Lymphoid tissue-specific cell production from hematopoietic progenitor cells in three-dimensional devices
A technology for hematopoietic progenitor cells and lymphoid tissue, which can be used in blood/immune system cells, measuring devices, tissue culture, etc., and can solve problems such as short half-life
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Embodiment 1
[0121] Example 1: Human cells produced in a co-culture system
[0122] Viability, immunophenotype and function of
[0123] The number of surviving cells produced in the co-culture system and their immunophenotypes are listed in Table 2. The greatest proliferation of human T cells was observed when human fetal thymic CD34+ cells and UCB CD34+ cells were co-cultured with murine thymic stroma cultured on Cellfoam. Data obtained from a direct comparison of CD34+ cell co-cultures on murine thymic stroma on Cellfoam and CD34+ cell co-cultures on murine stroma grown as simple monolayers are also presented in Table 2 .
[0124] T cells generated in co-culture systems have also been shown to be activated by T-tropic HIV-1 IIIB Infected, these cells were also transduced by the MFG vector with a transduction efficiency of 12-22% (n=3).
Embodiment 2
[0125] Example 2: Maintenance of immature progenitor cells
[0126] According to the present invention, it has also been found that cell foam cultures of thymic stromal cells can induce T cell differentiation of CD34+ progenitor cells while still retaining a partial composition of CD34- cells. Primate CD34+ progenitor cells were cultured on human or porcine thymus that had been established on Cellfoam tissue scaffolds. CD3+CD4+CD8+ triple positive cells and CD3+CD4+ and CD3+CD8+ double positive cells were reliably recovered after 14-21 days. Furthermore, after 14-21 days, CD3- cells were found to be composed in part with CD34+ progenitor cells. These CD34+ cells are not only CD3-, but many are also CD2+. This demonstrates that thymic cultures in Cellfoam tissue scaffolds can support T cell differentiation while preserving a long-lived CD34+ progenitor cell population. As will be apparent to those of ordinary skill in the art, this surprising finding demonstrates that it i...
Embodiment 3
[0127] Example 3: T cell function (proliferation / anergy) test
[0128] T cell function was assessed by proliferative potential to specific and nonspecific antigens using standard assays. Specifically, the test assessed T cell receptor response (TRR) mediated proliferation using an anti-CD3 antibody (Becton Dickinson), and baseline non-specific proliferation using concanavalin A (Con-A). Briefly, cells were washed with RPME containing 10% FCS and treated with a concentration of 10 6 cells / ml suspended in it. Add 100 μl (10 5 cells). 10 per culture dish in the presence of IL-2 (20 units / ml) and irradiated monocytes (MC) (100 ml RPMI containing 10% FCS). 5 cells), the cells were stimulated with Con-A (5 μg / ml) (non-specific reaction) or CD3 monoclonal antibody. For experimental conditions in which a monoclonal antibody to CD3 was used, purified goat anti-mouse F(ab') 2 Fragments (Kirkegard and Perry Laboratories, Gaithersberg, MD) were used as crosslinking reagents. Plat...
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