Proof of concept study for a closed ex vivo limb perfusion system for 24-hour subnormothermic preservation using acellular perfusate.

Abstract

The two approaches to vascularized tissue machine perfusion use either the open (nonpressurized) or closed (pressurized) perfusion system. Most studies describing isolated limb perfusion preservation rely on open perfusion systems and report tissue edema exceeding 40% after 12 to 14 hours of preservation. A variant of machine perfusion places the limb and perfusate into a reservoir closed to atmosphere. It is hypothesized that the reservoir pressure, acting as a transmural pressure, has the advantage of reducing edema formation by counteracting the hydrostatic pressure gradient from the perfusion pressure. This proof-of-concept study aim was to demonstrate feasibility of the Universal Limb Stasis System for Extended Storage (ULiSSES) device (closed, vertical perfusion system) to preserve forelimbs of Sus scrofa swine for 24 hours of subnormothermic perfusion compared with an open, horizontal perfusion system. The ULiSSES is a compact, practical device that applies pulsatile, pressurized perfusion through the novel use of a diaphragm pump powered by compressed oxygen. Forelimbs from swine were preserved in ULiSSES device (closed perfusion system) (n = 9) and in an open perfusion system (n = 4) using subnormothermic modified Krebs-Henseleit solution. Physiological parameters were measured at the start and every 3 hours for 24 hours. Limbs were weighed before and after perfusion to compare weight gain. Edema and cellular integrity were evaluated using histopathology pre and post perfusion. Closed perfusion system showed superiority compared with the open perfusion system in terms of oxygen consumption, reduction in vascular resistance, and overall tissue integrity. The closed perfusion system demonstrated a 21% reduction in weight gain compared with the open perfusion system and significantly reduced intracellular edema. The ULiSSES closed, pressurized perfusion technology has translatable military applications with the potential to preserve porcine limbs for 24 hours with improved results compared with an open perfusion system.