HEPATOCELLULAR TRANSPLANTATION AND TARGETING GENETIC MARKERS TO HEPATIC CELLS 
r 
C. Hepatocellular harvest, cultivation, and transplantation. li 
l| 
HCT has never been performed in human subjects. (Transplantation of cells harvested ’! 
from fetal liver has been reported (Touraine et al, 1979), though this procedure is , [ 
really a bone marrow transplant with extramedullary stem cells and hepatocellular ■ 
engraftment was not reported.) Considerable progress has been made, however, in ' 
developing methods for harvesting, cultivating, and transplanting hepatocytes in animal «i [ 
models. ;i| 
Hepatocytes are commonly harvested by treatment of donor organs with sequential 
perfusion using EDTA solution, which releases hepatocytes from their attachments, and 4j 
collagenase, which releases hepatocytes from matrix elements. Hepatocytes are cultured 1 ’ 
in hormonally defined media which have been developed to maintain hepatic morphology and m 
liver specific functions for weeks or months in culture (Jefferson et al, 1984; f 
Darlington et al, 1987; Jacob et al, 1989; Lanford, 1989) and will undergo 3-5 cell ■ 
divisions (Ledley et al, 1987a). A biomatrix support of collagen (Isom et al , 1987) or [ 
type IV collagen has been reported to enhance growth and differentiation (Enat et al, ; 
1984) . Consistent serial passage of hepatocytes has not been reported. Hepatocytes can j * 
be cryopreserved for culture or transplantation using conventional preservation *j 
techniques (Maganto et al, 1988; Fuller et al, 1983; Jamieson et al, 1988b). There have 
been extensive investigations of growth factors which may contribute to the considerable | j 
regenerative capacity of the liver in vivo (Francavilla et al, 1988). Recently, a «' 
putative hepatocyte growth factor has been cloned (Miyazawa et al, 1989), and its 
receptor has been identified. >§ | 
Human hepatocytes have been cultured from many laboratories including our own. The l[l 
conditions for a hormonally defined media which will support the growth and differentia- : 
tion of hepatocytes are somewhat different than those used for rodent cells. In the ! 
SUM media developed by Dr. Gretchen Darlington, and used in our laboratories, optimal 
human hepatocellular proliferation was observed with lower concentrations of insulin ^ 
10"’ vs lO"^ M and Dexamethesone (Darlington et al, unpublished data). A substantially ' 
diffeent media has been developed by Dr. Robert Lanford (Southwest Foundation for j 
Biomedical Research, San Antonio) for cultivation and transformation of hepatocytes from 
primates including baboons and chimpanzee (Lanford et al, 1989: Jacob et al, 1989). f 
This media has been used by Dr. Richard Haye and colleagues at the University of Chicago > 
to cultivate human hepatocytes (Haye et al, unpublished data). The method developed by 4 
Haye et al is particularly important because it is commonly used on human liver samples ■ 
prepared in Belzer's solution which may be necessary in the present work. 4 
Hepatocyte transplantation has been studied extensively in rodents. The most common « 
model is the Gunn rat which has hyperbilirubinemia due to UDP-glucuronyl transferase M 
deficiency and is homologous to Crigler-Najar syndrome in humans. Many laboratories A 
have demonstrated that HCT from a normal rat can lower bilirubin levels. Several 
techniques have been used including injection of hepatocytes into the peritoneum 
(Demetriou et al, 1986a; 1986b; 1986c; 1988), portal vein (Sutherland et al, 1977; Groth 
et al, 1977; Matas et al, 1976), spleen (Vroemen et al, 1985; 1986; 1988a; Fuller et al, m 
1983), fat pads (Jirtle and Michalopoulous , 1982), pancreas (Vroemen et al , 1988b), or # 
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Recombinant DNA Research, Volume 14 
