Evaluation of a new solution for Hypothermic Machine Perfusion (HMP) of the liver. I - Composition and physicochemical parameters.

PASCUCCI F.A.; CARNEVALE M.E.; BALABAN C.L.; MAMPRIN M.E.; GUIBERT E.E.; RODRIGUEZ J.V.

Rosario

Congreso; 49th meeting of the Society for Cryobiology; 2012

Society for Cryobiology / Centro Binacional (Argentina - Italia) de Investigaciones en Criobiología Clínica y Aplicada

There are many experimental as well as clinical reports indicating that HMP of marginal donor livers are superior to cold static preservation. However, there are still doubts as to the best perfusion preservation solution to use and the role of oxygenation during the HMP. For these reasons, we have developed a BES - Gluconate - Polyethylene Glycol based solution (BGP-HMP). This solution is used to protect the graft providing an appropriate oxygen concentration and by maintaining pH and vascular oncotic control during the HMP. The additions of substrates to decrease the ischemia reperfusion injury were also considered. Material and methods: BGP-HMP solution composition (mM) was; 100 Na+ gluconate, 7 K+ gluconate, 20 sucrose, 30 BES, 2.5 KH2PO4, 0.03 polyethylene glycol 35 kDA, 5 MgSO4, 3 glutathione, 5 adenosine, 15 glycine, 0.25 mg/mL streptomycin and 10 (U/mL) penicillin G. Osm: 297 ± 4 mOsm/Kg H2O, pH= 7.40, Na+ = 120 ± 2 and K+ = 10 ± 1 mEq/L. The freezing point and the buffering capacity (BC) ant 5 and 10 °C of the BGP-HMP solution were determined (Mamprin ME, Cryoletters 29:121-133, 2008) and compared with HTK (Bretschneider solution) as the reference solution. The solubility curves of O2 in BGP-HMP and HKT solutions were determined (Llarrull MS, Cryoletters 28:313-328, 2007). The O2 consumption of isolated rat livers perfused at 5 and 10°C with BGP-HMP or HTK solutions saturated with air were also measured (Rodriguez JV, Cryoletters 30:335-346 (2009). Results: The specific freezing point of the BGP-HMP sol. was -0.55 ± 0.05 °C and the BC were: BGP-HMP 15.44 and HTK 59 mEqH+/UpH/l solution respectively, there were no differences between 5 and 10°C. The O2 solubility at 5 and 10°C and 760 mmHg was 351 and 314 uM O2 for BGP-HMP and 370 and 325 uM O2 for HTK. The O2 consumption of rat livers after 5 hours of perfusion at 5 and 10 °C with BGP-HMP was: 33 ± 5 and 61 ± 12, (n=4) and with HTK 38 ± 7 and 57 ± 4, (n=4) nmol O2/min/g.liver respectively. The livers were perfused at constant pressure (70 ± 10 mmH2O) and the perfusion flow was 0.15 ± 0.02 mL/min/g.liver. The O2 extraction by the hypothermic perfused livers was 0.46 ± 0.16 and 0.67 ± 0.17 for BGP-HMP and 0.36 ± 0.01 and 0.45 ± 0.03 for HTK solution at 5 and 10°C respectively. Control livers perfused with Krebs-Henseleit-Dextran solution at 37 °C consume 1150 ± 10 nmol O2/min/g.liver (n=6), the extraction ratio was 0.86 ± 0.03. Conclusion: we have included three key components in the BGP-HMP solution, gluconate as a non-diffusible anion, the aminosulfonic acid BES to improve the BC and PEG 3500 to provide vascular oncotic support. The BC of our solution is lower than HTK solution but was sufficient to maintain the pH during 24 hr of hypothermic perfusion. The BGP-HMP solution could be saturated with medicinal air and deliver the adequate O2 to maintain the low respiration rate of rat livers at 5 or 10 °C.