Src and focal adhesion kinase mediate mechanical strain-induced proliferation and ERK1/2 phosphorylation in human H441 pulmonary epithelial cells.
Pulmonary epithelial cells are exposed to repetitive deformation during physiological breathing and mechanical ventilation. Such deformation may influence pulmonary growth, development, and barotrauma. Although deformation stimulates proliferation and activates extracellular signal-regulated kinases (ERK1/2) in human pulmonary epithelial H441 cells, the upstream mechanosensors that induce ERK activation are poorly understood. We investigated whether c-Src or focal adhesion kinase (FAK) mediates cyclic mechanical strain-induced ERK1/2 activation and proliferation in human pulmonary epithelial (NCI-H441) cells. The H441 and A549 cells were grown on collagen I-precoated membranes and were subjected to an average 10% cyclic mechanical strain at 20 cycles/min. Cyclic strain activated Src within 2 min by increasing phosphorylation at Tyr(418), followed by rapid phosphorylation of FAK at Tyr(397) and Tyr(576) and ERK1/2 at Thr(202)/Tyr(204) (n = 5, P < 0.05). Twenty-four (A549 cells) and 24-72 h (H441 cells) of cyclic mechanical strain increased cell numbers compared with static culture. Twenty-four hours of cyclic strain also increased H441 FAK, Src, and ERK phosphorylation without affecting total FAK, Src, or ERK protein. The mitogenic effect was blocked by Src (10 micromol/l PP2 or short interfering RNA targeted to Src) or MEK (50 micromol/l PD-98059) inhibition. PP2 also blocked strain-induced phosphorylation of FAK-Tyr(576) and ERK-Thr(202)/Tyr(204) but not FAK-Tyr(397). Reducing FAK by FAK-targeted short interfering RNA blocked mechanical strain-induced mitogenicity and significantly attenuated strain-induced ERK activation but not strain-induced Src phosphorylation. Together, these results suggest that repetitive mechanical deformation induced by ventilation supports pulmonary epithelial proliferation by a pathway involving Src, FAK, and then ERK signaling.
Chaturvedi
Lakshmi S
LS
John D. Dingell Veterans Affairs Medical Center, 4646 John R. St., Detroit, MI 48201, USA.
Marsh
Harold M
HM
Basson
Marc D
MD
D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005419:Flavonoids / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000187:drug effects; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D013314:Stress, Mechanical; D013997:Time Factors; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
C412373:AG 1879; D005419:Flavonoids; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D034741:RNA, Small Interfering; D014443:Tyrosine; D051417:Focal Adhesion Kinase 1; C496050:PTK2 protein, human; D019061:src-Family Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020929:Mitogen-Activated Protein Kinase Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one