CHARACTERIZATION AND EVALUATION OF CD34+ GENERATED LANGERHANS CELLS FOR TISSUE ENGINEERING.

Specialized antigen-presenting cells (APC), particularly Langerhans cells (LC) residing in the skin, mucosa, and lymphoid tissues play a key role in sensitization and other immunological reactions of the body. However, difficulty in harvesting, short survival time in culture, and variability in cytokine production of cultured LC has prevented researchers from widespread use of LC for allergenicity and immunogenicity studies. Likewise, these problems have hampered the development of in vitro tissue models containing LC. Recently, we have developed a new culture method to generate large numbers of LC from umbilical cord blood (UCB). UCB derived CD34+ progenitor cells are expanded to produce up to 200 million LC per 60 ml UCB sample. The generated LC express CD1a, HLA-DR, CD209 (DC-SIGN), and CD206 (mannose receptor), all characteristic of LC. Transmission electron microscopy shows the presence of Birbeck granules, a key ultrastructural marker of LC. The lifespan of these cells has been increased to 41 days with very little change in surface marker expression or phenotype. Upon stimulation with lipopolysaccharide (LPS) and phorbol-12-myristate-13-acetate (PMA), the LC show a reproducible, high level protein release for IL-12, MIP-1a, MIP-3a, IL-6, RANTES, and IL-6. In addition, recent experiments demonstrate that these cells can be incorporated into MatTek’s reconstructed skin model, EpiDerm™, and its ectocervical-vaginal tissue model, EpiVaginal™. LC within these tissue constructs are detected by immunostaining for HLA DR antigens. These new tissue models and the isolated LC alone will likely be useful in a variety of studies related to: 1) allergenicity, 2) microbial infection, 3) neutralizing antibodies, 4) antigen presentation, and 5) immuno-therapy, amongst others.

AIDS, Allergenicity, Antigen presentation, Antigen-presenting cells (APC), Birbeck granules, CD1a, CD209, CD34+ progenitor cells, Chemokine, Cytokine, DC-100-MM, DC-SIGN, Ecto-cervical, EpiDerm, EpiVaginal, HIV-1, HLA-DR, IL-12, IL-6, Immuno-therapy, Immunogenicity, LC, Langerhans cells, Macrophage tropic (MT), Mannose receptor, Microbial infection, Microbicidal blocking, Neutralizing antibodies, T-cell tropic (TCT), Transmission electron microscopy, Vaginal-ectocervical tissue models, Virucidal blocking

HIV-1 virus stocks, Lipopolysaccharide (LPS), Phorbol-12-myristate-13-acetate (PMA)