A role for mitogen kinase kinase 3 in pulmonary inflammation validated from a proteomic approach.
Proteomics is a powerful tool to ascertain which proteins are differentially expressed in the context of disease. We have used this approach on inflammatory cells obtained from patients with asthma to ascertain whether novel drugs targets could be illuminated and to investigate the role of any such target in a range of in vitro and in vivo models of inflammation. A proteomic study was undertaken using peripheral blood mononuclear cells from mild asthmatic subjects compared with healthy subjects. The analysis revealed an increased expression of the intracellular kinase, mitogen activated protein kinase (MKK3), and the function of this protein was investigated further in preclinical models of inflammation using MKK3 knockout mice. We describe a 3.65 fold increase in the expression of MKK3 in CD8(+) T lymphocytes obtained from subjects with asthma compared with healthy subjects using a proteomic approach which we have confirmed in CD8(+), but not in CD4(+) T lymphocytes or human bronchial epithelial cells from asthmatic patients using a Western blot technique. In wild type mice, bacterial lipopolysaccharide (LPS) caused a significant increase in MKK3 expression and significantly reduced airway neutrophilia in MKK3(-/-) mice (median, 25, 75% percentile; wild/LPS; 5.3 (0.7-9.9) × 10(5) cells/mL vs MKK3(-/-)/LPS; 0 (0-1.9) × 10(5) cells/mL, P < 0.05). In contrast, eosinophilia in sensitized wild type mice challenged with allergen (0.5 (0.16-0.65) × 10(5) cells/mL) was significantly increased in MKK3(-/-) mice (2.2 (0.9-3.5) × 10(5) cells/mL, P < 0.05). Our results suggest that asthma is associated with MKK3 over-expression in CD8(+) cells. We have also demonstrated that MKK3 may be critical for airway neutrophilia, but not eosinophilia, suggesting that this may be a target worthy of further consideration in the context of diseases associated with neutrophil activation such as severe asthma and COPD.
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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Holand
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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Riffo-Vasquez
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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Spina
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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O'Connor
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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Woisin
Institute of Pharmaceutical Science, King's College London, 3rd Floor, Franklin Wilkins Building, Waterloo Campus, London SE1 9NH, UK.
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Sand
Cardiovascular Research Division, The Rayne Institute, King's College London, St Thomas' Hospital Campus, Lambeth Palace Rd, London SE1 7EH, UK.
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Marber
Bayer HealthCare AG, Friedrich-Ebert-Str. 475, Wuppertal 42117, Germany.
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Bacon
Formerly Oxford Glycosciences, 208 Bath Road, Slough, Berkshire SL1 3WE, UK.
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Rohlff
Sackler Institute of Pulmonary Pharmacology, Institute of Pharmaceutical Science, King's College London, 5th Floor, Franklin-Wilkins Building, Waterloo Campus, London SE1 9NH, UK. Electronic address: clive.page@kcl.ac.uk.
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D000328:Adult; D000818:Animals; D001249:Asthma / Q000235:genetics* / Q000503:physiopathology; D015153:Blotting, Western; D018414:CD8-Positive T-Lymphocytes / Q000378:metabolism; D016022:Case-Control Studies; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D048371:MAP Kinase Kinase 3 / Q000235:genetics*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009504:Neutrophils / Q000378:metabolism*; D011014:Pneumonia / Q000235:genetics / Q000503:physiopathology; D040901:Proteomics / Q000379:methods*; D055815:Young Adult
D048371:MAP Kinase Kinase 3; C482530:MAP2K3 protein, human; C482531:Map2k3 protein, mouse
Asthma; Inflammation; MKK3; Neutrophils; Proteomics