Experimental studies of human fetal liver cells : in regard to in utero hematopoietic stem cell transplantation

Abstract

Successful in utero hematopoietic stem cell transplantation represents a potential step forward in the management of patients with severe inherited hematological, metabolic and immunological disorders. Animal data from a number of species have demonstrated evidence of engraftment, but to date the only effective in utero transplantations (IUTx) in humans have been performed in fetuses with severe immunodeficiencies, suggesting that the fetal immune system play a vital role in the success or failure of in utero transplantation.

To study the capacity of allogenic response of the human fetus in first and second trimesters, mixed lymphocyte culture (MLC) of fetal liver cells (FLC) was performed. FLC were able to recognize and react to allogenic stimulation already from 12 weeks' gestation and in some cases even earlier. This response varied according to not only gestational ages, but also between fetuses of the same gestational age. In all responding livers there was a tendency for increased responsiveness with increasing gestational age and the number of responding fetuses also increased with gestational age.

Depletion of human leukocyte antigen (HLA) class 11+ cells from the stimulator cells resulted in a decreased stimulation index (SI) in a majority of the experiments, indicating that the reactivity is HLA class H dependent. In order to further study the role of CD3+ (T-cells) and CD56+ (NK-cells) cells in the alloreactivity in fetal cells, MLC was performed prior to and after depletion of these cell subpopulations. The MLC response was reduced either after CD3+ or CD56+ cell depletion from responder cells. Both cell- types may exhibit complementary effects in fetal alloreactivity.

The possibility to decrease colony-forming capacity in FLC was also explored. Such regimens have been discussed as a way to create an advantage for transplanted cells to increase the potential for homing and engraftment after in utero stem cell transplantation. Parvovirus B 19 is known to inhibit erythropoietic colony formation in vitro. The hypothesis that recombinant B 19 empty capsid proteins (VP1, VP2) could themselves suppress erythropoietic colony formation in the absence of nonstructural proteins, was explored. The B 19 capsids were shown to inhibit colony formation of BFU-E (70-95%). When B 19 capsids were pre-incubated with anti-B 19 monoclonal antibody or IgG positive sera, the inhibitory effect on BFU-E was completely blocked. The B 19 capsids inhibitory effect could also be reduced by at least 25% by pre-incubating the FLC with anti-P monoclonal antibody.

The sensitivity of fetal hematopoietic stem cells to chemotherapeutic and immunomodulative agents, measured as colony-forming capacity was tested. FLC were incubated with doxorubicin, daunorubicin, antithymocyte-globulin (ATG), Orthoclone OKT-3 and betamethasone. All drugs tested were capable in variable degrees to decrease colony-forming capacity of fetal hematopoietic progenitor cells. However, cytotoxic drugs may adversely affect the fetus. These potential risks needs to be compared to the severity of the disease to be treated and the gains of transplantation.

A case of IUTx with cryopreserved FLC given at 14 weeks by a single intraperitoneal injection to a male fetus with X-linked SCID is reported. Engraftment was detected at 24 weeks, and the fetus had a normal T-cell count at 33 weeks. Complete reconstitution of T-cells was achieved before birth, and the clinical course during the first two years of life has been completely uneventful. Although experience of IUTx in X-linked SCID is still limited, IUTx might be offered as a therapeutic option in cases where the parents are considering termination of an affected fetus identified at prenatal diagnosis.