Flow cytometric measurement of CFTR-mediated chloride transport in human neutrophils​Alejandra Montanez-Barragan on 21 de January de 2025 at 11:00

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J Leukoc Biol. 2025 Jan 22:qiaf006. doi: 10.1093/jleuko/qiaf006. Online ahead of print.

ABSTRACT

Immune cells express a variety of ion channels and transporters in the plasma membrane and intracellular organelles, responsible of the transference of charged ions across hydrophobic lipid membrane barriers. The correct regulation of ion transport ensures proper immune cell function, activation, proliferation, and cell death. Cystic fibrosis (CF) is a genetic disease in which the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) chloride channel gene is defective, consequently, the CFTR protein is dysfunctional, and the chloride efflux in CF cells is markedly impaired. Cystic fibrosis is characterized by chronic inflammation in the airways, mainly triggered by neutrophilic infiltration and recurring bacterial infections, causing the decline of lung function and eventually respiratory failure. Novel modulator-based therapies have improved lung function in people with Cystic Fibrosis (pwCF) by increasing expression and function of CFTR in the plasma membrane of lung cells, however, the effects of these drugs in the lung recruited inflammatory cells, specifically neutrophils, remains unknown. Given the complex biology of neutrophils and their short lifespan, we aimed to develop a fluorometric method to evaluate CFTR-mediated chloride transport in human neutrophils by using flow cytometry and the intracellular chloride-binding MQAE dye. Our results show that CFTR-mediated chloride transport in human neutrophils or human neutrophil-like cell lines can be consistently evaluated in vitro by this methodology. Additionally, this assay measured increased chloride efflux in neutrophils collected from pwCF under modulator therapy, as compared to healthy donors, indicating this method can evaluate restoration of CFTR-mediated chloride transport in CF neutrophils.

PMID:39837350 | DOI:10.1093/jleuko/qiaf006

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