Dataset Information

Phagosomal chloride dynamics in the alveolar macrophage.

ABSTRACT: Acidification in intracellular organelles is tightly linked to the influx of Cl^- counteracting proton translocation by the electrogenic V-ATPase. We quantified the dynamics of Cl^- transfer accompanying cargo incorporation into single phagosomes in alveolar macrophages (AMs). Phagosomal Cl^- concentration and acidification magnitude were followed in real time with maximal acidification achieved at levels of approximately 200 mM. Live cell confocal microscopy verified that phagosomal Cl^- influx utilized predominantly the Cl^- channel CFTR. Relative levels of elemental chlorine (Cl) in hard X-ray fluorescence microprobe (XFM) analysis within single phagosomes validated the increase in Cl^- content. XFM revealed the complex interplay between elemental K content inside the phagosome and changes in Cl^- during phagosomal particle uptake. Cl^- -dependent changes in phagosomal membrane potential were obtained using second harmonic generation (SHG) microscopy. These studies provide a mechanistic insight for screening studies in drug development targeting pulmonary inflammatory disease.

SUBMITTER: Riazanski V

PROVIDER: S-EPMC8733233 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Phagosomal chloride dynamics in the alveolar macrophage.

Riazanski Vladimir V Mauleon Gerardo G Zimnicka Adriana M AM Chen Si S Nelson Deborah J DJ

iScience 20211216 1

Acidification in intracellular organelles is tightly linked to the influx of Cl- counteracting proton translocation by the electrogenic V-ATPase. We quantified the dynamics of Cl- transfer accompanying cargo incorporation into single phagosomes in alveolar macrophages (AMs). Phagosomal Cl- concentration and acidification magnitude were followed in real time with maximal acidification achieved at levels of approximately 200 mM. Live cell confocal microscopy verifi ...[more]

PMID: 35024579

Similar Datasets

Project description:Pulmonary Alveolar Proteinosis (PAP) patients exhibit an acquired deficiency of biologically active granulocyte-macrophage colony stimulating factor (GM-CSF) attributable to GM-CSF specific autoantibodies. PAP alveolar macrophages are foamy, lipid-filled cells with impaired surfactant clearance and markedly reduced expression of the transcription factor peroxisome proliferator-activated receptor gamma (PPAR?) and the PPAR?-regulated ATP binding cassette (ABC) lipid transporter, ABCG1. An open label proof of concept Phase II clinical trial was conducted in PAP patients using rituximab, a chimeric murine-human monoclonal antibody directed against B lymphocyte specific antigen CD20. Rituximab treatment decreased anti-GM-CSF antibody levels in bronchoalveolar lavage (BAL) fluid, and 7/9 patients completing the trial demonstrated clinical improvement as measured by arterial blood oxygenation.This study sought to determine whether rituximab therapy would restore lipid metabolism in PAP alveolar macrophages.BAL samples were collected from patients pre- and 6-months post-rituximab infusion for evaluation of mRNA and lipid changes.Mean PPAR? and ABCG1 mRNA expression increased 2.8 and 5.3-fold respectively (p???0.05) after treatment. Lysosomal phospholipase A2 (LPLA2) (a key enzyme in surfactant degradation) mRNA expression was severely deficient in PAP patients pre-treatment but increased 2.8-fold post-treatment. In supplemental animal studies, LPLA2 deficiency was verified in GM-CSF KO mice but was not present in macrophage-specific PPAR? KO mice compared to wild-type controls. Oil Red O intensity of PAP alveolar macrophages decreased after treatment, indicating reduced intracellular lipid while extracellular free cholesterol increased in BAL fluid. Furthermore, total protein and Surfactant protein A were significantly decreased in the BAL fluid post therapy.Reduction in GM-CSF autoantibodies by rituximab therapy improves alveolar macrophage lipid metabolism by increasing lipid transport and surfactant catabolism. Mechanisms may involve GM-CSF stimulation of alveolar macrophage ABCG1 and LPLA2 activities by distinct pathways.

Dataset Information

Phagosomal chloride dynamics in the alveolar macrophage.

Publications

Phagosomal chloride dynamics in the alveolar macrophage.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets