Received on February 21, 2007
Accepted on July 23, 2007

Proteomics and Protein Markers

Novel Neutrophil-Derived Proteins in Bronchoalveolar Lavage Fluid Indicate an Exaggerated Inflammatory Response in Pediatric Cystic Fibrosis Patients

¹ Menzies Research Institute, University of Tasmania, Hobart, Australia
² Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC
³ Murdoch Children's Research Institute, Royal Children's Hospital, Melbourne, Australia, and Department of Paediatrics, University of Melbourne, Melbourne, Australia
⁴ Department of Paediatrics and Child Health, Wellington School of Medicine and Health Sciences, University of Otago, Wellington, New Zealand
⁵ Institute for Molecular Bioscience, University of Queensland, Brisbane, Australia
⁶ Department of Paediatrics, Monash University, Melbourne, Australia
⁷ Murdoch Children's Research Institute, Royal Children's Hospital, Melbourne, Australia
⁸ Children's Hospital, Sydney, Australia
⁹ Starship Hospital, Auckland, New Zealand
¹⁰ Royal Children's Hospital, Brisbane, Australia
¹¹ University of Victoria-Genome BC Proteomics Centre, Victoria, British Columbia, Canada
¹² Royal Children's Hospital, Brisbane, Australia, and School of Medicine, University of Queensland, Brisbane, Australia

^* To whom correspondence should be addressed. E-mail: brendan.mcmorran{at}utas.edu.au.

Background: Airway inflammation in cystic fibrosis (CF) is exaggerated and characterized by neutrophil-mediated tissue destruction, but its genesis and mechanisms remain poorly understood. To further define the pulmonary inflammatory response, we conducted a proteome-based screen of bronchoalveolar lavage fluid (BALF) collected from young children with and without CF experiencing endobronchial infection.

Methods: We collected BALF samples from 45 children younger than 5 years and grouped them according to the presence of respiratory pathogens: 1 x 10⁵ colony-forming units (CFU)/mL BALF (18 and 12 samples with and without CF, respectively) and <1 x 10⁵ CFU/mL (23 and 15 samples). BALF proteins were analyzed with SELDI-TOF mass spectrometry (MS) and H4 ProteinChips^®. Proteins were identified and characterized using trypsin digestion, tandem MS, Fourier transform ion cyclotron resonance MS, immunoblotting, and ELISA.

Results: The SELDI-TOF MS BALF profiles contained 53 unique, reliably detected proteins. Peak intensities of 24 proteins differed significantly between the CF and non-CF samples. They included the neutrophil proteins, -defensin 1 and 2, S100A8, S100A9, and S100A12, as well as novel forms of S100A8 and S100A12 with equivalent C-terminal deletions. Peak intensities of these neutrophil proteins and immunoreactive concentrations of selected examples were significantly higher in CF than non-CF samples.

Conclusions: Small-neutrophil–derived BALF proteins, including novel C-terminal truncated forms of S100A proteins, are easily detected with SELDI-TOF MS. Concentrations of these molecules are abnormally high in early CF lung disease. The data provide new insights into CF lung disease and identify novel proteins strongly associated with CF airway inflammation.