Long-Term Hematopoietic Engraftment of Congenic Amniotic Fluid Stem Cells After in Utero Intraperitoneal Transplantation to Immune Competent Mice

Abstract

Clinical success of in utero transplantation (IUT) using allogeneic hematopoietic stem cells (HSCs) has been limited to fetuses that lack an immune response to allogeneic cells due to severe immunological defects, and where transplanted genetically normal cells have a proliferative or survival advantage. Amniotic fluid (AF) is an autologous source of stem cells with hematopoietic potential that could be used to treat congenital blood disorders. We compared the ability of congenic and allogeneic mouse AF stem cells (AFSC) to engraft the hematopoietic system of time-mated C57BL/6J mice (E13.5). At 4 and 16 weeks of age, multilineage donor engraftment was higher in congenic versus allogeneic animals. In vitro mixed lymphocyte reaction confirmed an immune response in the allogeneic group with higher CD4 and CD8 cell counts and increased proliferation of stimulated lymphocytes. IUT with congenic cells resulted in 100% of donor animals having chimerism of around 8% and successful hematopoietic long-term engraftment in immune-competent mice when compared with IUT with allogeneic cells. AFSCs may be useful for autologous cell/gene therapy approaches in fetuses diagnosed with congenital hematopoietic disorders.

In Utero Transplantation of Expanded Autologous Amniotic Fluid Stem Cells Results in Long-Term Hematopoietic Engraftment

“…AFSCs exhibited significant clonogenic potential, producing colonies from all three hematopoietic lineages when cultured in semisolid media. This is in keeping with what we have previously demonstrated , and further highlights the potential of obtaining fetal HSCs from an easily accessible source during gestation . Most importantly, we proved here that following 8 days of culture and a fourfold rise in the number of CD117+ cells, AFSCs maintained a similar phenotype and exhibited significant hematopoietic potential equal to that of freshly isolated AFSCs (similar numbers of hematopoietic colonies were observed when identical numbers of fresh and cultured AFSCs were placed in semisolid media).…”

“…This remarkable engraftment potential may also be related to the intravenous injection technique which is superior to previously described intraperitoneal delivery and is more applicable to possible clinical translation . This is also demonstrated by our recently published preliminary study in which intraperitoneal IUT of 10 4 congenic AFSCs per fetus resulted in long‐term hematopoietic engraftment only of around 10%, which is approximately half to that achieved in the present study using intravenous IUT . Multilineage long‐term engraftment equivalent to that observed in the host was confirmed in the recipient groups.…”

“…I, J). We have observed similar results in fresh AFSCs isolated from mouse strains other than B6‐GFP, including B6.SJL‐Ptprc a Pepc b /BoyJ (CD45.1) and Balb/c (Supporting Information Fig. S3A, S3B).…”

“…Freshly isolated murine AFSCs have a phenotype similar to fetal liver (FL) HSCs and can engraft after postnatal transplantation into adult immunocompromised hosts . We have also recently demonstrated that intraperitoneal IUT of AFSCs generates multilineage hematopoietic engraftment . Similarly, freshly isolated CD34+ sheep AFSCs can be genetically modified overnight and are able to engraft after IUT in the hematopoietic system in an autologous setting .…”

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“…Previous research from the laboratory of Paolo De Coppi (UCL Great Ormond Street Institute of Child Health, London, UK) provided proof‐of principle for the hematopoietic potential of AFSCs and their multilineage hematopoietic engraftment following IUT . However, the loss of hematopoietic potential after in vitro expansion under adherent culture conditions has hindered the development of hAFSCs as a therapy for inherited hematopoietic disorders.…”