Ogerin mediated inhibition of TGF-β(1) induced myofibroblast differentiation is potentiated by acidic pH

Abstract

Transforming growth factor beta (TGF-β) induced myofibroblast differentiation is central to the pathological scarring observed in Idiopathic Pulmonary Fibrosis (IPF) and other fibrotic diseases. Our lab has recently identified expression of GPR68 (Ovarian Cancer Gene Receptor 1, OGR1), a pH sensing G-protein coupled receptor, as a negative regulator of TGF-β induced profibrotic effects in primary human lung fibroblasts (PHLFs). We therefore hypothesized that small molecule activators of GPR68 would inhibit myofibroblast differentiation. Ogerin is a positive allosteric modulator (PAM) of GPR68, inducing a leftward shift of the dose response curve to proton induced signaling. Using PHLFs derived from patients with both non-fibrotic and IPF diagnoses, we show that Ogerin inhibits, and partially reverses TGF-β induced myofibroblast differentiation in a dose dependent manner. This occurs at the transcriptional level without inhibition of canonical TGF-β induced SMAD signaling. Ogerin induces PKA dependent CREB phosphorylation, a marker of Gαs pathway activation. The ability of Ogerin to inhibit both basal and TGF-β induced collagen gene transcription, and induction of Gαs signaling is enhanced at an acidic pH (pH 6.8). Similar findings were also found using fibroblasts derived from dermal, intestinal, and orbital tissue. The biological role of GPR68 in different tissues, cell types, and disease states is an evolving and emerging field. This work adds to the understanding of Gαs coupled GPCRs in fibrotic lung disease, the ability to harness the pH sensing properties of GPR68, and conserved mechanisms of fibrosis across different organ systems.

Persistent hypoxia promotes myofibroblast differentiation via GPR‐81 and differential regulation of LDH isoenzymes in normal and idiopathic pulmonary fibrosis fibroblasts

“…The increased expression of HIF‐1α supports the premise that fibroblasts within fibroblastic foci and the fibrotic interstitium of IPF tissue are subjected to cellular hypoxia due to impaired diffusion (Higgins et al, 2007). Whether due to metabolic reprogramming and aerobic glycolysis or to impaired oxygen diffusion to fibroblasts in the alveolar interstitium, elevated lactic acid levels have been found in lung tissues from IPF patients (Bell et al, 2022; Kottmann et al, 2012). However, the precise mechanism(s) driving lactate generation in fibrotic lung fibroblasts and the potential role of lactate‐induced cellular signaling on fibroblast phenotype are poorly understood.…”

“…For example, an acidic microenvironment can liberate latent TGF‐β from the extracellular matrix and contribute to fibrosis by increasing the pool of active TGF‐β (Kottmann et al, 2012). However, a recent study also reported that an acidic pH regulates Ogerin (GPR‐68)‐mediated inhibition of TGF‐β signaling (Bell et al, 2022), demonstrating the complexity of TGF‐β activation and signaling responsiveness. GPR‐68 is a G‐protein‐coupled receptor, and GPRs comprise the largest transmembrane receptor family in humans (Bockaert & Pin, 1999).…”

“…GPR‐68 is a G‐protein‐coupled receptor, and GPRs comprise the largest transmembrane receptor family in humans (Bockaert & Pin, 1999). As a class, GPRs are implicated in a variety of human diseases and targeting G‐protein‐coupled receptors and their downstream signaling has been proposed for the treatment of lung fibrosis, although GPRs can activate pro‐fibrotic or anti‐fibrotic signaling depending on the interaction of the associated G‐protein isoforms (Bell et al, 2022; Choi et al, 2021; Haak et al, 2020; Walker & Fisher, 2014; Zmajkovicova et al, 2020). Lactate is the ligand of GPR‐81 (HCAR1) (Kuei et al, 2011).…”

Recent advances in acid sensing by G protein coupled receptors

“…OGR1 has been implicated in allergen-induced hyper-responsiveness and idiopathic pulmonary fibrosis [ 3 , 10 , 56 , 57 ] and was reported to be pro-inflammatory [ 29 , 56 ]. It is expressed in lung fibroblasts [ 3 , 56 ] and bronchial smooth muscle cells [ 10 , 29 ].…”

“…OGR1 has been implicated in allergen-induced hyper-responsiveness and idiopathic pulmonary fibrosis [ 3 , 10 , 56 , 57 ] and was reported to be pro-inflammatory [ 29 , 56 ]. It is expressed in lung fibroblasts [ 3 , 56 ] and bronchial smooth muscle cells [ 10 , 29 ]. Whilst OGR1 expression is increased in bronchial smooth muscle cells in an ovalbumin antigen-challenged murine asthma model [ 10 ], it is downregulated in fibroblasts from patients with idiopathic pulmonary fibrosis [ 65 ], which promotes myofibroblast differentiation, suggesting that OGR1 has anti-fibrotic effects in lung fibroblasts [ 56 ].…”