Abstract
Efficient hematopoietic reconstitution of wild type mice requires preconditioning. Established experimental protocols exist to transplant hematopoietic stem cells into lethally irradiated or chemically myeloablated adult mice or unirradiated immunodeficient mice. We sought to develop a protocol to reconstitute immuno-replete neonatal mice. We describe irradiation and injection procedures for two-day old mice that lead to efficient long-term reconstitution of primary and secondary lymphoid organs. We demonstrate that the frequencies of lymphoid and myeloid cells in primary and secondary lymphoid organs are indistinguishable from unirradiated uninjected sex- and age-matched control animals by 5 weeks post-reconstitution. Thus, this system will facilitate studies aimed at understanding the developmental and environmental mechanisms that contribute to conditions that have a window of susceptibility during the perinatal period.
Highlights
In recent years, the importance of early life events on the subsequent development of human disease that may not manifest until late childhood or even adulthood has become increasingly appreciated[1]
For a brief time during the perinatal period the liver continues to be a source of extra-medullary hematopoiesis until eventually the bone marrow is fully established and takes over[20]
We reasoned that donor bone marrow injected into the liver of lethally irradiated newborn mice should be able to seed primary lymphoid organs and eventually populate the peripheral immune system of wild type mice
Summary
The importance of early life events on the subsequent development of human disease that may not manifest until late childhood or even adulthood has become increasingly appreciated[1]. Interactions between immune cells and nonhematopoietic cells are essential for maintaining overall tissue homeostasis, and disruption of this communication may have critical importance in the pathogenesis of disease. Generation of bone marrow chimeras allows mixing and matching of recipient and bone marrow donors so that the role of hematopoietic and non-hematopoietic components can be studied alone or in combination, and in mice with a fixed genetic background without the 3-year delay incurred by completing 20 backcrosses[4]. We sought to establish an experimental protocol that allows complete hematopoietic reconstitution of newborn immunocompetent mice so that the relative contribution of hematopoietic and nonhematopoietic lineages could be evaluated during the perinatal period using any wild type or genetically modified strain as the donor or host. Taking into account previous data on mortality, frequency of mice with detectable donor cells, and the frequency of donor cells among total CD45+, as well as practical considerations such as time, expense, special training, and side effects, we chose to optimize an irradiation regimen
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