Human hematopoietic progenitors engraft in fetal canine recipients and expand with neonatal injection of fibroblasts expressing human hematopoietic cytokines.
OBJECTIVE
The development of large-animal models for human hematopoiesis will facilitate the study of human hematopoietic stem cells and their progenitors in vivo. In previous studies, human hematopoietic progenitors engrafted in fetal dogs and contributed to hematopoiesis for one year. Despite initially high levels of human cells, the proportion declined to less than 0.1% at 6 months, possibly due to inability of the canine hematopoietic microenvironment to support ongoing human hematopoiesis. In the current experiments we examined the potential of co-transplanting fibroblasts expressing human hematopoietic cytokines with the hematopoietic graft to increase the contribution of human progenitors to chimeric hematopoiesis.


METHODS
Mid-gestation canine fetuses were injected with 1-3 x 10(7) human cord blood cells and 1 x 10(7) murine fibroblasts engineered to express human cytokines. Neonatal pups were boosted with additional injections of cytokine-expressing fibroblasts. Human cell engraftment was monitored by PCR amplification of human-specific DNA sequences from recipient hematopoietic tissues.


RESULTS
Human hematopoietic cells were detected in 13/15 fetal recipients for at least 7 months. At time points up to 30 weeks of age, human DNA was detected in stimulated lymphocyte cultures, approximately 0.1% of blood leukocytes and 1.5% (85/5757) of myeloid colonies. Eight months postinfusion, 1.7% of colony-forming units (CFUs) were of human origin. By one year 0.5% or less of myeloid colonies and less than 0.01% of blood leukocytes carried human DNA. Following an infusion of cytokine-expressing fibroblasts at one year, the proportion of human myeloid progenitors rose to 11.5% and remained detectable for 8 months.


CONCLUSIONS
These studies confirm that human hematopoietic progenitors can engraft in fetal pups and contribute to multilineage hematopoiesis. Infusion of cells expressing human cytokines is one approach to stimulate human hematopoietic progenitors in vivo and thus increase their contributions to chimeric hematopoiesis.
Department of Clinical Pathology, Sunnybrook and Women's Health Sciences Centre, Toronto, Ontario, Canada.
Carolyn
C
Lutzko
Lisa
L
Meertens
Liheng
L
Li
Yongjun
Y
Zhao
Anthony
A
Abrams-Ogg
J Paul
JP
Woods
Stephen
S
Kruth
Margaret R
MR
Hough
Ian D
ID
Dubé
D019264:Adoptive Transfer; D000818:Animals; D000831:Animals, Newborn; D019070:Cell Lineage; D004247:DNA / Q000032:analysis; D004285:Dogs / Q000196:embryology*; D005312:Fetal Blood / Q000166:cytology; D005347:Fibroblasts / Q000378:metabolism / Q000637:transplantation*; D005865:Gestational Age; D006082:Graft Enhancement, Immunologic; D006085:Graft Survival; D016179:Granulocyte Colony-Stimulating Factor / Q000235:genetics / Q000378:metabolism*; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D007274:Injections, Intraperitoneal; D007377:Interleukin-3 / Q000235:genetics / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000637:transplantation*; D008214:Lymphocytes / Q000166:cytology; D051379:Mice; D022423:Myeloid Cells / Q000166:cytology; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013045:Species Specificity; D019089:Stem Cell Factor / Q000235:genetics / Q000378:metabolism*; D017154:Stromal Cells / Q000378:metabolism / Q000637:transplantation; D014162:Transfection; D014183:Transplantation, Heterologous / Q000276:immunology
D007377:Interleukin-3; D011993:Recombinant Fusion Proteins; D019089:Stem Cell Factor; D016179:Granulocyte Colony-Stimulating Factor; D004247:DNA