NORMAL HUMAN DENDRITIC CELLS: A TOOL TO STUDY ALLERGIC AND IMMUNOLOGICAL REACTIONS IN VITRO.

Dendritic cells (DC) or Langerhans cells (LC) are immunological cells of the body which play a key role in allergic reactions and infectious diseases. However, widespread experimental use of DC has not occurred since previously-used methods of harvesting DC directly from tissue result in poor yields, short survival time, and rapid phenotypic changes. Currently, we have developed a new method of generating DC from CD34+ progenitor cells harvested from human umbilical cord blood. This method resulted in an average increase of 205 ± 86 fold (n = 15) in DC number versus previous methods. The DC express CD1a, HLA-DR, and co-stimulatory molecules (CD40 and CD80), and can be cultured for up to 36 days with no significant change in cell number or phenotype. Transmission electron microscopy showed the presence of Birbeck granules, a key ultrastructural marker of DC, over the duration of the culture period. The DC contain plasmacytoid (CD123+/cd11c-) and myeloid (CD11c+/cd123+) populations and can be frozen and recovered with a viability of 80-90%. Exposure of the DC to external stimuli such as lipopolysaccharide (LPS), phorbol-12-myristate-13-acetate (PMA), and interferon-γ (IFN-γ), caused a reproducible (n = 4), high level of gene and protein responsiveness in terms of IL-12, macrophage inflammatory protein (MIP)-1α, MIP-3α, IL-6, and TNF-α expression. DC pulsed with the neo-antigen, keyhole-limpet hemocyanin (KLH), or the recall-antigen, tetanus toxoid, induced autologous T-cell proliferation as measured by 5-bromo-2-deoxyuridine (BrdU) incorporation. Functionally, the DC were also shown to: 1) express foreign genes following transfection, 2) migrate in response to chemotactic factors such as granulocyte/monocyte colony stimulating factor (GM-SCF) and MIP-3α, 3) become infected following exposure to HIV-1. Thus, these cells are likely to be useful in a broad variety of allergic and immunological studies.

AIDS, Allergenicity, Antigen presentation, Antigen-presenting cells (APC), Birbeck granules, CD1a, CD209, CD34+ progenitor cells, Chemokine, Cytokine, DC-100-MM, Ecto-cervical, EpiDerm, EpiVaginal, HIV-1, HLA-DR, IL-12, IL-6, Immuno-therapy, Immunogenicity, Inflammatory, LC, Langerhans cells, MIP-3a, Macrophage inflammatory protein (MIP)-1a, Macrophage tropic (MT), 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)