Abstract
Cigarette smoke extract (CSE) decreases the expression of the CFTR. Furthermore, CSE has profound effects on epithelial cells but different results are reported with cell lines and primary cells. This study aimed to compare the effects of CSE on spontaneous and induced IL-8 release from primary nasal epithelial cells isolated from CF patients possessing two different classes of CFTR mutation (F508del homozygous; R1 17H heterozygote) and healthy volunteers. Nasal epithelial cells were obtained via nasal brushings of the inferior turbinate of CF patients; F508del homozygote (n=5), R1 17H heterozygote (n=7) and healthy volunteers (n=6). The cells were expanded in culture and used at passage 3. Monolayer cultures were pre-exposed for 4h in the presence or absence of 5% CSE. The cells were then stimulated with LPS from P. aeruginosa (LPS-PA 50 or 100 ?g/ml); Cytomix 1 (TNF-? 10 ng/ml, IL-1? 5 ng/ml, LPS-PA 5 ?g/ml) or Cytomix 2 (TNF-? 10 ng/ml, IL-1? 10 ng/ml, IFN-? 1000 U/ml) for 24 h. Culture supernatants were harvested and IL-8 release measured using ELISA (R & D systems). LPS (100 ?g/ml) induced IL-8 release in control cells was increased by CSE (1188?292 vs. 1774?501.6, p< 0.05). In cells from F508del homozygous patients CSE increased cytomix 2 and LPS 50 ?g/ml induced IL-8 secretion. In the R1 17H heterozygotes only the spontaneous IL-8 secretion was significantly increased by CSE exposure. Comparing the responses from the various genetic backgrounds of the subjects; the only significant differences between the groups were with cytomix 2 where the releases followed R1 17H< F508del< control (8157?1746, vs 16031?3729 vs 21296?2576). R1 17H cells were also significantly less responsive to cytomix 2 compared to F508del cells in the presence of CSE.In contrast to our data using cell lines, where CSE consistently decreased IL-8 secretion from the CF cell line (CFBE41o-), preincubation of primary cells with CSE only caused sporadic increases in IL-8 release. Cytomix 2 induced significantly different levels of IL-8 secretion depending on the genetic background of the cells. As yet, we have no explanation for these data but further studies will investigate the action of IFN-? alone on primary cells. Williams MTS et al. Ped Pulmon 2008;287:246