Abstract
NGF, whose expression is increased in asthma, may participate into airway inflammation and hyperresponsiveness. We have here studied whether NGF may have a role in human airway smooth muscle cell (HASMC) proliferation.
Proliferation of HASMC was assessed by the XTT and BrdU techniques in the absence or presence of NGF (0.076-22pM). Involvement of the TrkA receptor was assessed by use of its antagonist ALE-540 (300µM), and of a Trk kinase inhibitor K252a (100nM). TrkA phosphorylation was studied by immunoprecipitation and Western blotting. Involvement of the MAP kinase pathway was determined by use of PD98059 (MEK inhibitor: 20µM), SB203580 (p38 inhibitor: 5µM), SP600125 (JNK inhibitor: 1.1µM), and of PI3K inhibitors, wortmannin (100nM) and LY294002 (15µM).
NGF induced proliferation of HASMC (12% at 2.2pM, p<0.01), which was blocked by pretreatment with both ALE-540 (78 ± 11% inhibition) and K252a (97 ± 9%), suggesting involvement of the TrkA receptor. TrkA phosphorylation was revealed at 2.2pM NGF, with maximal activation at 5-15 min. This was totally inhibited by ALE-540 and K252a. PD98059 and SB203580 blocked NGF-induced proliferation (84 ± 16% and 95 ± 3%, respectively). SP600125, wortmannin and LY294002 had no effect.
In conclusion, NGF in the picomolar range induces proliferation of HASMC through activation of the TrkA receptor and the ERK and p38 pathways. NGF might therefore participate into the airway remodelling in asthma, by increasing proliferation of the airway smooth muscle.