title: Autologous transplantation of amniotic fluid-derived mesenchymal stem cells into sheep fetuses.
abstract: Long-term engraftment and phenotype correction has been difficult to achieve in humans after in utero stem cell transplantation mainly because of allogeneic rejection. Autologous cells could be obtained during gestation from the amniotic fluid with minimal risk for the fetus and the  mother. Using a sheep model, we explored the possibility of using amniotic fluid mesenchymal stem cells (AFMSCs) for autologous in utero stem cell/gene therapy. We collected amniotic fluid (AF) under ultrasound-guided amniocentesis in early gestation pregnant sheep (n = 9, 58 days of  gestation, term = 145 days). AFMSCs were isolated and expanded in all sampled fetal sheep. Those cells were transduced using an HIV vector encoding enhanced green fluorescent protein (GFP) with 63.2% (range 38.3-96.2%) transduction efficiency rate. After expansion, transduced AFMSCs  were injected into the peritoneal cavity of each donor fetal sheep at 76 days under ultrasound guidance. One ewe miscarried twin fetuses after amniocentesis. Intraperitoneal injection was successful in the remaining 7 fetal sheep giving a 78% survival for the full procedure. Tissues were sampled  at postmortem examination 2 weeks later. PCR analysis detected GFP-positive cells in fetal tissues including liver, heart, placenta, membrane, umbilical cord, adrenal gland, and muscle. GFP protein was detected in these tissues by Western blotting and further confirmed by cytofluorimetric  and immunofluorescence analyses. This is the first demonstration of autologous stem cell transplantation in the fetus using AFMSCs. Autologous cells derived from AF showed widespread organ migration and could offer an alternative way to ameliorate prenatal congenital disease.
authors.affiliation: Prenatal Cell and Gene Therapy Group, Institute for Women’s Health, University College London, London, UK.
authors.forename: S W Steven
authors.initials: SW
authors.lastname: Shaw
authors.forename: Sveva
authors.initials: S
authors.lastname: Bollini
authors.forename: Khalil Abi
authors.initials: KA
authors.lastname: Nader
authors.forename: Annalisa
authors.initials: A
authors.lastname: Gastaldello
authors.forename: Annalisa
authors.initials: A
authors.lastname: Gastadello
authors.forename: Vedanta
authors.initials: V
authors.lastname: Mehta
authors.forename: Elisa
authors.initials: E
authors.lastname: Filppi
authors.forename: Mara
authors.initials: M
authors.lastname: Cananzi
authors.forename: H Bobby
authors.initials: HB
authors.lastname: Gaspar
authors.forename: Waseem
authors.initials: W
authors.lastname: Qasim
authors.forename: Paolo
authors.initials: P
authors.lastname: De Coppi
authors.forename: Anna L
authors.initials: AL
authors.lastname: David
mesh_terms: D000649:Amniocentesis; D000653:Amniotic Fluid / Q000166:cytology*; D000818:Animals; D000013:Congenital Abnormalities / Q000517:prevention & control; D005260:Female; D005822:Genetic Vectors / Q000378:metabolism; D005865:Gestational Age; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D045164:Mesenchymal Stem Cell Transplantation; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D023421:Models, Animal; D011247:Pregnancy; D011270:Pregnancy, Animal; D012756:Sheep; D014182:Transplantation, Autologous
chemical_list: C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins