The effect of induced sputum and bronchoalveolar lavage fluid from patients with chronic obstructive pulmonary disease on neutrophil migration in vitro

Agnė Babušytė¹, Jolanta Jeroch^{1, 2}, Rimantas Stakauskas³, Kristina Stravinskaitė⁴, Kęstutis Malakauskas^{1, 4}, Raimundas Sakalauskas⁴

¹Laboratory of Pulmonology, Institute for Biomedical Research, Kaunas University of Medicine, Lithuania, ²Department of Laboratory Medicine, Kaunas University of Medicine, Lithuania, ³Institute of Immunology, University of Veterinary Medicine, Hannover, Germany, ⁴Department of Pulmonology and Immunology, Kaunas University of Medicine, Lithuania

Key words: neutrophils; migration; chronic obstructive pulmonary disease; induced sputum; bronchoalveolar lavage fluid.

Summary. Objective. The aim of study was to investigate a chemotactic effect of induced sputum and bronchoalveolar lavage fluid on blood neutrophils in patients with chronic obstructive pulmonary disease (COPD) and healthy individuals.

Material and methods. Forty-three smokers with COPD, 19 ex-smokers with COPD, 13 healthy smokers, and 17 healthy nonsmokers were recruited to the study.

Neutrophils were isolated from peripheral blood of study individuals. For the same experimental conditions, pooled induced sputum and bronchoalveolar lavage fluid of 20 COPD patients were used.

Neutrophil chemotaxis in vitro was performed in cell-transmigration chamber. Substances tested for chemoattraction (interleukin-8, induced sputum, bronchoalveolar lavage fluid directly or in addition to interleukin-8) were added to lower wells. Upper wells were filled with 2.5×10⁶/mL of neutrophil culture and incubated for 2 hours. Migration was analyzed by flow cytometry.

Results. Interleukin-8 (10–100 ng/mL) induced a dose-dependant neutrophil migration in all the groups. Only 100 ng/L of interleukin-8 induced more intensive chemotaxis of neutrophils from COPD smokers as compared to ex-smokers (P<0.05). Such difference between healthy individuals was obtained using 30 ng/mL of interleukin-8 (P<0.05).

Induced sputum/interleukin-8 (10–100 ng/mL), as well as induced sputum directly, induced neutrophil migration (P<0.05). Chemotaxis of neutrophils isolated from COPD patients and healthy nonsmokers did not depend on additional interleukin-8 concentration.

Bronchoalveolar lavage fluid/interleukin-8 (30–100 ng/mL) induced more intensive migration of neutrophils from COPD patients than bronchoalveolar lavage fluid (P<0.05) alone.

Conclusions. Migration of neutrophils isolated from patients with COPD was more intensive compared to healthy individuals. Induced sputum and bronchoalveolar lavage fluid directly and with addition of interleukin-8 stimulated chemotaxis, and it was higher in neutrophils from COPD patients. Migration of neutrophils did not depend on smoking status.

Correspondence to A. Babušytė, Laboratory of Pulmonology, Institute for Biomedical Research, Kaunas University of Medicine, Eivenių 4, 50161 Kaunas, Lithuania. E-mail: agne.babusyte@gmail.com

Received 17 June 2009, accepted 7 May 2010