One approach of regenerative medicine to generate functional hepatic tissue in vitro is decellularization and recellularization, and several protocols for the decellularization of livers of different species have been published. This appears to be the first report on rat liver decellularization by perfusion under oscillating pressure conditions, intending to optimize microperfusion and minimize damage to the ECM. Four decellularization protocols were compared: perfusion via the portal vein (PV) or the hepatic artery (HA), with (+P) or without (-P) oscillating pressure conditions. All rat livers (n = 24) were perfused with 1% Triton X-100 and 1% sodium dodecyl sulphate, each for 90 min with a perfusion rate of 5 ml/min. Perfusion decellularization was observed macroscopically and the decellularized liver matrices were analysed by histology and biochemical analyses (e.g. levels of DNA, glycosaminoglycans and hepatocyte growth factor). Livers decellularized via the hepatic artery and under oscillating pressure showed a more homogeneous decellularization and less remaining DNA, compared with the livers of the other experimental groups. The novel decellularization method described is effective, quick (3 h) and gentle to the extracellular matrix and thus represents an improvement of existing methodology. Copyright © 2014 John Wiley & Sons, Ltd.

Decellularization of livers is a well-established procedure. Data on different reseeding techniques or the functional evolution and reorganization processes of repopulated grafts remains limited. A proprietary, customized bioreactor was established to repopulate decellularized rat livers (n = 21) with primary rat hepatocytes (150 × 10 cells) via the hepatic artery and to subsequently evaluate graft morphology and function during 7 days of ex vivo perfusion. Grafts were analysed at 1 h, 6 h, 12 h, 24 h, 3 days, 5 days and 7 days after recellularization (all n = 3) by immunohistological evaluation, hepatocyte-related enzyme (aspartate transaminase, alanine transaminase and lactate dehydrogenase) and albumin measurement in the perfusate. This appears to be the first available protocol for repopulation of rat livers via the hepatic artery. Within the first 24 h after repopulation, the hepatocytes seemed to migrate out of the vascular network and form clusters in the parenchymal space around the vessels. Graft function increased for the first 24 h after repopulation with a significantly higher function compared to standard two-dimensional culture after 24 h. Thereafter, graft function constantly decreased with significantly lower values after 6 days and 7 days of perfusion, although histologically viable hepatocytes were found even after this period. The data suggests that, owing to a constant loss of function, repopulated grafts should potentially be implanted as soon as cell engraftment and graft re-organization are completed. Copyright © 2016 John Wiley & Sons, Ltd.

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