Umbilical Cord Blood Derived CD133+ Hematopoietic Stem Cells Are Defective Antigen Presenting Cells (APC), Lack Expression of Co-Stimulatory Receptors, and Generate TH₂ T-Lymphocyte Responses In Vitro.

Early clinical studies have demonstrated benefit in patients receiving cellular therapy utilizing bone marrow derived (BM)-CD133⁺ hematopoietic stem cells (HSC) for treatment of cardiovascular disease. Umbilical cord blood (UCB) is a potential source of CD133⁺ cells for use as an allogeneic cell source for therapeutic angiogenesis, however, the benefits must be weighed against potential adverse immunologic responses. We tested the hypothesis that allogeneic CD133⁺ cells derived from UCB may be defective as professional antigen-presenting-cells (APC) and thus may mediate TH₂ T-cell immune response. CD133⁺ cells were isolated from UCB mononuclear cells (MNC) by magnetic autoMACs bead selection (Miltenyi Biotech, Auburn CA) and analyzed for purity and surface expression of MHC and co-stimulatory antigens by flow cytometry. We have demonstrated induced immune reactivity by mixed lymphocyte reactions (MLR) using healthy adult peripheral blood MNC as responders stimulated by irradiated CD133⁺ from UCB and BM (ratio 3:1) with both ³H-thymidine and CFSE staining. Surface expression of both MHC class I (57.6±17.2%) and MHC class II (66.5±12.5%) are present on the majority of UCB CD133⁺ cells. To test the ability of CD133⁺ cells to function as APC, a modified MLR was performed. Briefly, isolated UCB CD133⁺ cells were cultured for 96h in the presence of adult peripheral blood (AB) derived-MNC or isolated CD3 AB T-cells as responders at a 3:1 ratio (responder:effector) in RPMI 1640 supplemented with 10% FBS and 1% L-glutamine. CD133⁺ cells induced proliferation in both non-selected AB MNC (21.7±6.4e3cpm) and selected AB T-cell (85.6±15.1e3 cpm) cultures as measured by ³H-thymidine incorporation. UCB CD133⁺ cells were noted to lack surface expression of co-stimulatory antigens including: CD40 (0.93±1.0%), CD80 (0.85±0.15%), and CD86 (0.88±0.64%). Because primary T-cell receptor stimulation in the absence of a co-stimulatory response is known to induce TH₂ responses we compared UCB MNC and CD133⁺ derived from the same unit as stimulators for responding HLA mismatched allogeneic AB MNC. Preliminary results demonstrate elevated levels of IL-4 and IL-10 produced by responder AB MNC stimulated by CD133⁺ cells as compared to UCB MNC, with measured increases of 964±538pg/mL (IL-10) and 141±45.7pg/mL (IL-4). In conclusion, though CD133⁺ cells derived from UCB express both MHC class I and II they lack surface expression of co-stimulatory receptors, and are defective as professional APC. Additionally, UCB CD133⁺ cells induce elevated levels of IL-4 and IL-10 by responding allogeneic adult MNC in modified mixed lymphocyte in vitro cultures. MNC stimulation by selected UCB CD133⁺ cells elicit elevated levels of tolerance-associated cytokines IL-4 and IL-10 as compared to UCB MNC used as stimulator cells. Taken together, UCB CD133 lack co-stimulatory receptor expression and elicit TH₂ T-lymphocyte responses, and may allow for immune tolerance responses when used as a source of allogeneic stem cells for therapeutic use in vasculogenesis.