Despite intensive research studies, theories have yet to focus on the contribution of hypoxia to patency differences observed clinically between arterial vs. venous grafts. This study investigates the differential hypoxic response of endothelial cell (EC) derived growth factor on modulation of smooth muscle cell (SMC) proliferation and migration under hypoxia and its contribution to graft patency.
Our study shows differential regulation of arterial vs. venous smooth muscle cell proliferation and migration under hypoxia. Initiation of SMC proliferation under hypoxia (<5% O2) occurred only after incubation with hypoxic endothelial cell conditioned media (H-ECM). After investigating several possible growth factors in the H-ECM that may be responsible for SMC proliferation, the greatest difference was observed in vascular endothelial growth factor (VEGF-A) and platelet derived growth factor homodimer B (PDGF-BB) expression. Under hypoxia, two-fold increase in VEGF-A was observed in arterial derived SMC (A-SMC) in comparison to venous derived (V-SMC), which showed no significant change. V-SMC showed higher vascular endothelial growth factor receptor -2 (VEGFR-2) expression under hypoxia, while A-SMC had significantly lower (p<0.05) receptor expression. Incubation with VEGFR-2 neutralizing antibody / platelet derived growth factor receptor (PDGFR) antagonist in V-SMC prior to addition of H-ECM resulted in decreased proliferation. A-SMC proliferation under hypoxia did not decrease with incubation of VEGFR-2 neutralizing antibody but did decrease upon PDGFR antagonist incubation. Proliferation in A-SMC and V-SMC was regulated by an ERK1/2 dependant mechanism with V-SMC showing greater ERK1/2 expression under hypoxia upon addition of hypoxic endothelial cell conditioned media.
Our work was also focused on investigating the impact of hypoxia on smooth muscle cell migration. To summarize our work, we determined that SMC migration was occurring via an autocrine and paracrine mechanism under hypoxia. Migration in V-SMC under hypoxia was regulated via VEGFR-1 since there was an induction in VEGFR-1 expression under hypoxia and migration in V-SMC decreased upon neutralization with a VEGFR-1 antibody. V-SMC migration was also regulated by PDGF-BB since there was a partial reduction in V-SMC migration under hypoxia upon neutralization with PDGF-BB antibody. Migration in A-SMC was induced more by PDGF-BB since neutralization with PDGF-BB antibody decreased A-SMC migration to basal levels.
In conclusion, our studies illustrate that origin of cellular growth factors play an important role in how the pathology of a disease develops. Therefore, combination therapies focusing on the use of anti-VEGFR-1 antagonist to combat migration and anti VEGFR-2 and PDGFR â to combat proliferation, would work best to help alleviate intimal hyperplasia, a result of smooth muscle cell migration and proliferation in venous derived grafts and surgical interventions.
University of Minnesota Ph.D. dissertation. September 2011. Major: Pharmacology. Advisor: Sabita Roy, PhD. 1 computer file (PDF); viii, 126 pages.
Differential modulation of arterial vs venous smooth muscle cell proliferation and migration by hypoxia and hypoxia inducible endothelial cell growth factors..
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