Insulin and local growth factor PDGF induce intimal hyperplasia in bypass graft culture models of saphenous vein and internal mammary artery.

OBJECTIVE: In arteriosclerosis and bypass graft stenosis, intimal proliferation is controlled by local and systemic growth factors, such as platelet derived growth factor (PDGF) or insulin. Intimal hyperplasia can be produced in organ culture models. Our aim was to compare neointima formation in two organ culture models of internal mammary artery (IMA) and saphenous vein (SV), with special reference to the influence of systemic and local growth stimuli. METHODS: Rings of freshly isolated human SV and IMA were cultured over a 3-, 6- or 8-day period. They were distributed into five groups of incubation protocols: incubation with 10% serum; insulin 50 ng/ml and 100 ng/ml; PDGF-BB 5 ng/ml and 10 ng/ml. Frozen sections of cultured rings and pre-culture segments were subjected to elastic stain and immunohistochemistry. Antibodies directed against beta-actin and smooth muscle alpha-actin were used to characterize smooth muscle cell phenotype and against proliferating cell nuclear antigen (PCNA) to demonstrate proliferating cells. RESULTS: Growth factor incubation caused massive intimal hyperplasia with increased elastic fibers in SV and intimal smooth muscle cell as well as matrix accumulation in IMA. Intimal thickening, PCNA and beta-actin expression reached their maximum on day 6 of culture. In both culture models, serum, insulin and PDGF caused increasing intimal thickening, with more pronounced effects in SV. CONCLUSIONS: These organ culture models demonstrate the effects of insulin and PDGF on intimal hyperplasia in IMA and SV representing models for arteriosclerosis and bypass graft stenosis and stressing the role of insulin and growth factors for neointima development.

ETA receptor antagonists inhibit intimal smooth muscle cell proliferation in human vessels.

We have determined the ability of the endothelin (ET)(A) receptor antagonist, PD156707 (CI 1020), to inhibit intimal proliferation in human saphenous veins maintained in organ culture. After 28 days in culture, veins exposed to 1 microM PD156707 exhibited a significant reduction in intima to intima-plus-media ratio (I:I+M ratio) (0.14+/-0.02, n=15) and an increase in lumen area (3.1+/-0.8 mm(2)) compared with veins cultured without the antagonist (I:I+M, 0.29+/-0.02; lumen area, 2.5+/-0.7 mm(2); n=23) but were not significantly different from pre-cultured controls (I:I+M, 0.15+/-0.02; lumen area, 4.4+/-1.2 mm(2); n=17) (Dunn's test for non-parametric multiple comparisons: alpha<0.05). In organ bath experiments, ET-1 and 5-hydroxytryptamine constricted pre-cultured control vessels with pD(2) values (where pD(2) is defined as the negative logarithm of the molar EC(50) value of an agonist) of 8.9+/-0.4 and 7.0+/-0.4 (n=3) and E(max) (efficacy) values of 86+/-3% and 71+/-13% (compared with constriction induced by KCl) respectively. There was no difference in the responsiveness of veins cultured for 14 days to either agonist, indicating that the vessels maintained in organ culture remain viable. Crucially, vein segments cultured with 1 microM PD156707 (a concentration that antagonized ET-1 responses in pre-cultured control vessels) contracted to ET-1 with a potency comparable to that obtained in vessels cultured in the absence of the antagonist (pD(2)=8.9+/-0.4 and 8.0+/-0.6 respectively, n=3) confirming that PD156707 was not toxic to the tissue at the concentration used. In conclusion we have shown that the ET(A)-selective antagonist, PD156707, completely blocked intimal hyperplasia in human saphenous veins in organ culture, suggesting that ET(A) antagonists may be beneficial in preventing or delaying saphenous vein graft disease in patients receiving bypass grafts for coronary artery disease.