Increased Neutrophil H 2 O 2 Production and Enhanced Pulmonary Clearance of Klebsiella pneumoniae in G6PD A- Mice.

The X-linked A ^- variant (rs1050828, Val68Met) in G6PDX accounts for glucose-6-phosphate (G6PD) deficiency in approximately 11% of African American males. This common, hypomorphic variant may impact pulmonary host defense and phagocyte function during pneumonia by altering levels of reactive oxygen species produced by host leukocytes. We used CRISPR-Cas9 technology to generate novel mouse strain with "humanized" G6PD A- variant containing non-synonymous Val68Met single nucleotide polymorphism. Male hemizygous or littermate wild-type (WT) controls were inoculated intratracheally with K. pneumoniae (KP2 serotype, ATCC 43816 strain,10 ³ CFU inoculum). We examined leukocyte recruitment, organ bacterial burden, bone marrow neutrophil and macrophage (BMDM) phagocytic capacity, and hydrogen peroxide (H ₂ O ₂ ) production. Unexpectedly, G6PD-deficient mice showed decreased lung bacterial burden (p=0.05) compared to controls 24-h post-infection. Extrapulmonary dissemination and bacteremia were significantly reduced in G6PD-deficient mice 48-h post-infection. Bronchoalveolar lavage fluid (BALF) IL-10 levels were elevated in G6PD-deficient mice (p=0.03) compared to controls at 24-h but were lower at 48-h (p=0.03). G6PD A- BMDMs show mildly decreased in vitro phagocytosis of pHrodo-labeled KP2 (p=0.03). Baseline, but not stimulated, H ₂ O ₂ production by G6PD A- neutrophils was greater compared to WT neutrophils. G6PD A- variant demonstrate higher basal neutrophil H ₂ O ₂ production and are protected against acute Klebsiella intrapulmonary infection.
Competing Interests: Competing Interests: The authors BEZ, HFP, ZX, LW, ECP, ER, MY, SG, and JSL have no conflicts of interest to report. MTG receives research support from NIH grants R01HL098032, R01HL125886, UH3HL143192, R01 HL168775, the Department of Defense, and Globin Solutions, Inc. He previously received research support from Bayer Corp. MTG is a co-inventor of patents and patent applications directed to the use of recombinant neuroglobin and heme-based molecules as antidotes for CO poisoning, which have been licensed by Globin Solutions, Inc. MTG is a shareholder, advisor, and director in Globin Solutions, Inc. MTG is also co-inventor on patents directed to the use of nitrite salts in cardiovascular diseases, which were previously licensed to United Therapeutics, and now licensed to Globin Solutions and Hope Pharmaceuticals. MTG is an inventor on an unlicensed patent application directed at the use of nitrite for halogen gas poisoning and smoke inhalation. MTG was a principal investigator in a research collaboration with Bayer Pharmaceuticals to evaluate riociguate as a treatment for patients with SCD, which has concluded. MTG is a textbook author and receives royalties from MedMaster Inc., and is a textbook editor and receives royalties from McGraw-Hill. MTG has an active consulting agreement in place with Third Pole Therapeutics. Agreements that concluded more than 12 months ago: He has previously served as a consultant for Forma Therapeutics, Acceleron Pharma, Inc., Sujana Biotech, Epizyme, Inc., Catalyst Biosciences, Inc. Complexa, Novartis, United Therapeutics, Fulcrum, Actelion, Bayer Healthcare, Pfizer, and Modus Therapeutics. MTG also previously served on Bayer HealthCare LLC’s Heart and Vascular Disease Research Advisory Board.