Chronic high pressure potentiates the antiproliferative effect and abolishes contractile phenotypic changes caused by endothelial cells in cocultured smooth muscle cells.
High in vitro pressures have been reported to alter smooth muscle cell (SMC) and endothelial cell (EC) phenotype, while endothelial cells (ECs) can influence the proliferation, phenotype, and contractile features of smooth muscle cells (SMC) in coculture systems. However, little is known about the in vitro effects of pressure on EC/SMC cocultures. We therefore sought to compare SMC proliferation in independent and EC coculture under ambient and high pressure, and identify changes in the contractile phenotype of SMCs by measuring levels of the L-type Ca(2+) channel a(1) subunit (dihydropyridine-DHP receptor) which is critical for Ca(2+) transients, differentiation and contractility in SMC.


METHODS
Rat aortic SMCs in independent culture (SMC/0) and coculture with ECs (SMC/EC) were maintained in 5% CO(2) under either atmospheric or high pressure (130 mmHg). SMC were counted at 0, 1, 3, and 5 days and compared to initial cell counts of day 0 before the exposure to experimental conditions. DHP receptor levels were quantitated by Western blotting (three similar studies).


RESULTS
ECs suppressed SMC proliferation on day 1 of coculture in both atmospheric and high pressure (20% inhibition vs independent culture, P < or = 0.05). By day 3, cocultured SMC under atmospheric pressure displayed no EC-mediated inhibition, and at day 5, atmospheric cocultured SMCs revealed statistically significant enhanced proliferation as compared with SMCs in independent cultures. However, cocultured SMCs exposed to 130 mmHg pressure displayed sustained sensitivity to EC growth inhibition at both days 3 and 5 of the experiment. Coculture decreased SMC DHP-receptor levels under atmospheric pressure. However, this effect was abolished in cocultures exposed to high pressure.


CONCLUSIONS
High pressure substantially alters the regulatory influence of EC on SMC proliferation and contractile potential. This pressure/coculture model should increase our understanding of cellular interaction in hypertensive vasculopathy.
Vouyouka
Angela G
AG
Departments of Surgery and Cardiology, John D. Dingell VA Medical Center, Detroit, MI 48201-1932, USA. Angela.Vouyouka@med.va.gov
Salib
Shady Selim
SS
Cala
Steven
S
Marsh
James D
JD
Basson
Marc D
MD
D000818:Animals; D001011:Aorta / Q000166:cytology / Q000503:physiopathology; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000502:physiology*; D018929:Cell Culture Techniques; D002455:Cell Division / Q000502:physiology; D002465:Cell Movement / Q000502:physiology; D002908:Chronic Disease; D018920:Coculture Techniques; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D006973:Hypertension / Q000378:metabolism / Q000503:physiopathology*; D008954:Models, Biological; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000502:physiology*; D010641:Phenotype; D011312:Pressure; D051381:Rats; D014664:Vasodilation / Q000502:physiology*
D020746:Calcium Channels, L-Type; D002118:Calcium