Microchimerism does not induce tolerance after in utero transplantation and may lead to the development of alloreactivity

In utero transplantation is a new technology that may provide non-toxic treatment for congenital disorders. However, a decade of research on in utero transplantation has demonstrated a low degree of chimerism and tolerance in small and large animal models as well as in human beings. We hypothesized that if large numbers of purified stem cells/progenitors were injected, a higher degree of tolerance would be induced. We have performed a 2-year experiment designed to study chimerism and tolerance after in utero transplantation with large numbers of cytokine-recruited C-kit+ cells. Chimerism in the blood and tissues was tested through the lifespan of the animals, and in vitro immunologic assays were performed at the end of life. C-kit(+) cells obtained from the peripheral blood of C57BL/6 mice were injected intraperitoneally into 12- to 13-day-old Balb/c murine fetuses. The injected populations contained 5% to 20% of Sca-1+ and 1% to 5% of CD3+ cells, Twenty-three percent of mice that received transplants showed circulating donor cells in the blood, and 7% to 14% showed donor cells in the tissues. The percent of donor cells in the blood and tissues was low (<0.01%). Timing of injection or cell dose did not affect chimerism or tolerance. Fifty percent (13 of 26) showed accelerated skin graft rejection and 5 of 26 (19%) had prolonged acceptance as compared with control mice not receiving transplants in utero. All mice that rejected skin grafts showed significantly increased natural killer function as compared with the mice with delayed graft acceptance. Fifty percent of tested recipient mice showed reactivity against donor cells in the cytotoxicity assay, which could be related to the prenatal sensitization. We conclude that microchimerism does not lead to the induction of a high degree tolerance after in utero transplantation and instead may lead to the development of alloreactivity to donor cells.