Platelet-derived growth factor-AA-mediated functional angiogenesis in the rat epigastric island flap after genetic modification of fibroblasts is ischemia dependent


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Abstract

Background.The aim of this study was to induce therapeutic angiogenesis in ischemically challenged flap tissue by means of gene transfer.Methods.Isogenic rat fibroblasts were retrovirally transfected to produce platelet-derived growth factor (PDGF)-AA. Stable gene expression was monitored by PDGF-AA enzyme-linked immunosorbent assay. Eighty animals were divided into 2 groups (1 and 2), each with 4 subgroups. The angiogenic target was a 7 × 7-cm epigastric island flap used as a necrosis model. Group 1 received flap treatment 1 week before flap elevation: 107 genetically modified fibroblasts, expressing PDGF-AA (genetically modified fibroblasts) plus 1 mL of Dulbecco's modified Eagle's medium (DMEM) (1A), 107 nonmodified fibroblasts (NMFB) plus 1 mL of DMEM (1B), 1 mL of DMEM (1C), and 1 mL of sodium chloride 0.9% (1D). All substances were injected at evenly distributed spots into the panniculus carnosus of the entire flap. Group 2 had the same flap treatment at the day of flap elevation. All flaps were sutured back. Seven days later, the flaps were harvested and examined both clinically, histologically, and immunohistochemically.Results.In vitro, the GMFB produced up to 117.9 ± 57.2 ng of PDGF-AA/mL medium during a 4-day period, compared with 0.7 ± 0.6 ng of PDGF-AA/mL medium produced by NMFB in the same time period. In vivo production of PDGF-AA in flaps amounted to 1.3 ± 0.7 ng of PDGF-AA/1 μL flap tissue for group 1A and 1.7 ± 1.1 ng of PDGF-AA/1 μL flap tissue for group 2A seven days after cell transplantation. Fibroblasts persisted in all flaps from groups 1A, 1B, 2A, and 2B without major inflammatory reaction. Clinically, group 2A developed significantly less flap necrosis compared with all other groups, including group 1A. Accordingly, only group 2A gave significant histologic and immunhistochemical evidence for enhanced angiogenesis within the flap tissue.Conclusions.After retroviral gene transfer, isogenic rat fibroblasts produce high amounts of PDGF-AA in vitro. In vivo, PDGF-AA can be detected in flaps receiving genetically modified fibroblasts, which suggests survival of the implanted fibroblasts in this model. PDGF-AA produced by GMFB can induce flap angiogenesis only under ischemic conditions in this model. Transplantation of PDGF-AA-overexpressing fibroblasts results in higher flap survival in this model. (Surgery 2002;131:393-400.)

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