Rectus Abdominis Flap Replantation after 18 h Hypothermic Extracorporeal Perfusion-A Porcine Model

Abstract

Cold storage remains the clinical standard for composite tissue preservation but is time-limited. A long ischemia time during surgery will adversely affect postoperative outcomes due to ischemia-reperfusion injury. Extracorporeal perfusion (ECP) seems to be a promising alternative for prolonged preservation, but more evidence is needed to support its use and to identify optimal perfusion fluids. This article assessed musculocutaneous flap vitality after prolonged ECP and compared outcomes after replantation to short static cold storage (SCS). Unilateral musculocutaneous rectus abdominis flaps were raised from 15 pigs and preserved by 4 h SCS (n = 5), 18 h mid-thermic ECP with Histidine-Tryptophan-Ketoglutarate (HTK, n = 5) or University of Wisconsin solution (UW, n = 5). Flaps were replanted and observed for 12 h. Skeletal muscle histology was assessed (score 0-12; high scores equal more damage), blood and perfusate samples were collected and weight was recorded as a marker for oedema. Mean histological scores were 4.0 after HTK preservation, 5.6 after UW perfusion and 5.0 after SCS (p = 0.366). Creatinine kinase (CK) was higher after ECP compared to SCS (p < 0.001). No weight increase was observed during UW perfusion, but increased 56% during HTK perfusion. Following 12 h reperfusion, mean weight gain reduced 39% in the HTK group and increased 24% in the UW group and 17% in the SCS group. To conclude, skeletal muscle seemed well preserved after 18 h ECP with HTK or UW perfusion, with comparable histological results to 4 h SCS upon short reperfusion. The high oedema rate during HTK perfusion remains a challenge that needs to be further addressed.

Keywords: VCA; composite tissue transplantation; ex vivo preservation; free flap; mid-thermic storage.

Figure 1

Schematic illustration of the extracorporeal perfusion set-up. The semi-closed perfusion set-up (closable box) with a flap in the perfusion box on top of a metal grid. This allows passive venous drainage of the preservation fluid into the collection reservoir. The centrifugal pump is pressure regulated at in-line ≤ 30 mmHg and the fluid is infused with a mix of 95% O₂ and 5% CO₂ in the oxygenator. The heater–cooler machine cools the fluid to 8–10 °C. There is a drug administration point and fluid sampling point. Before entering the flap via the artery, the ultrasonic flow meter measures flow per minute.

Figure 2

Morphological features of degenerative muscle fibres due to ischemia-reperfusion injury in transverse muscle sections stained with H&E at 12 h after flap replantation. Upper left: variation in muscle fibre shape and size. Oedema and inflammation are also seen. Upper right: hypoxic myocyte (rounded and hypereosinophilic). Lower left: neutrophil influx in the myocyte. Lower middle: necrotic, pale fibres and phagocytized myocytes. Lower right: Masson’s trichrome staining, fibrotic myocytes are stained green.

Figure 3

Photographs of all flaps at 12 h after replantation. * Flap 1 was excluded due to therapy-resistant venous congestion and later arterial obstruction. ** Flap 5 at 11 h (left) and 12 h (right) after flap replantation (before and after acute arterial thrombosis). *** Flap 14 underwent successful revision of the arterial pedicle at 8 h after replantation and had normal appearance afterwards. Abbreviations: CS; cold storage, UW; University of Wisconsin solution, HTK; histidine-tryptophan-ketoglutarate solution.

Figure 4

Mean CK (a), potassium (b) and lactate (c) per pig. Arterial measurements were taken during flap retrieval and after flap replantation. Porcine reference values: CK 153–5427 U/L, potassium 3.7–6.1 mmol/L, lactate unknown [24].

Figure 5

Boxplot showing an overview of histological changes at the study endpoint indicating ischemia-reperfusion injury per flap and group. Cell count per 10 high-power fields at 20× magnification. Scoring for inflammation and interstitial oedema: 0 = no, 1 = minimal, 2 = intermediate, 3 = diffuse.

Figure 6

Mean flap weight over time per group. Time points of flap weight measurements: directly after flap retrieval, after 4 h cold storage or 18 h ECP and after 12 h reperfusion.