OBJECTIVE: To study the effects of an amelogenin mixture on integrin-dependent adhesion, DNA synthesis and apoptosis of cultured human dermal microvascular endothelial cells and angiogenesis in an organotypic assay. METHOD: Immobilised antibodies against specific integrins (alpha-1, alpha-2, alpha-3, alpha-4, alpha-5, alpha-v, ß1, ß2, ß3, ß4, ß6, alpha-vß3, alpha-vß5 and alpha-5ß1) were used to capture treated human dermal microvascular endothelial cells, which were detected colourimetrically. DNA synthesis of the cells was monitored by 5-bromo-2'- deoxyuridine incorporation and apoptosis by a TdT-mediated dUTP nick-end labelling technique. Tubule formation from aortic arches of 13-d-old chick embryos were followed over 48h. RESULTS: The amelogenin mixture increased microvessel outgrowth by 76% (p < 0.01, n=12) from the aortic explants. Also, amelogenins increased the adhesion (p < 0.01, n = 5) by multiple angiogenesis associated integrin subunits and alpha-vß3, alpha-vß5 and alpha-5ß1 heterodimers on human dermal microvascular endothelial cells at a non-mitogenic concentration (100 µg/ml). Conversely, amelogenins at 1,000 µg/ml decreased microvessel formation possibly due to attenuation of corresponding integrins despite increasing (p < 0.001, n = 8) DNA synthesis. No significant apoptosis was detected in human dermal microvascular endothelial cells cultured on Matrigel with and without amelogenins. CONCLUSION: Increased surface expression of integrins on endothelial cells may contribute to the proangiogenic property of amelogenins.
OBJECTIVE: To study the effects of an amelogenin mixture on integrin-dependent adhesion, DNA synthesis and apoptosis of cultured human dermal microvascular endothelial cells and angiogenesis in an organotypic assay. METHOD: Immobilised antibodies against specific integrins (alpha-1, alpha-2, alpha-3, alpha-4, alpha-5, alpha-v, ß1, ß2, ß3, ß4, ß6, alpha-vß3, alpha-vß5 and alpha-5ß1) were used to capture treated human dermal microvascular endothelial cells, which were detected colourimetrically. DNA synthesis of the cells was monitored by 5-bromo-2'- deoxyuridine incorporation and apoptosis by a TdT-mediated dUTP nick-end labelling technique. Tubule formation from aortic arches of 13-d-old chick embryos were followed over 48h. RESULTS: The amelogenin mixture increased microvessel outgrowth by 76% (p < 0.01, n=12) from the aortic explants. Also, amelogenins increased the adhesion (p < 0.01, n = 5) by multiple angiogenesis associated integrin subunits and alpha-vß3, alpha-vß5 and alpha-5ß1 heterodimers on human dermal microvascular endothelial cells at a non-mitogenic concentration (100 µg/ml). Conversely, amelogenins at 1,000 µg/ml decreased microvessel formation possibly due to attenuation of corresponding integrins despite increasing (p < 0.001, n = 8) DNA synthesis. No significant apoptosis was detected in human dermal microvascular endothelial cells cultured on Matrigel with and without amelogenins. CONCLUSION: Increased surface expression of integrins on endothelial cells may contribute to the proangiogenic property of amelogenins.