RESUMEN
CI-1023 (AdGVVEGF121.10) is a replication-deficient adenovirus vector (complete E1a-, partial E1b-, partial E3-) delivering human vascular endothelial growth factor-121 gene. Previous studies from this group have established that CI-1023 can successfully transfer human vascular endothelial growth factor-121 gene resulting in local tissue expression of vascular endothelial growth factor protein. The purpose of this study was to evaluate neovascularization-promoting potency and efficacy of CI-1023 in a wide dose range. In a rat hindlimb ischaemic model, we measured neovascularization-promoting effect of CI-1023 using three end-points: post mortem angiography, immuno-histochemistry and Laser Doppler scanning of tissue blood perfusion. Neovascularization-promoting activity of CI-1023 over the dose range of 4 x 10(6) pu-4 x 10(10) pu was evaluated. Our data demonstrated an obvious dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu. The neovascularizing effect is somewhat plateaued at the levels between 4 x 10(8) pu and 4 x 10(10) pu. We conclude CI-1023 is a potent neovascularization-promoting compound, with a dose-dependent effect between 4 x 10(6) pu-4 x 10(8) pu in the rat hindlimb ischaemic model.
Asunto(s)
Proteínas Angiogénicas/farmacología , Relación Dosis-Respuesta a Droga , Miembro Posterior/efectos de los fármacos , Linfocinas/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Proteínas Angiogénicas/administración & dosificación , Proteínas Angiogénicas/uso terapéutico , Angiografía/métodos , Animales , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos/métodos , Vectores Genéticos/administración & dosificación , Vectores Genéticos/química , Vectores Genéticos/farmacología , Miembro Posterior/irrigación sanguínea , Linfocinas/administración & dosificación , Linfocinas/uso terapéutico , Masculino , Neovascularización Fisiológica/fisiología , Perfusión , Ratas , Ratas Sprague-Dawley , Factores de Tiempo , Factores de Crecimiento Endotelial Vascular/genética , Factores de Crecimiento Endotelial Vascular/metabolismoRESUMEN
We have studied an age-related impairment in angiogenesis and evaluated the effect of overexpressing VEGF in this situation. Polyvinyl alcohol sponges were implanted subcutaneously into aged (24-month), adult (12-month), and young (2-month) rats. Blood vessel ingrowth and proliferative activity in the sponges were assessed by histology with immunostaining for von Willebrand's factor and proliferating cell nuclear antigen (PCNA), respectively. The percentage of total sponge area filled with ingrowing fibrovascular tissue was minimal in aged rats, intermediate in adult rats and highest in young rats. A similar pattern was observed for the total blood vessel numbers in the sponges from old to young animals. The percentage of total sponge endothelial cells (ECs) showing proliferative activity (PCNA positive) was lowest in the aged animals, intermediate in the adult rats and highest in the young rats. To further explore the mechanism of impaired angiogenesis in aged animals, we investigated and found a reduced level of endogenous VEGF protein expression in 12-month-old rats compared to that in 2-month-old rats. VEGF121 gene transfer significantly enhanced blood vessel and fibrovascular tissue ingrowth in adult/aged rats. Adenoviral-VEGF gene transfer also significantly stimulated EC proliferation in aged and adult rats. However, identical treatment failed to further stimulate the already more robust angiogenesis in young animals. The different angiogenic response in adult vs. young rats was not due to differences in gene transfer efficiency, since similar levels of human VEGF121 protein was detected in adult and young rats. Our results indicate that the decreased angiogenic response with aging is associated with reduced EC proliferation and reduced endogenous VEGF production. Adenoviral-VEGF121 gene transfer is effective in augmenting angiogenesis, particularly in older animals.
Asunto(s)
Adenoviridae/genética , Envejecimiento , Técnicas de Transferencia de Gen , Neovascularización Fisiológica , Factor A de Crecimiento Endotelial Vascular/genética , Animales , Northern Blotting , División Celular , Endotelio Vascular/citología , Ensayo de Inmunoadsorción Enzimática , Humanos , Immunoblotting , Inmunohistoquímica , Neovascularización Patológica , Ratas , Factores de Tiempo , Transfección , Factor A de Crecimiento Endotelial Vascular/biosíntesis , Receptor 2 de Factores de Crecimiento Endotelial Vascular/biosíntesisRESUMEN
This study examined whether retarded angiogenesis in a hypertension animal model was associated with impaired VEGF signaling. Furthermore, we sought to determine whether this impairment could be overcome by VEGF addition. Using a rat sponge implantation model, we confirmed impaired angiogenesis in spontaneous hypertensive rats (SHRs). Fourteen days after sponge implantation, the level of angiogenesis in SHRs was approximately half of those in age-matched normotensive Wistar-Kyoto or Sprague-Dawley rats. Significantly, expression of kinase-insert domain-containing receptor (KDR) and membrane type 1 matrix metalloproteinase (MT1-MMP) was reduced in SHRs compared to controls. Immunohistological analysis indicated endothelial proliferation was decreased in SHRs. Gene transfer of human VEGF(121) increased KDR and MT1-MMP expression in SHRs. VEGF(121) also up-regulated endothelial proliferation and angiogenesis. Our results indicate down-regulated KDR and MT1-MMP expression is associated with an impaired angiogenesis in SHRs. VEGF gene transfer is effective in ameliorating the impaired angiogenesis in SHRs.