TY - JOUR
T1 - Machine perfusion enhances hepatocyte isolation yields from ischemic livers
AU - Izamis, Maria Louisa
AU - Perk, Sinem
AU - Calhoun, Candice
AU - Uygun, Korkut
AU - Yarmush, Martin L.
AU - Berthiaume, François
N1 - Funding Information:
Funding from the US National Institutes of Health ( R01DK59766 , R01DK096075 , R00DK088962 ), the Shriners Hospitals for Children, and Lifeblood Medical Inc. is gratefully acknowledged.
Publisher Copyright:
© 2015 Elsevier Inc.
PY - 2015/10/1
Y1 - 2015/10/1
N2 - Background: High-quality human hepatocytes form the basis of drug safety and efficacy tests, cell-based therapies, and bridge-to-transplantation devices. Presently the only supply of cells derives from an inadequate pool of suboptimal disqualified donor livers. Here we evaluated whether machine perfusion could ameliorate ischemic injury that many of these livers experience prior to hepatocyte isolation. Methods: Non-heparinized female Lewis rat livers were exposed to an hour of warm ischemia (34°C) and then perfused for 3h. Five different perfusion conditions that utilized the cell isolation apparatus were investigated, namely: (1) modified Williams Medium E and (2) Lifor, both with active oxygenation (95%O2/5%CO2), as well as (3) Lifor passively oxygenated with ambient air (21%O2/0.04%CO2), all at ambient temperatures (20±2°C). At hypothermic temperatures (5±1°C) and under passive oxygenation were (4) University of Wisconsin solution (UW) and (5) Vasosol. Negative and positive control groups comprised livers that had ischemia (WI) and livers that did not (Fresh) prior to cell isolation, respectively. Results: Fresh livers yielded 32. ±. 9. million cells/g liver while an hour of ischemia reduced the cell yield to 1.6. ±. 0.6. million cells/g liver. Oxygenated Williams Medium E and Lifor recovered yields of 39. ±. 11 and 31. ±. 2.3. million cells/g liver, respectively. The passively oxygenated groups produced 15. ±. 7 (Lifor), 13. ±. 7 (Vasosol), and 10. ±. 6 (UW). million cells/g liver. Oxygenated Williams Medium E was most effective at sustaining pH values, avoiding the accumulation of lactate, minimizing edematous weight gain and producing bile during perfusion. Conclusions: Machine perfusion results in a dramatic increase in cell yields from livers that have had up to an hour of warm ischemia, but perfusate choice significantly impacts the extent of recovery. Oxygenated Williams Medium E at room temperature is superior to Lifor, UW and Vasosol, largely facilitated by its high oxygen content and low viscosity.
AB - Background: High-quality human hepatocytes form the basis of drug safety and efficacy tests, cell-based therapies, and bridge-to-transplantation devices. Presently the only supply of cells derives from an inadequate pool of suboptimal disqualified donor livers. Here we evaluated whether machine perfusion could ameliorate ischemic injury that many of these livers experience prior to hepatocyte isolation. Methods: Non-heparinized female Lewis rat livers were exposed to an hour of warm ischemia (34°C) and then perfused for 3h. Five different perfusion conditions that utilized the cell isolation apparatus were investigated, namely: (1) modified Williams Medium E and (2) Lifor, both with active oxygenation (95%O2/5%CO2), as well as (3) Lifor passively oxygenated with ambient air (21%O2/0.04%CO2), all at ambient temperatures (20±2°C). At hypothermic temperatures (5±1°C) and under passive oxygenation were (4) University of Wisconsin solution (UW) and (5) Vasosol. Negative and positive control groups comprised livers that had ischemia (WI) and livers that did not (Fresh) prior to cell isolation, respectively. Results: Fresh livers yielded 32. ±. 9. million cells/g liver while an hour of ischemia reduced the cell yield to 1.6. ±. 0.6. million cells/g liver. Oxygenated Williams Medium E and Lifor recovered yields of 39. ±. 11 and 31. ±. 2.3. million cells/g liver, respectively. The passively oxygenated groups produced 15. ±. 7 (Lifor), 13. ±. 7 (Vasosol), and 10. ±. 6 (UW). million cells/g liver. Oxygenated Williams Medium E was most effective at sustaining pH values, avoiding the accumulation of lactate, minimizing edematous weight gain and producing bile during perfusion. Conclusions: Machine perfusion results in a dramatic increase in cell yields from livers that have had up to an hour of warm ischemia, but perfusate choice significantly impacts the extent of recovery. Oxygenated Williams Medium E at room temperature is superior to Lifor, UW and Vasosol, largely facilitated by its high oxygen content and low viscosity.
KW - Hepatocytes
KW - Isolation
KW - Lifor
KW - Perfusion
KW - Temperature
KW - UW
KW - Vasosol
KW - Viscosity
KW - Williams E
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U2 - 10.1016/j.cryobiol.2015.07.006
DO - 10.1016/j.cryobiol.2015.07.006
M3 - Article
C2 - 26188080
AN - SCOPUS:84942293170
SN - 0011-2240
VL - 71
SP - 244
EP - 255
JO - Cryobiology
JF - Cryobiology
IS - 2
M1 - 3643
ER -