Engineering innate immunity: CRISPR-Cas9 MPO knockout in hematopoietic stem cells abolishes chlorination and PMA-induced NET formation while preserving non-oxidative effector functions
Abstract
IntroductionMyeloperoxidase (MPO) is a key neutrophil heme peroxidase that contributes to antimicrobial oxidative killing. Emerging evidence implicates MPO in diverse inflammatory disorders, including atherosclerosis, autoimmune diseases, and neurodegenerative diseases. Since inherited MPO-deficiency is typically compatible with preserved host defense, inhibition or genetic deletion of MPO represents a promising strategy to mitigate MPO-driven tissue injury.MethodsHuman HSPCs were edited using CRISPR-Cas9 to generate MPO-knockout (MPO-KO) neutrophils, followed by in vitro differentiation. We systematically evaluated canonical neutrophil effector functions to determine which of them require MPO. Additionally, RAG2-KO was generated and used as a control for knockout procedure.ResultsMPO-KO did not significantly impair major neutrophil effector functions, including neutrophil elastase activity, fMLP-induced NE and MMP9 degranulation, phagocytosis of E. coli and S. aureus bioparticles, and superoxide production. In contrast, MPO-dependent chlorination was abolished and PMA-induced neutrophil extracellular trap (NET) release was profoundly reduced in MPO-KO neutrophils, while RAG2-KO control cells retained all of these functions.ConclusionThis strategy may offer a durable means to attenuate MPO-driven inflammatory damage, warranting further preclinical evaluation in relevant disease models.
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BibTeX
@article{baron2026engineering,
title = {{Engineering innate immunity: CRISPR-Cas9 MPO knockout in hematopoietic stem cells abolishes chlorination and PMA-induced NET formation while preserving non-oxidative effector functions}},
author = {Szilvia Baron and Aurélien B. L. Colamartino and Jenny Shapiro and Yeshai Schary and Zulema Romero Garcia and Donald B. Kohn and Anat Kahana and Ronit Elhasid and Rachel Shukrun},
journal = {Frontiers in Immunology},
year = {2026},
month = nov,
doi = {10.3389/fimmu.2026.1970785},
url = {https://www.frontiersin.org/articles/10.3389/fimmu.2026.1970785},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Engineering innate immunity: CRISPR-Cas9 MPO knockout in hematopoietic stem cells abolishes chlorination and PMA-induced NET formation while preserving non-oxidative effector functions
AU - Szilvia Baron
AU - Aurélien B. L. Colamartino
AU - Jenny Shapiro
AU - Yeshai Schary
AU - Zulema Romero Garcia
AU - Donald B. Kohn
AU - Anat Kahana
AU - Ronit Elhasid
AU - Rachel Shukrun
PY - 2026
DA - 2026/11/04
JO - Frontiers in Immunology
DO - 10.3389/fimmu.2026.1970785
UR - https://www.frontiersin.org/articles/10.3389/fimmu.2026.1970785
AB - IntroductionMyeloperoxidase (MPO) is a key neutrophil heme peroxidase that contributes to antimicrobial oxidative killing. Emerging evidence implicates MPO in diverse inflammatory disorders, including atherosclerosis, autoimmune diseases, and neurodegenerative diseases. Since inherited MPO-deficiency is typically compatible with preserved host defense, inhibition or genetic deletion of MPO represents a promising strategy to mitigate MPO-driven tissue injury.MethodsHuman HSPCs were edited using CRISPR-Cas9 to generate MPO-knockout (MPO-KO) neutrophils, followed by in vitro differentiation. We systematically evaluated canonical neutrophil effector functions to determine which of them require MPO. Additionally, RAG2-KO was generated and used as a control for knockout procedure.ResultsMPO-KO did not significantly impair major neutrophil effector functions, including neutrophil elastase activity, fMLP-induced NE and MMP9 degranulation, phagocytosis of E. coli and S. aureus bioparticles, and superoxide production. In contrast, MPO-dependent chlorination was abolished and PMA-induced neutrophil extracellular trap (NET) release was profoundly reduced in MPO-KO neutrophils, while RAG2-KO control cells retained all of these functions.ConclusionThis strategy may offer a durable means to attenuate MPO-driven inflammatory damage, warranting further preclinical evaluation in relevant disease models.
SN - 1664-3224
ER -