技术资料

Umbilical cord blood (UCB) is increasingly being used for human hematopoietic stem cell (HSC) transplantation in children but often requires pooling multiple cords to obtain sufficient numbers for transplantation in adults. To overcome this limitation,we have used an ex vivo two-week culture system to expand the number of hematopoietic CD34(+) cells in cord blood. To assess the in vivo function of these expanded CD34(+) cells,cultured human UCB containing 1 x 10(6) CD34(+) cells were transplanted into conditioned NOD-scid IL2rgamma(null) mice. The expanded CD34(+) cells displayed short- and long-term repopulating cell activity. The cultured human cells differentiated into myeloid,B-lymphoid,and erythroid lineages,but not T lymphocytes. Administration of human recombinant TNFalpha to recipient mice immediately prior to transplantation promoted human thymocyte and T-cell development. These T cells proliferated vigorously in response to TCR cross-linking by anti-CD3 antibody. Engrafted TNFalpha-treated mice generated antibodies in response to T-dependent and T-independent immunization,which was enhanced when mice were co-treated with the B cell cytokine BLyS. Ex vivo expanded CD34(+) human UCB cells have the capacity to generate multiple hematopoietic lineages and a functional human immune system upon transplantation into TNFalpha-treated NOD-scid IL2rgamma(null) mice. View Publication

Two critical concerns in clinical cord blood transplantation are the initial time to engraftment and the subsequent restoration of immune function. These studies measured the impact of progenitor cell dose on both the pace and strength of hematopoietic reconstitution by transplanting nonobese diabetic/severe combined immunodeficiency/interleukin-2 receptor-gamma-null (NSγ) mice with lineage-depleted aldehyde dehydrogenase-bright CD34(+) human cord blood progenitors. The progress of each transplant was monitored over an extended time course by repeatedly analyzing the peripheral blood for human hematopoietic cells. In vivo human hematopoietic development was complete. After long-term transplantation assays (≥ 19 weeks),human T-cell development was documented within multiple tissues in 16 of 32 NSγ mice. Human T-cell differentiation was active within NSγ thymuses,as documented by the presence of CD4(+) CD8(+) T-cell progenitors as well as T-cell receptor excision circles. It is important to note that although myeloid and B-cell engraftment was detected as early as 4 weeks after transplantation,human T-cell development was exclusively late onset. High progenitor cell doses were associated with a robust human hematopoietic chimerism that accelerated both initial time to engraftment and subsequent T-cell development. At lower progenitor cell doses,the chimerism was weak and the human hematopoietic lineage development was frequently incomplete. View Publication