HEPATOCELLULAR TRANSPLANTATION AND TARGETING GENETIC MARKERS TO HEPATIC CELLS 
measured by trypan blue exclusion or replating efficiency. We have also observed no 
toxicity from exposure to protamine or recombinant vectors. Therefore we feel that 
plating and cultivation of hepatocytes presently represents the most effective means for 
storing hepatocytes between the time of harvest and transplantation. There is no reason 
to believe we will be endangering the viability of these cells or the probability of a 
successful transplant (which is the primary goal of this research) by exposing these 
cells to protamine and vector during this cultivation. 
The number of cells which will be infused has been calculated from murine and canine 
experiments and adjusted for the recipient's weight. In the murine experiments it was 
possible to infuse 2.5 x 10’ cells/kg into the portal vein and 1 x 10® cells/kg into the 
spleen. In the canine model it has been possible to infuse 1 x 10® cells into 5-6 kg 
animals (2 x 10®/kg) via the splenic vein or direct splenic injection without 
complications. We propose to use 2 x 10® cells/kg in these experiments. 
There are several ways to assess this number relative to the number of cells in the 
patient's liver. Pervious calculations in murine experiments (Ponder et al , 1991) were 
based on an estimate of 10® cells in a mouse (25 grams) or 4 x 10® cells/gm. 
Calculations of the number of cells transplanted were made by quantitation of a secreted 
transgene marker (alphaj-antitrypsin) , £-galactosidase staining of the transduced organ, 
and cell sorting with reasonably consistent results. 
Extrapolating from murine data based on the number of cells/mass (a mouse weighs 
(0,025 kg) we would predict that a 5 kg animal would have 2 x 10^° cells and a 
conventional 70 kg human would have 2.8 x 10“ cells. This number is consistant with an 
order of magnitude of cells in the mature human liver of 10“-10^^. The calculations 
below show data on cell numbers from the murine and canine experiments, numbers proposed 
for this protocol, and numbers extrapolated to the "ideal" 70kg individual for 
comparison. These calculations suggest that the number of cells transplanted will 
represent between 1.4 and 14% of the normal host hepatocytes, and with 50% survival we 
might expect 0.7% to 7% reconstitution with transplanted cells. 
Table 7 . Number of hepatocytes transplanted as a fraction of hepatic cells in 
various experimental systems. 
MOUSE EXPERIMENTS (Data from Ponder et al. 1991) 
portal vein 
spleen 
animal mass (kg) 
total cells 
cells 
5.00E+05 
2 . OOE-t-06 
0.024 
l.OOE+08 
cells/kg 
2.08E-I-07 
8.33E+07 
% cells 
0.50% 
2.00% 
DOG EXPERIMENTS ("Data from 
Woo et al. 
unnublished data) 
cells l.OOE+09 l.OOE+09 5 2.08E+10 
cells/kg 2.00E+08 2.00E+08 
% cells 4.80% 4.80% 
Recombinant DNA Research, Volume 14 
[807] 
