Locally administered vascular endothelial growth factor cDNA increases survival of ischemic experimental skin flaps.

Microvascular surgery has emerged as an attractive area for recent advances in the field of gene therapy. The present study investigated the survival of ischemic, experimental skin flaps after treatment with the gene encoding vascular endothelial growth factor (VEGF). In 30 Sprague-Dawley rats, anterior abdominal skin flaps supplied by the epigastric artery and vein were created. Ten animals were treated with a mixture of liposomes and the cDNA encoding the 121-amino acid isoform of VEGF. Another 10 animals were treated with control plasmid DNA and liposome transfection medium; a third group of 10 animals was given physiologic saline. Each solution was injected directly into the femoral artery distal to the origin of the epigastric pedicle supplying the flap. Four days after injection, the pedicle was ligated and blood flow in the flap was approximated using dye fluorescence. Seven days later, the amount of viable tissue within the flap was measured by planimetry. After the animals were killed, specimens from both the operated and nonoperated sides of the abdomen were harvested for immunohistologic evidence of VEGF protein expression. Average dye fluorescence indices of the three groups (VEGF cDNA, control plasmid, and saline) 2 hours after pedicle ligation were 35.9, 23.9, and 53.9 percent, respectively (p < 0.05). Compared with the two control groups, flaps receiving VEGF cDNA had significantly greater tissue viability at the end of 7 days: 93.9 versus 28.1 percent for the control plasmid DNA group and 31.9 percent for the saline group (p < 0.05). Immunohistochemical staining documented increased deposition of VEGF protein in flaps that were infused with the VEGF cDNA versus saline alone (p < 0.05). The results indicated that the survival of ischemic tissues can be enhanced by administration of a cDNA encoding VEGF, a protein known to be important in the process of angiogenesis and wound healing.

Effect of time on the viability of ischemic skin flaps treated with vascular endothelial growth factor (VEGF) cDNA.

This study examined the efficacy of gene therapy on wound healing. The authors investigated whether delivery of the gene encoding a particular cytokine, known to be important in angiogenesis, could affect ischemic skin flaps. Anterior abdominal skin flaps, based solely on the epigastric artery and vein, were created in the Sprague-Dawley rat model. At the time of elevation, the arterial pedicle supplying each flap was infused either with the gene for vascular endothelial growth factor (VEGF) or physiologic saline alone. The flaps were resutured into place and observed for a period of either 4 or 3 days, at which time the pedicle was ligated. Twenty minutes following ligation, blood flow in the flaps was measured by dye fluorescence. Tissue viability of the flaps was subsequently measured by planimetry after a period of 7 days. Flaps that received the VEGF gene and were ligated at 4 days had an average dye fluorescence index (DFI) of 31.1 following ligation, and 93.9 percent viable tissue after 7 days. Flaps that received saline alone, and were ligated following a similar interval, had an average DFI of 14.0 and 31.9 percent viable tissue. Among the subjects that were ligated at 3 days, only a single, gene-infused flap had any noticeable viable tissue after 7 days. The DFI of these groups was 11.0 for the gene-infused group and 22.1 for the saline-infused group. The results suggest that delivery of the gene for VEGF can improve the survival of ischemic skin flaps, but that the effect of gene therapy is not limitless.

Microsurgical reconstruction of difficult orocutaneous fistulas.

We present 5 cases from a series of eight orocutaneous fistulas that were successfully reconstructed through the use of free tissue transfer. Reconstruction was performed in 5 patients with a radial forearm flap and in 3 patients with a free jejunal transfer. This report addresses the approach to reconstructing the difficult orocutaneous fistula with emphasis on free flap design and the use of a bilaminar flap.

Evaluation of a continuous systemic infusion of iloprost, a stable PGI-2 analog, on the survival of experimental skin flaps.

Numerous investigators have attempted to enhance the survival of ischemic experimental skin flaps using various pharmacologic manipulations. Recently, the authors' laboratory demonstrated the beneficial effect of iloprost, a stable PGI2 analogue, as a post-ischemic perfusion washout, in improving the survival of ischemic skin flaps. The rat unilateral abdominal skin flap, based on the superficial epigastric vessels, was utilized in this study involving 30 animals. The animals were divided into three different treatment groups, with ischemic periods of 16 and 18 hr. Perfusion washouts were performed at the completion of the various ischemic periods. Alzet osmotic pumps were used to deliver a continuous systemic infusion of iloprost for 7 days postoperatively. The groups consisted of the following: Group 1 (single ILO)--perfusion washout with iloprost only; Group 2 (continuous LD ILO)--low-dose systemic iloprost infusion (0.066 mcg/kg/min) and perfusion washout with iloprost; and Group 3 (continuous HD ILO)--high-dose systemic iloprost infusion (0.1 mcg/kg/min) and perfusion washout with iloprost. The percentage of flap survival was assessed on postoperative day 7. Skin flaps of the animals receiving the continuous systemic infusion of iloprost were noted to have varying percentages of survival, while skin flaps undergoing perfusion washout only were found to have either complete survival using a continuous systemic infusion of iloprost, compared to iloprost perfusion washout alone. In addition, the hypotensive side effects of systemic iloprost infusion limit its use in the rat skin-flap model.

