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Open Access Research

Rituximab therapy in pulmonary alveolar proteinosis improves alveolar macrophage lipid homeostasis

Anagha Malur1, Mani S Kavuru13, Irene Marshall1, Barbara P Barna1, Isham Huizar1, Reema Karnekar1 and Mary Jane Thomassen12*

Author Affiliations

1 Program in Lung Cell Biology and Translational Research, Division of Pulmonary, Critical Care and Sleep Medicine, East Carolina University, Greenville, NC, USA

2 Division of Pulmonary, Critical Care and Sleep Medicine, East Carolina University, Brody School of Medicine, 3E-149 Brody Medical Sciences Building, Greenville, NC, 27834, USA

3 Current address: Division of Pulmonary & Critical Care Medicine, Thomas Jefferson University & Hospital, 834 Walnut St., Suite 650, Philadelphia, PA, 19107, USA

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Respiratory Research 2012, 13:46  doi:10.1186/1465-9921-13-46

Published: 14 June 2012

Abstract

Rationale

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 (PPARγ) and the PPARγ-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.

Objectives

This study sought to determine whether rituximab therapy would restore lipid metabolism in PAP alveolar macrophages.

Methods

BAL samples were collected from patients pre- and 6-months post-rituximab infusion for evaluation of mRNA and lipid changes.

Results

Mean PPARγ 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 PPARγ 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.

Conclusions

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.

Keywords:
PAP; Rituximab; Alveolar macrophages; Surfactant; PPARγ; ABCG1; LPLA2