https://www.jacionline.org/article/S0091-6749(24)00520-7/abstract
The Journal of Allergy and Clinical Immunology, Published: May 19, 2024. DOI: https://doi.org/10.1016/j.jaci.2024.05.009
James Krempski, PhD, Amnah Yamani, PhD, Lakshmi Narasimha Rao Thota, PhD, Sahiti Marella, B Sc, Varsha Ganesan, B Sc, Ankit Sharma, PhD, Atsunori Kaneshige, Ph.D, Longchuan Bai, Ph.D, Haibin Zhou, PhD, Paul S. Foster, PhD, Shaomeng Wang, PhD, Andrea T. Obi, MD, Simon P. Hogan, PhD
Abstract
Background
Mast cell (MC)-derived mediators induce vasodilatation and fluid extravasation, leading to cardiovascular failure in severe anaphylaxis. We have previously revealed a synergistic interaction between the cytokine IL-4 and the MC-derived mediator histamine in modulating vascular endothelial (VE) dysfunction and severe anaphylaxis. The mechanism by which IL-4 exacerbates histamine-induced VE dysfunction and severe anaphylaxis are unknown.
Objective
To identify the IL-4-induced molecular processes regulating the amplification of histamine-induced VE barrier dysfunction and the severity of IgE-mediated anaphylaxic reactions.
Methods
RNAseq, Western blot, Ca2+ imaging and barrier functional analyses on the vascular endothelial cell line (EA.hy926). Pharmacologic degraders (selective PROTAC (proteolysis-targeting chimera) and genetic (lentiviral shRNA) inhibitors were used to determine the roles of STAT3 and STAT6 in conjunction with in vivo model systems of histamine-induced hypovolemic shock.
Results
IL-4 enhancement of histamine-induced VE barrier dysfunction was associated with increased VE-Cadherin degradation, intracellular calcium flux, phospho-Src levels and required transcription and de novo protein synthesis. RNAseq analyses of IL-4 stimulated VE cells identified dysregulation of genes involved in cell proliferation, cell development, and cell growth and transcription factor motif analyses revealed a significant enrichment of differential expressed genes (DEGs) with putative STAT3 and STAT6 motif. IL-4 stimulation in EA.hy926 cells induced both STAT3Y705 and STAT3S727 phosphorylation. Genetic and pharmacologic ablation of VE STAT3 activity revealed a role for STAT3 in basal VE barrier function, however IL-4 enhancement and histamine-induced VE barrier dysfunction was predominantly STAT3-independent. In contrast, IL-4 enhancement and histamine-induced VE barrier dysfunction was STAT6-dependent. Consistent with this finding, pharmacologic knockdown of STAT6 abrogated IL-4-mediated amplification of histamine-induced hypovolemia.
Conclusions
These studies unveil a novel role of the IL-4/ STAT6 signaling axis in the priming of VE cells predisposing to exacerbation of histamine-induced anaphylaxis.
Key words: anaphylaxis vascular endothelium histamine interleukin-4 transcription
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