Iloprost improves survival of ischemic experimental skin flaps.

The ability of the prostacyclin analogue iloprost to improve survival of ischemic experimental skin flaps was investigated. Unilateral island skin flaps based on the superficial inferior epigastric vessels were raised in 70 rats and subjected to varying lengths of primary ischemia. The flaps were divided into the following four groups: group I, no perfusion washout; group II, postischemic washout with lactated Ringer's solution; group III, postischemic washout with urokinase; and group IV, postischemic washout with iloprost. Flap survival rates for group IV were significantly higher than all other groups (p < 0.05). The primary ischemia time at which 50% of the flaps failed was 8.9 hours for group I, 9.5 hours for group II, 13.3 hours for group III, and 15.3 hours for group IV. This is the first study to investigate the effect of iloprost on skin flap survival. Iloprost was found to be significantly more effective than urokinase in salvaging ischemic experimental skin flaps.

Use of urokinase during secondary ischemia in experimental skin flaps.

The ability of urokinase to salvage experimental flaps after a secondary ischemic insult was investigated in a rat model. Unilateral abdominal island skin flaps based on the superficial inferior epigastric vessels were raised and subjected to either 4 or 6 hours of primary ischemia followed by 12 hours of reperfusion and varying lengths of secondary ischemia. At the conclusion of secondary ischemia, the flaps were perfused with either lactated Ringer's solution or urokinase. One group of flaps served as a control and received no postischemic perfusion washout. The secondary critical ischemia time at which 50% of the flaps failed clinically was greater for flaps perfused with urokinase. Furthermore, the survival rates for all flaps perfused with urokinase were significantly greater than either control flaps or flaps perfused with lactated Ringer's solution (p < 0.05). Flap survival decreased significantly in all groups with increasing primary and/or secondary ischemia time (p < 0.05).

Enhancement of fluorescein perfusion in experimental skin flaps following postischemic washout with iloprost, urokinase, verapamil, and University of Wisconsin solution.

The enhancement of blood flow in experimental skin flaps following postischemic perfusion washout was investigated in rats. Unilateral island skin flaps based on the superficial epigastric vessels were raised and subjected to 6 hr of primary ischemia. Group 1 was designated as a control and did not undergo postischemic perfusion washout. In the remaining rats, postischemic washout was performed with one of five agents: Group 2--lactated Ringer's solution; Group 3--University of Wisconsin solution, an organ preservation medium; Group 4--verapamil, a calcium channel blocker; Group 5--urokinase, a thrombolytic agent; Group 6--iloprost, a stable prostacyclin analog. Two hours following perfusion washout, fluorometric analysis revealed a statistically significant enhancement of blood flow in Groups 4, 5, and 6, compared to Groups 2 and 3 (p < 0.05). Furthermore, a significant increase in skin surface fluorescence was demonstrated in all the flaps that underwent perfusion washout, compared to the control flaps (p < 0.05). By analyzing skin surface fluorescence, the enhancement of nutritive blood flow in flaps, following postischemic perfusion washout, was evaluated. This is the first study in which the above pharmacologic agents were compared in a quantitative manner.

The additive beneficial effect of UW solution and urokinase on experimental microvascular free-flap survival.

Pharmacologic manipulation of free flaps to enhance tolerance to ischemia has become a subject of great interest in the research literature. In an effort to improve survival, perfusion washout of experimental free flaps was performed following an episode of primary ischemia. The perfusates utilized were lactated Ringer's solution (LR), University of Wisconsin solution (UW), a high-molecular-weight medium used in organ preservation, and urokinase, a thrombolytic agent. Seventy-five rats were used in this study and divided into groups of 5 each. A 3 x 6-cm abdominal free flap based on the superficial inferior epigastric vessels was raised in each rat. The free flaps were subjected to either 12 or 18 hr of primary ischemia. Following the period of ischemia, perfusion washout was performed with either LR, UW solution, or urokinase at increasing concentrations alone or in combination with UW solution. Urokinase was first evaluated as a perfusate alone at increasing concentrations. In the 12-hr ischemia group, free-flap survival was shown to increase from 0 percent in the LR-perfused flaps to 20 percent, 60 percent, and 80 percent in flaps perfused with 12,500, 25,000, and 100,000 U of urokinase, respectively (p < 0.05). A similar increase in survival was demonstrated in the 18-hr ischemia group, where 0 percent, 20 percent, and 40 percent of flaps survived following perfusion with 12,500, 25,000, and 100,000 U of urokinase, respectively (p < 0.05). Urokinase was then perfused along with UW solution to evaluate the combined effect on flap survival.(ABSTRACT TRUNCATED AT 250 WORDS)