A murine model for human cord blood transplantation: near-term fetal and
neonatal peripheral blood cells can achieve long-term bone marrow
engraftment in sublethally irradiated adult recipients
A Scaradavou, L Isola, P Rubinstein, Y Galperin, V Najfeld, D Berlin, J Gordon and RS Weinberg
Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, New York,
USA.
The purposes of the research reported here were first to explore a murine
model for human placental and umbilical cord blood transplantation and
second to evaluate the engraftment ability of ex vivo cultured
hematopoietic cells. Murine near-term fetal and neonatal peripheral blood
(FNPB) cells, genetically marked with the human multiple drug resistance
transgene (MDR1) were used for syngeneic transplants into sublethally
irradiated adult mice. Donor cells were transplanted either fresh and
untreated, or after ex vivo culture in the presence of the hematopoietic
growth factors recombinant murine stem cell factor, recombinant human
interleukin-3 (rHu IL-3), and rHu IL-6, in a liquid culture system. To
evaluate, count, and characterize FNPB progenitor cell-derived colonies,
neonatal mouse mononuclear cells were cultured directly in methylcellulose
with growth factors. To assess their ex vivo expansion ability, FNPB
mononuclear cells were first cultured in liquid medium for 3 to 8 days and
then transferred to semisolid assay plates. Evaluation of the cell counts
after liquid culture showed a 1.4- to 11.6-fold increase, and the numbers
of colonies observed in methylcellulose were similar to those produced by
fresh FNPB cells. Donor-type engraftment was demonstrated by polymerase
chain reaction (PCR) amplification of the human MDR1 transgene in the
peripheral blood of all surviving animals (5 of 7 recipients of the fresh,
and 3 of 8 recipients of the ex vivo-cultured cells) 2 to 4 months after
transplantation. The proportion of donor leukocytes in the peripheral blood
of the recipients (chimerism) was evaluated using fluorescence in situ
hybridization (FISH) analysis 4 to 6 months after transplantation and
ranged from 2% to 26%. In addition, bone marrow cultures were obtained from
two recipient animals: one had received fresh-untreated cells and was
evaluated 8 months after transplant, the other had received ex vivo
cultured cells and was tested 14 months after grafting. The derived
hematopoietic colonies were tested by PCR and the transgene was detected,
conclusively proving long-term engraftment of donor cells. These results
indicate that FNPB transplants can be successfully performed in sublethally
irradiated mice with and without ex vivo culture. Long-term donor-type
engraftment with sustained chimerism has been demonstrated. Thus, murine
neonatal blood grafts can be used as an animal model for cord blood
transplantation for gene therapy studies where complete myeloablation is
not desirable and partial replacement of defective marrow may be
sufficient. Furthermore, the possibility of numerically expanding
hematopoietic progenitor cells contained in neonatal blood without
affecting their engraftment ability could facilitate use of cord blood
grafts in adult recipients.
Volume 89,
Issue 3,
pp. 1089-1099,
02/01/1997
Copyright © 1997 by The American Society of Hematology
