Isolation of human hepatocytes from livers rejected for liver transplantation on a national basis: Results of a 2-year experience

Abstract

The offer of liver transplantation to many patients affected by liver failure is limited by organ shortage. Clinical application of human-based liver cell therapies, such as bioartificial liver and hepatocyte transplantation, might support liver transplantation, allowing more patients to be treated and decreasing mortality in the waiting list. The development of a standardized method of hepatocyte isolation is a mainstay for large-scale application of liver cell therapy. The aim of this study is to analyze retrospectively a 2-year experience of human hepatocyte isolation from livers rejected from transplantation at organ harvesting, performed on a national basis in Italy. All the livers judged unsuitable for transplantation were considered for hepatocyte isolation. Macrosteatosis greater than 60% was the most common reason of refusal, followed by nonviral cirrhosis. Fifty-four organs were used. Human hepatocyte isolation resulted in more that 7 million liver cells/g of tissue digested with 73% ؎ 14% viability. Steatotic organs gave better results in terms of cell yield than cirrhotic livers. Isolated hepatocytes were able to perform specific liver functions, and evidence of factor IX and albumin messenger RNA (mRNA) production was reported when cells were plated in culture. Modifications of the traditional method of hepatocyte isolation, aimed at reducing ischemia-reperfusion damage and improving post-isolation cell conditions, showed improvements in post-isolation viability. In conclusion, we show that it is possible to use the vast majority of livers not suitable for transplantation on a national basis for human hepatocyte isolation, obtaining a large amount of viable functioning human hepatocytes that might be used for cell transplantation and therapy. (Liver Transpl 2003;9:506-512.)

Hepatocyte transplantation

“…With occasional exceptions, hepatocytes for transplantation are derived from cadaver livers that had been offered for organ transplantation, but were not accepted because of prolonged ischemia or excessive fat content. Thus, the viability of these hepatocytes varies greatly [21]. The ability to cryopreserve hepatocytes is also related, in great part, to the initial quality of the cells.…”

Potential Impact of Steatosis on Cytochrome P450 Enzymes of Human Hepatocytes Isolated from Fatty Liver Grafts

“…Healthy human liver is only occasionally available for experimental purposes, and this scarcity is increased by the competing demands for human cells for clinical applications, such as hepatocyte transplantation programs or artificial liver devices. Normal liver resected during therapeutic hepatectomies, remnants of livers after reduction or split-liver procedures, and human cadaveric livers thought unsuitable for transplantation are potentially good sources of high-quality isolated hepatocytes (Baccarani et al, 2003;Mitry et al, 2003;Serralta et al, 2003;Barbich et al, 2004;Haghighi et al, 2004). High-grade macrosteatosis and cirrhosis are the two principal reasons for organ refusal for transplantation.…”

“…High-grade macrosteatosis and cirrhosis are the two principal reasons for organ refusal for transplantation. Steatotic livers give better results, in terms of cell yield, than cirrhotic organs from hepatocyte isolation procedures (Baccarani et al, 2003). A key question is the functionality of cells isolated from fatty livers.…”

“…However, the impact of hepatic steatosis on the outcome of hepatocyte transplantation and/or culture has not yet been determined. The quality of isolated cells to be used for transplantation has been usually assessed based on cellular viability, plating efficiency, or plasma protein mRNA expression after a short period of culture (Baccarani et al, 2003;Mitry et al, 2003;Lloyd et al, 2004). Some discrepancies have been found in predictions of hepatocyte functionality after transplantation based on viability of isolated cells and survival in primary culture (Olinga et al, 2000;Nishitai et al, 2005).…”

Hepatocytes isolated from neoplastic liver-immunomagnetic purging as a new source for transplantation

“…Currently, the standard approach is to isolate hepatocytes from livers rejected for liver transplantation due to excessive steatosis, cirrhosis and prolonged ischemia. However, availability of hepatocytes from this source is in increasingly short supply and concerns with the functional capability of such cells have been raised [12] . Our group recently reported the feasibility of harvesting tumor-free hepatocytes from macroscopically nor mal liver unavoidably removed during hepatic resection for malignancy [13] .…”