Activation of Th2 cells downregulates CRTh2 through an NFAT1 mediated mechanism

Abstract

CRTh2 (encoded by PTGDR2) is a G-protein coupled receptor expressed by Th2 cells as well as eosinophils, basophils and innate lymphoid cells (ILC)2s. Activation of CRTh2, by its ligand prostaglandin (PG)D2, mediates production of type 2 cytokines (IL-4, IL-5 and IL-13), chemotaxis and inhibition of apoptosis. As such, the PGD2-CRTh2 pathway is considered important to the development and maintenance of allergic inflammation. Expression of CRTh2 is mediated by the transcription factor GATA3 during Th2 cell differentiation and within ILC2s. Other than this, relatively little is known regarding the cellular and molecular mechanisms regulating expression of CRTh2. Here, we show using primary human Th2 cells that activation (24hrs) through TCR crosslinking (αCD3/αCD28) reduced expression of both mRNA and surface levels of CRTh2 assessed by flow cytometry and qRT-PCR. This effect took more than 4 hours and expression was recovered following removal of activation. EMSA analysis revealed that GATA3 and NFAT1 can bind independently to overlapping sites within a CRTh2 promoter probe. NFAT1 over-expression resulted in loss of GATA3-mediated CRTh2 promoter activity, while inhibition of NFAT using a peptide inhibitor (VIVIT) coincided with recovery of CRTh2 expression. Collectively these data indicate that expression of CRTh2 is regulated through the competitive action of GATA3 and NFAT1. Though prolonged activation led to NFAT1-mediated downregulation, CRTh2 was re-expressed when stimulus was removed suggesting this is a dynamic mechanism and may play a role in PGD2-CRTh2 mediated allergic inflammation.

Comparative efficacy of glucocorticoid receptor agonists on Th2 cell function and attenuation by progesterone

“…1 (Fig. S 1 )) [ 42 , 43 ] cultured in IL-2 (4 days) were treated (24 h) with equimolar concentrations of either CS (10 − 9 -10 − 7 M) as in [ 15 , 18 ]. Moreover, these concentrations are within the pharmacological range [ 44 ].…”

Revisiting the Concept of Targeting NFAT to Control T Cell Immunity and Autoimmune Diseases

“…RUNX3 physically interacts with NFAT2 and suppresses IL-4 production ( 64 ). NFAT1 competitively binds to the CRTh2 promoter with GATA3 and negatively regulates CRTh2 expression, which mediates the production of Th2 cytokines such as IL-4, IL-5, and IL-13 ( 65 ). Nfat1 deficiency increased Th2 cytokine levels, enhanced chromatin accessibility, and increased DNA demethylation in the Il4 promoter region, inducing preferential recruitment of JUNB/SATB1 to the Il4 promoter ( 51 , 52 ).…”

Lipid‐Embedded Molecular Dynamics Simulation Model for Exploring the Reverse Prostaglandin D2 Agonism of CT‐133 towards CRTH2 in the Treatment of Type‐2 Inflammation Dependent Diseases

“…This underscores the role that CRTH2 directly plays in the recruitment and release of inflammatory cells and pro‐inflammatory cytokines such as interleukin‐4 (IL‐4), IL‐5 and IL‐13 in response to its native agonist, prostaglandin D2 (PGD2) . CRTH2 activation has therefore been implicated in the pathogenesis of type 2 inflammation diseases involving allergies such as asthma, rhinitis and cigarette smoke (CS) induced inflammation . Recent studies has also established the activities of CRTH2 beyond allergy and asthma to involve pro‐fibrotic and antifibrotic effects in the respiratory tract, abetting renal fibrosis and inflammation through repression of interleukin 4 and 13 as well as inflammation induced cancer .…”

“…Being a G‐protein coupled receptor, the conformational changes that occur upon the binding of these ligands is of interest to understand the mechanism of action of the agonists as well as the antagonists. These insights have facilitated the development of several selective CRTH2 antagonists directed towards the treatment of type 2 inflammation‐induced asthma, alopecia and recently cigarette‐induced acute lung injury, with few advancing to phase 2 and 3 clinical trials. Notable among these selective inhibitors is fevipiprant, which has advanced to phase 3 of clinical trials…”

“…CT‐133 mediated the reduction of infiltrating pro inflammatory cells and macrophages in both in vivo and in vitro studies thereby suggesting a defensive role in acute lung injury . CT‐133 was found to significantly reduce invading inflammatory cells and their effects on the lungs as well as supporting the production of IL‐10 and attenuating the chemotaxis of neutrophils by PGD2 in cigarette smoking‐induced diseases in mice . As such, many PGD2 antagonists target the same binding pocket as PGD2 on CRTH2 in order to negate the biological impacts of PGD2…”