G protein-coupled receptors (GPCRs) are key signal transducers and the target of about one-third of all FDA-approved drugs. Many structural and pharmacological studies rely on disrupted membrane conditions, such as purified receptors in artificial systems or radioligand binding assays using membrane fragments, even though it had not been systematically studied whether membrane integrity affects GPCR function. To address this, we developed Förster resonance energy transfer (FRET)-based GPCR conformation sensors to directly measure receptor activation in both intact and disrupted membranes. Our results show that while some GPCRs remain unaffected, prostanoid receptor conformation sensors exhibit a strong dependence on membrane integrity: their agonist and antagonist potencies decrease up to 30-fold upon membrane disruption, revealing a crucial role of the membrane integrity in ligand-receptor affinity. Validation with wildtype receptors in functional signaling assays confirmed that these effects reflect genuine receptor characteristics rather than unspecific signals from the sensor design. We ruled out several factors that could explain the loss of affinity but were unable to fully elucidate the mechanism behind this phenomenon. Nevertheless, this effect may introduce bias into structural and pharmacological studies. It is therefore essential to account for membrane integrity and to employ optimized experimental strategies to ensure robust and reliable data interpretation.
© 2025 The Author(s).

In this study, we observe that in human and murine melanomas, T-cell activation abates hematopoietic prostaglandin-D2 synthase (HPGDS) transcription in tumor-associated macrophages (TAM) through TNFα signaling. Mechanistically, HPGDS installs a prostaglandin D2 (PGD2) autocrine loop in TAMs via DP1 and DP2 activation that sustains their protumoral phenotype and promotes paracrine inhibition of CD8+ T cells via a PGD2-DP1 axis. Genetic or pharmacologic HPGDS targeting induces antitumoral features in TAMs and favors CD8+ T-cell recruitment, activation, and cytotoxicity, altogether sensitizing tumors to αPD1. Conversely, HPGDS overexpression in TAMs or systemic TNFα blockade sustains a protumoral environment and αPD1 resistance, preventing the downregulation of HPGDS by T cells. Congruently, patients and mice resistant to αPD1 fail to suppress HPGDS in TAMs, reinforcing the evidence that circumventing HPGDS is necessary for efficient αPD1 treatment. Overall, we disclose a mechanism whereby T-cell activation controls the innate immune system, and we suggest HPGDS/PGD2 targeting to overcome immunotherapy resistance.
In this study, we show a mechanism whereby T-cell activation controls the innate immune system and shapes the tumor microenvironment by reducing PGD2 production in TAMs. We suggest HPGDS inhibition as a promising strategy to treat refractory tumors to current immunotherapies or to overcome acquired resistance to immune checkpoint blockade.
©2025 The Authors; Published by the American Association for Cancer Research.

20-carbon fatty acid-derived eicosanoids are versatile signaling oxylipins in mammals. In particular, a group of eicosanoids termed prostanoids are involved in multiple physiological processes, such as reproduction and immune responses. Although some eicosanoids such as prostaglandin E2 (PGE2) have been detected in some insect species, molecular mechanisms of eicosanoid synthesis and signal transduction in insects have not been thoroughly investigated. Our phylogenetic analysis indicated that, in clear contrast to the presence of numerous receptors for oxylipins and other lipid mediators in humans, the Drosophila genome only possesses a single ortholog of such receptors, which is homologous to human prostanoid receptors. This G protein-coupled receptor, named Prostaglandin Receptor or PGR, is activated by PGE2 and its isomer PGD2 in Drosophila S2 cells. PGR mutant flies die as pharate adults with insufficient tracheal development, which can be rescued by supplying high oxygen. Consistent with this, through a comprehensive mutagenesis approach, we identified a Drosophila PGE synthase whose mutants show similar pharate adult lethality with hypoxia responses. Drosophila thus has a highly simplified eicosanoid signaling pathway as compared to humans, and it may provide an ideal model system for investigating evolutionarily conserved aspects of eicosanoid signaling.
Copyright: © 2025 Fujinaga et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Lung cancer is characterized by a poor prognosis and is a significant comorbidity of chronic obstructive pulmonary disease (COPD). Therefore, effective chemopreventive agents are warranted. We evaluated the effects of the cyclooxygenase-2 (COX-2) inhibitor celecoxib on the prevention of lung-carcinoma development using an intermittent smoking-induced lung-carcinoma mouse model. Additionally, we explored COX-2's role in lipid metabolism.
Male A/J mice were exposed to sham air or mainstream cigarette smoke for 20 weeks. Vehicle or celecoxib was administered via intragastric feeding once daily. Lung tissues were analyzed for tumor nodules and emphysema; the bronchoalveolar lavage fluid was collected for cell counting. COX-2 expression was measured using real-time polymerase chain reaction and western blotting; lipidomic analysis was conducted using liquid chromatography-tandem mass spectrometry. Cell proliferation and colony-forming assays were performed on LA-4 cells to assess the effects of prostaglandins and COX-2 inhibitors.
Intermittent smoking exposure increased lung adenomas, adenocarcinomas, and COX-2 expression. Lung adenomas were characterized by abundant COX-2-positive cells. Celecoxib reduced intermittent smoking-induced inflammation, emphysema, and cell counts in the bronchoalveolar lavage fluid and decreased the incidence of lung adenocarcinomas, whereas the total number of observed lung tumors was unchanged. Celecoxib markedly suppressed single-smoke-induced prostaglandin E2 (PGE2) production in the airway. PGE2 increased LA-4 cell viability via the EP4 receptor and promoted colony formation.
Celecoxib effectively inhibited lung-carcinoma development, inflammation, and emphysema, demonstrating the potential for chemoprevention in smokers and patients with COPD. Further studies on EP4 inhibitors for the prevention of emphysema and lung cancer are warranted.
Copyright © 2025 Sakurai, Chubachi, Miyata, Hamamoto, Naganuma, Shimada, Otake, Nakayama, Irie, Tsutsumi, Kameyama, Hegab, Shimoda, Terai, Yasuda, Kanai, Arita and Fukunaga.

Since the discovery of fatty acid hydroxy fatty acids (FAHFAs), significant progress has been made in understanding their regulation, biochemistry, and physiological activities. Here, we contribute to this understanding by revealing that inflammation induces the production of fatty acid hydroxy stearic acids and fatty acid hydroxyoctadecadienoic acids in white adipose tissue depots and in adipocytes cocultured with macrophages. In lipopolysaccharide (LPS)-induced coculture systems, we confirm that adipose triglyceride lipase is required for inflammation-induced FAHFA generation and demonstrate that inflammation is necessary for producing hydroxy fatty acids. Chemically synthesized fatty acid hydroxyoctadecadienoic acids show anti-inflammatory activities in vivo, but only at supraphysiological concentrations. While endogenous FAHFAs are unlikely to be anti-inflammatory due to their low concentrations, conversion of proinflammatory hydroxy fatty acids into FAHFAs may dampen inflammation. Indeed, we demonstrate that proinflammatory lipids, such as hydroxyeicosatetraenoic acids (HETEs) and leukotriene B4 (LTB4), can be converted by cells in culture to weakly anti-inflammatory FAHFAs.
Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.