Hematopoietic stem cell engraftment by early-stage in utero transplantation in a mouse model.
A novel intrauterine transplantation (IUT) approach was developed to improve the efficiency of engraftment of hematopoietic stem cells (HSCs). HSCs with a green fluorescent protein (GFP) reporter gene were transplanted in utero on days 12.5, 13.5 and 14.5 post coitum (p.c.). The degree of chimerism of donor cells in recipient newborn mice was examined using fluorescent microscopy, polymerase chain reaction (PCR), fluorescence-activated cell sorting (FACS), and fluorescence in situ hybridization (FISH) analyses. Microscopic examination revealed the presence of green fluorescent signal in the peripheral blood of the chimeric mice. The highest survival rate (47%) as well as the highest chimerism rate (73%) were achieved by our new approach in the newborn mice that were subjected to in utero transplantation (IUT) on day 12.5 p.c. (E12.5) compared to the conventional IUT method. FACS analysis indicated that 1.55+/-1.10% of peripheral blood cells from the newborn mice were GFP-positive donor cells. FISH showed that cells containing the donor-specific GFP sequence were present in the bone marrow (BM) of the chimeric mice. Thus, the efficiency of chimera production with this new method of IUT was significantly improved over the existing IUT techniques and instruments.
Chen
Xin
X
Shanghai Institute of Medical Genetics, Shanghai Children's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China.
Gong
Xiu-li
XL
Katsumata
Makoto
M
Zeng
Yi-tao
YT
Huang
Shu-zhen
SZ
Zeng
Fanyi
F
D000818:Animals; D002469:Cell Separation; D002470:Cell Survival; D002678:Chimera; D046528:Chimerism; D005260:Female; D005434:Flow Cytometry; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000097:blood / Q000235:genetics; D018380:Hematopoietic Stem Cell Transplantation / Q000295:instrumentation / Q000379:methods*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000502:physiology*; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D008815:Mice, Inbred Strains; D008856:Microscopy, Fluorescence; D023421:Models, Animal; D014599:Uterus
D049452:Green Fluorescent Proteins