Endothelial c-IAP2 Loss Amplifies P2X7 Receptor-Driven Inflammation and Worsens Infection-Associated Pulmonary Hypertension

Villarreal, E. S.; Marinho, Y.; Loya, O.; Aboagye, S. Y.; Williams, D. L.; Sun, J.; Erzurum, S.; de Jesus Perez, V.; Oliveira, S. D.

doi:10.1101/2025.05.22.655387

bioRxiv · 10.1101/2025.05.22.655387

Endothelial c-IAP2 Loss Amplifies P2X7 Receptor-Driven Inflammation and Worsens Infection-Associated Pulmonary Hypertension

Villarreal, E. S.·Marinho, Y.·Loya, O.·Aboagye, S. Y.·Williams, D. L.·Sun, J.·Erzurum, S.·de Jesus Perez, V.·Oliveira, S. D.

View original source ↗Export metadata ↓

Abstract

Schistosomiasis-associated Pulmonary Hypertension (Sch-PH) is the most common form of group I PH worldwide. Recently, data revealed that the preclinical animal model of Sch-PH exhibited gut and lung microbiome dysbiosis linked to significant endothelial dysfunction and microvascular apoptosis, but the role of pro/anti-apoptosis sensors, such as the inhibitor of apoptosis protein 2 (c-IAP2) and purinergic receptor P2X7 (P2X7R), remained unclear. Using a novel Cdh5cre-ERT2;cIAP1-/-;cIAP2fl/fl animal model, this study investigated the contribution of endothelial c-IAP2 in this process, revealing P2X7R overexpression as a putative target in the onset of Sch-PH. Pharmacologically, inhibition of P2X7R function confirmed its role in promoting lung endothelial death and disease progression. Moreover, data suggest that microbiome-associated metabolic alterations in Sch-PH seem linked to microvascular endothelial apoptosis driven by ATP/P2X7R overactivation and suppressed c-IAP2 expression. Indeed, genetic ablation of endothelial c-IAP2 expression was sufficient to induce PH-like features in mice, with echocardiography indicating a higher pulmonary acceleration time (PAT), PAT/pulmonary ejection time (PET), and right ventricular free wall thickness after IP/IV-Egg challenge compared to controls. These findings suggest a significant contribution of lung endothelial P2X7R activation and c-IAP2 suppression to Sch-PH pathology, highlighting them as promising novel therapeutic targets for this life-threatening illness.

Explore related subjects

pharmacology and toxicology Preprint

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

Villarreal, E. S., Marinho, Y., Loya, O., Aboagye, S. Y., Williams, D. L., Sun, J., Erzurum, S., de Jesus Perez, V., Oliveira, S. D.. 2025-05-27. Endothelial c-IAP2 Loss Amplifies P2X7 Receptor-Driven Inflammation and Worsens Infection-Associated Pulmonary Hypertension. https://doi.org/10.1101/2025.05.22.655387

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

A metastable GαS/β-arrestin complex sustains endosomal PTH1R signaling

G protein-coupled receptors (GPCRs) initiate cellular signaling by activating heterotrimeric G proteins at the plasma membrane and are silenced when beta-arrestin binds the phosphorylated receptor, preventing G protein binding. However, many receptors continue to generate cAMP after internalization, even while barr is bound, a paradox that lacked a molecular mechanism in live cells. A direct barrestin1-Gas interface in the PTH1R-barr1-Gas complex has recently been defined structurally, but whether the complex operates in endosomes, and how it behaves there, has not been established. Using intact cells, we examine the assembly at three spatial resolutions. Confocal imaging with a nanobody selective for nucleotide-free Gas places activated Gas, barr, and agonist-occupied PTH1R on the same endosomes. Single-molecule localization microscopy, with illumination modes that separate the plasma membrane from the endosomal membrane in single cells, shows that the agonist recruits Gas and barr at both membranes. FRET, however, reveals a specific interaction only on endosomes, where a Gas inhibitor restricted to that compartment abolishes the sustained cAMP phase without affecting receptor internalization. Weighted-ensemble-enhanced molecular simulations show that the complex is metastable, releasing Gas through a large rigid-body rotation away from barr while barr itself tightens its interactions with the receptor. These findings provide a molecular basis for endosomal Gs/cAMP signaling and open a new avenue for drug development targeting this pathway.

pharmacology and toxicology↗

bioRxiv2026

Advancing AOPs for genotoxicity through quantifying KERs linking DNA alkylation to increase in mutations and chromosome aberrations

Quantitative Adverse Outcome Pathways (qAOPs) help translate mechanistic knowledge into quantitative toxicity predictions; however, their development is constrained by the limited availability and heterogeneity of quantitative data. Here, we investigated the feasibility of developing a qAOP for genotoxicity by modeling quantitative relationships linking DNA alkylation to increased mutations and structural chromosome aberrations through intermediate biological key events (KEs). We compiled and harmonized published concentration-response data from human lymphoblastoid TK6 cells exposed to ethyl methanesulfonate, methyl methanesulfonate, or N-ethyl-N-nitrosourea, covering eight genotoxicity assays representing five KEs with well-established dependencies. We evaluated sources of variability, compared points of departure (PoDs), and quantified relationships between successive events using response-response modeling. Substantial between-study variability could to a limited extent be explained through differences in experimental design while batch effects for individual studies and researchers were substantial. PoD estimates that were calculated from combined results of independent studies with similar experimental design, were often uncertain due to insufficient concentration-response characterization or conflicting results between studies. Nevertheless, response-response modeling suggested a threshold effect for certain key-event relationships (KERs). Specifically, DNA adducts seemed to be triggered at lower concentrations of alkylating agents compared to the subsequent KEs of DNA strand breaks and chromosome aberrations. Furthermore, these fitted response-response models can be used for prediction of downstream KEs from earlier ones. The general approach we developed for BMC analysis followed by response-response modeling is applicable to other AOPs as well. These findings highlight both the feasibility and limitations of using existing data for qAOP development.

pharmacology and toxicology↗

bioRxiv2026

Denaturation-sensitive BRET probes enable intracellular thermal shift measurements across diverse protein targets and biomolecular complexes

Thermal shift assays have revolutionized drug discovery, but can be limited in sensitivity and scalability. Combining the unique attributes of differential scanning fluorimetry (DSF) and cellular thermal shift (CETSA), we report the methodological framework for the bioluminescence resonance energy transfer shift assay (BRETSA). BRETSA utilizes denaturation sensitive BRET probes, in combination with luciferase-tagged target proteins, to track and longitudinally record thermal shift in living cells. Denaturation profiles were successfully measured for over 500 globular proteins, spanning 24 target families in diverse cellular compartments. Among these, target engagement was confirmed for over 200 proteins; including validated drug targets as well as understudied members of the kinase subfamily. Leveraging orthosteric, allosteric, and covalent ligands in potency studies, this method offers improved sensitivity and scalability compared to other thermal shift methods. We extend the utility of BRETSA to interrogate target engagement at biomolecular complexes and query cooperative binding for molecular glues and emerging tricomplex drug modalities.

pharmacology and toxicology↗