1. Chitinase activation in patients with fungus-associated cystic fibrosis lung disease 
Andreas Hector, MD, Sanjay H. Chotirmall, MD, PhD, Gillian M. Lavelle, PhD, Bojana Mirković, PhD, Deirdre Horan, MD, Laura Eichler, MD, Markus Mezger, MD, PhD, Anurag Singh, PhD, Anjai Ralhan, PhD, Sina Berenbrinker, MD, Ines Mack, MD, Regina Ensenauer, MD, Joachim Riethmüller, MD, Ute Graepler-Mainka, MD, Michelle A. Murray, MD, Matthias Griese, MD, N. Gerry McElvaney, MD, Dominik Hartl, MD 
Journal of Allergy and Clinical Immunology 
Volume 138, Issue 4, Pages e4 (October 2016)
DOI: /j.jaci
Copyright © 2016 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2. Fig 1
Chitinase activities in patients with CF. Chitinase activities were quantified in blood (serum; A and B) and BALF (C; n = 15) from patients with CF (serum, n = 105), asthmatic patients (serum, n = 20), patients with Gaucher disease (serum, n = 3), or healthy control subjects (serum, n = 17). Bars represent means with SEMs. Fig 1, B and C, Patients with CF were stratified into patients colonized (intermittent or chronic19) or not with C albicans (blood: Cand+, n = 68 and Cand−, n = 37 [Fig 1, B]; BALF: Cand+, n = 8 and Cand−, n = 7 [Fig 1, C]) or A fumigatus (blood: Asp+, n = 23 and Asp−, n = 82 [Fig 1, B]; BALF: Asp+, n = 6 and Asp−, n = 9 [Fig 1, C]). *P < .05 versus control subjects. n.s., Nonsignificant.
Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci )
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3. Fig 2
Chitinase-protease interactions in airway fluids from patients with CF. A, Effect of airway proteases from patients with CF on the integrity of recombinant chitotriosidase. Upper panel, Time-course incubation of 1 μg of chitotriosidase with 8 μmol/L neutrophil elastase (NE; lanes 2-4), cathepsin L (CatL; lanes 5-7), and cathepsin B (CatB; lanes 8-10) for up to 120 minutes. Chitotriosidase was incubated with water as a control standard (lanes 1 and 11). A representative blot of 3 individual experiments is shown. The 50-kDa chitotriosidase protein is depicted with an arrow. Lower panel: Left panel, Chitotriosidase (1 μg) was incubated with BALF from 3 different adults with CF (lanes 2, 4, and 6; high elastase content), 3 different adults without CF (HC; lanes 3, 5, and 7; low elastase content), and 3 different pediatric patients with CF (lanes 8–10; low elastase content, 9 μL) for 1 hour at 37°C. Chitotriosidase was incubated with water as a control standard (lane 1). Right panel, Chitotriosidase was added to BALF from an adult with CF that had been preincubated for 1 hour at 37°C with water (lane 11) or various protease inhibitors: phenylmethylsulfonyl fluoride (PMSF; NE inhibitor, lane 12), CAA0225 (cathepsin L inhibitor, lane 13), and CA074 (cathepsin B inhibitor, lane 14). A representative blot of 3 individual experiments is shown. The 50-kDa chitotriosidase protein is depicted with an arrow. B, Chitinase and proteases activities in airway fluids from patients with CF. Chitotriosidase and elastase activities were analyzed in BALF from patients with CF (n = 15).
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4. Fig 3
Genetic chitinase deficiency and fungal colonization in patients with CF. DNA samples from 521 patients with CF were genotyped for genetic chitotriosidase deficiency (24-bp duplication, CHIT1, rs ). C albicans status was determined according to the method of Chotirmall et al.19 Bars show means with SEMs. CHIT+/+ (wild-type; n = 330, 63.3%), CHIT+/− (heterozygous; n = 162, 31.1%), and CHIT−/− (homozygous; 24-bp duplication, chitinase deficiency, n = 29, 5.6%).
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5. Fig E1
Protease-chitinase interactions in patients with CF, patients with COPD, and asthmatic patients. Pooled BALF (n = 4 each) from patients with CF, patients with COPD, or asthmatic patients was incubated with 8 μmol/L neutrophil elastase (NE) for 120 minutes. Afterward, chitotriosidase activity was quantified in BALF by using a commercially available substrate assay (Sigma-Aldrich). *P < .05.
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6. Fig E2
Fungus-mediated chitotriosidase release by human neutrophils. Isolated human peripheral blood neutrophils (1 × 106 cells/mL) were incubated for 120 minutes at 37°C and 5% CO2 with medium only (RPMI 1640) or with C albicans yeasts (1 × 105/mL), zymosan depleted (10 μg/mL), or Curdlan (100 μg/mL) or pretreated for 60 minutes with an anti–Dectin-1 antibody (20 μg/mL) and then stimulated with C albicans yeasts (1 × 105/mL) for 120 minutes. Afterward, chitotriosidase activity was quantified in BALF by using a commercially available substrate assay (Sigma-Aldrich). *P < .05, C albicans–stimulated only versus Dectin-1 blocked and C albicans–stimulated; **P < .05 stimulated versus medium.
Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci )
Copyright © 2016 American Academy of Allergy, Asthma & Immunology Terms and Conditions