model of bleomycin-induced dermal fibrosis, adenoviral overexpression of GRK5 exacerbated skin fibrosis. RNA-Seq unraveled GRK5 as a novel upstream regulator of numerous profibrotic genes including COL1A1, ACTA2, CTGF and PAI-1. Mechanistically, RNA-Seq also revealed that GRK5 modulated signal transducer and activator of transcription 3 (STAT3) and histone deacetylases (HDACs) activity to induce fibroblast activation.

Conclusion: We demonstrate that GRK5 is sufficient and required for fibroblast activation in response to TGFβ and serves as an upstream regulator of multiple profibrotic signals. Inactivation of GRK5 exerts potent antifibrotic effects in various preclinical models of SSc. Inactivation of GRK5 may thus offer potential as a target for antifibrotic therapies.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/g-protein-coupled-receptor-kinase-5-in-fibrotic-tissue-remodeling/