The naturally occurring stem cell migratory patterns, the availability of expanding homing and engraftment sites, and the presence of tissue/organ-specific signals in the developing mammalian fetus provide the ideal setting for stem cells to exhibit their full biological potential. These characteristics combined with the relative immunological naivete of the early gestational age fetus that permits the engraftment and long- term persistence of allogeneic and xenogeneic donor stem cells make it possible to use the developing fetus to assess the in vivo potential of a variety of stem cells. We have taken advantage of these permissive characteristics of the fetus to develop a large animal model of human hematopoiesis in sheep that permits not only the long-term engraftment of human hematopoietic stem cell/progenitor cells and their differentiation into the full range of lymphohematopoietic elements, but also the relatively robust expression of their potential to contribute to the formation of non-hematopoietic tissues.
Animals ; Cell Differentiation ; Humans ; *Models, Animal ; Sheep/*embryology ; *Stem Cell Transplantation ; Stem Cells/*cytology ; Transplantation, Heterologous

In order to enhance the efficiency of sheep somatic cell nuclear transfer, we used a chemically assisted enucleation with colchicine to study the effects of the concentration of colchicine, the incubation time of oocytes in colchicine and the maturation time of oocytes on the enucleation rates and the development of reconstructed embryos. The results showed that 1) there were no significant differences in the rates of cytoplast protrusion and enucleation between oocytes that were incubated in colchicine (0.4 microg/mL) for 0.5 h and oocytes that were incubated in colchicine (0.4 microg/mL) for 1 h, and the rate of cytoplast protrusion can be 85.4% while the rate of cytoplast enucleation is 100%. 2) There was no significant difference in oocyte enucleation between oocytes treated with medium containing 0.2 microg/mL colchicine for 0.5 h and oocytes treated with medium containing 0.4 microg/mL colchicine for 0.5 h. 3) A maturation time of 18-23 h did not affect the rates of cytoplast protrusion and enucleation by chemically assisted enucleation, whereas the rate of enucleation of oocytes by blind enucleation was found to decrease with a prolonged incubation time. 4) The development rates of reconstructed embryos could not be influenced by these two enucleation methods, increased from oocytes matured for 21-23 h. These results demonstrate that sheep oocytes can be enucleated fast and effectively by optimized colcholine chemically assisted enucleation, which can enhance the enucleation rate of sheep oocytes and the early development of reconstructed embryos in vitro.
Animals ; Cloning, Organism ; methods ; Colchicine ; pharmacology ; Embryo, Mammalian ; embryology ; Female ; Nuclear Transfer Techniques ; veterinary ; Oocytes ; cytology ; drug effects ; Sheep

The purpose of this study was to determine if mild hypoxia alters the responsiveness to vasoactive agents in the renal and the femoral arteries in the fetal sheep. Ten pregnant sheep were operated under halothane anesthesia at 116 to 124 days' gestation. A maternal tracheal catheter was placed for infusing compressed air (control group, n=5) or nitrogen (hypoxia group, n=5) starting on post operative day 6 and maintained for 5 days. Femoral and renal arteries were harvested from the fetus to study the constriction response to phenylephrine (PE 10(-9) to 10(-5) mol/L). To determine the involvement of nitric oxide as a modulator of vessel constriction, N-nitro-Larginine methyl ester (L-NAME) was used at a concentration of 10(-4) mol/L in parallel chambers. In the hypoxia group, maternal Pao2 significantly decreased from a base-line of 110.4 +/-1.4 to 80.5 +/-1.6 (mmHg, p <0.01), fetal Pao2 significantly decreased from a baseline of 20.9 +/-0.3 to 15.5 +/-0.1 (mmHg, p <0.01). Hypoxia was associated with a significant increase in PE maximal response in the absence (184.5 +/-6.6 vs. 146.2 +/-4.3) and presence (166.9 +/-6.3 vs. 145.0 +/-4.5) of L-NAME, and a decrease in EC50 in the absence (6.0 +/-1.1 vs. 27.0 +/-4.1) of L-NAME of femoral arteries. However, there were no significant differences in PE maximal response and EC50 in the absence and presence of L-NAME of renal arteries. We concluded that mild chronic hypoxia seems to increase the fetal femoral artery response to PE, but not in the fetal renal artery. This observation is consistent with a redistribution of cardiac output away from the carcass.
Animals ; *Anoxia ; Blood Glucose/metabolism ; Dose-Response Relationship, Drug ; Enzyme Inhibitors/pharmacology ; Femoral Artery/*embryology/*pathology ; Hematocrit ; Hydrogen-Ion Concentration ; Kidney/blood supply ; Lactates/blood/metabolism ; NG-Nitroarginine Methyl Ester/pharmacology ; Nitric Oxide/metabolism ; Nitric-Oxide Synthase/antagonists & inhibitors ; Phenylephrine/chemistry/metabolism/pharmacology ; Renal Artery/pathology ; Sheep/*embryology ; Time Factors ; Vasoconstrictor Agents/*pharmacology