A perfusion system for preparing the victim and the ischemic limb for reimplantation

Abstract

We have developed and tested an original perfusion system for rapid and effect ive simultaneous preparation of the victim and the traumatically amputated limb, after a long period of ischemia, for reimplantation. The extracorporeal circuit of the circulatory system (Fig. 1) includes the main drainage channels 3 and 6, reservoirs for holding the perfusion fluid 7 and 14, peristaltic (roller) pumps 8 and 15, a column 9 containing the hemosorbent, a low-output membrane oxygenator with heat exchanger (LMO-HE) (International Standard) 10, the isolated donor's spleen 11, an ultraviolet source (UVS) 12 for irradiating the blood, the rotor of the RK-0.5 blood fractionator 13, and transducers for measuring perfusion pressure 16 and temperature 17 and 18. During development of the system we initially assumed that the hemosorbent and living isolated hepatocytes can temporarily perform the detoxicating function of the liver. Besides this function, hepatocytes utilize toxic substrates of lipoprotein complexes, which are essential for the restoration of disturbed metabolic processes (aerobic oxidation, etc.) in cells. The action of the donor's spleen, UV irradiation, the AUVM-Dnepr -MN-1 activated carbon fiber adsorbent, and of dioxidine was aimed at neutralizing the damaging action of microorganisms and leading to their elimination. In addition, the UVS stimulates processes of energy metabolism and oxidative phosphorylation, as well as transmembrane mass exchange in isolated hepatocytes, and this must unquestionably make the working of the whole system more efficient. The perfusion medium was obtained by diluting whole blood by one-third of its volume with native plasma, and also by adding heparin in a dose of 5000 U and 30 mi of a 1% solution of dioxidine for every 400 ml of fluid. The final hematoerit reading of the perfusion fluid was 28-32%. The extracorporeal circulation was filled with perfusion fluid, the volume of which corresponded to the mass of the amputated limb (1:1). The temperature of the perfusion fluid entering the main artery was 37"C, the perfusion pressure was 70-80 mm Hg, and the volume velocity of regional perfusion was 40-60 ml/(min.kg). The pO 2 of the perfusion fluid entering the arterial system of the amputated limb fluctuated from 160 to 300 mm Hg. The duration of isolated regional perfusion of the amputated limb and of the limb stump below the tourniquet was 1 h. The AUVM-Dnepr -MN-I activated carbon-f iber adsorbent, soaked with 1% dioxidine solution, was applied to the infected wound surface of the amputated limb. The extracorporeal system (Fig. 1) works in the following way. Perfusion fluid draining from the veins of the limb and stump enters the first reservoir 7, which may be equipped with a volume self-regulator, and it is pumped from the reservoir by the pump 8, through the column 9, which has a volume of 100-250 ml, and which is filled with hemosorbent ( S K N I K or SKN-2K) . Next, at the rate of 30-40 ml/min the perfusion fluid passes through the blood chamber of the LMO-HE (International Standard) chamber 10, where it is saturated with oxygen and heated. The perfusion fluid then passes through the regional vascular bed of the isolated donor's spleen 11 and, passing through the UV source 12, it enters the rotor 13 of the RK-0.5 blood fractionator, containing 100-350 ml of a dense suspension of hepatoeytes. The speed of rotation of the rotor is 600-800 rpm. The design of the RK-0.5 rotor is intended to ensure direct contact between the flowing perfusion fluid and hepatocytes. After the rotor the perfusion fluid drains into the second reservoir 14, which may be equipped likewise with a servo-operated volume regulator. From the reservoir the perfusion fluid is pumped by the roller pump 15 into the arteries of the isolated limb and stump. Isolated living xenogeneic hepatocytes were isolated by a combined enzymic-mechanical method [1, 2]. During regional perfusion of the limb, a dense suspension of hepatocytes containing 2 x 108 cells in 10 mi was used. Hepatocytes kept in the extracorporeal circuit by centrifugal force or by capsules of semipermeable membrane functioned actively on direct contact with the perfusion fluid throughout the period of perfusion (results of electron-