Ex Vivo Subnormothermic Preservation of Porcine Superior Epigastric Artery Perforator Flaps

PURPOSE: Reconstruction of complex head and neck, breast, and extremity defects often require autologous transfer of vascularized tissues and restoration of perfusion microsurgical repair of the vascular pedicle. Flap failure, caused by vascular thrombosis (~5.1%), leads to significant patient morbidity, longer hospitalizations, and greater health care costs. In vessel-depleted or sick patients, free flap reconstruction is challenging. The aim of this study was to develop a protocol for Ex Vivo Subnormothermic Preservation (EVSNP) of flaps. We hypothesized that EVSNP can maintain flaps in near physiologic conditions for at least 12 hours. METHODS: Twelve superior epigastric artery perforator flaps were procured from Yorkshire pigs. Flaps were preserved using ex vivo subnormothermic perfusion (EVSNP, n=6) with an oxygenated colloid solution containing HBOC-21 as oxygen carrier or at 4° C (static cold storage control) for 12h (n=6). Outcome measures, including perfusate dynamics, temperature, gases, metabolites, electrolytes, and flap weight, were monitored and evaluated using Pearson correlations and paired t-tests. Skin biopsies were taken every 6 hours for Hematoxylin and Eosin (H&E) histological evaluation (perivascular inflammation, spongiosis, vacuolization of epidermal cells and epidermal necrosis). Indocyanine Green (ICG) angiography was utilized to analyze skin perfusion before division of the flap pedicle and after 12 hours of EVNP. RESULTS: Mean perfusate flow was 10±0 ml/min at baseline and increased to 16±2 ml/min at TP12 (p=0.002). Mean arterial pressure (45±13 mmHg) remained stable during EVNP (r=0.08, p=0.78). Mean perfusate and flap temperatures were 30.9±1.4° C and 28.4±1.6° C, respectively. Mean arterial PaO2 was 490±74 mmHg and decreased from 566±41 at baseline to 448±56 at perfusion end (p=0.004). PaCO2 was 21±1 mmHg on average, not changing significantly from baseline (TP0 21±1 vs. TP12 21±1, p=0.19). PvCO2 was 24±3 at baseline and decreased to 19±1 at TP12 (p=0.04). The average pH was 7.37±0.02 and was comparable to baseline at TP12 (TP0.5 7.36±0.02 vs. TP12 7.37±0.02, p=0.14). Mean arterial glucose was 4.7±0.7mmol/L. Venous lactate was 5.1±0.8 mmol/L and remained comparable at perfusion end (TP0.5 4.8±0.5 mmol/L vs. TP12 5.4±1.3 mmol/L, p=0.18). Creatine kinase increased over time (TP0.5 864±468 U/L vs. TP12 6340±1764 U/L, p=0.001; r=0.99, p=0.01). Venous methemoglobin was 32.3±10.0%, and increased during perfusion (r=0.96, p<0.0001). Potassium remained in a physiologic range (mean 3.9±0.22 mEq/L), increasing from 3.7±0.2 mEq/L at baseline to 4.1±0.2 at TP12 (p=0.0005). Sodium was slightly elevated (mean 159±3 mEq/L) and increased from baseline 157±0.4 mEq/L to 162±2 mEq/L (p=0.0004). Flap weight did not change from beginning to end of perfusion (TP0 0.222±0.041 kg vs. TP12 0.223±0.044 kg, p=0.48) or in controls (TP0 0.136±0.067 kg vs. TP12 0.135±0.067 g, p=0.48). There was no difference between the percent change in perfused and control flap weight from baseline to 12 hours (p=0.09). H&E of perfused skin biopsies did not reveal histopathological damage. Perfusion TP12 ICG angiography revealed well-perfused flaps with regional differences. CONCLUSION: Ex vivo subnormothermic perfusion preserved flaps in physiologic conditions for 12 hours. Physiologic parameters were maintained without development of edema and weight gain.