Contemporary studies have established that Langerhans cells (LC) are an immature subset of leukocytes closely related to lymphoid dendritic cells (DC). As resident antigen presenting cells (APC) in skin, LC play a key role in the induction of T cell-mediated immunity. In addition, ultraviolet B (UVB) radiation (290-320 nm wavelength), which causes skin cancer in humans, prevents LC from presenting epicutaneously applied haptens in an immunogenic fashion and depletes LC from the epidermis. An important goal in determining the mechanism by which UVB and other eternal agents produce these effects in skin is to identify the relevant target of that radiation. This question is the subjects of the applicant's currently funded NIH R01 research grant. Results from these studies will elucidate immunologic mechanism in contact dermatitis and in the destruction of skin cancer, leading to the development of novel therapies to treat dermatitis and to prevent the development of skin cancer in humans. Recently, new techniques of molecular biology have permitted the study of other epidermal that were cloned from the skin. Unfortunately, these techniques have been applied to LC with only modest success, primarily because of the unavailability of cell lines and clones. It has also been established recently that the related lymphoid DC will proliferate vigorously in culture when appropriate cytokines are provided, suggesting new strategies to establish lone-term LC lines and clones. The objective in the work supported by this Fogarty mechanism, is to develop long-term, stale lines or clones of LC, thereby allowing us to study UVB-induced effects more effectively. The current proposal will take advantage of the extensive expertise in tissue culture, cell phenotyping, and molecular biology within the host department.
The aims are: 1. To study growth factor requirements of LC. 2. To establish stable, long-term cell lines and clones that are derived from epidermal LC> 3. To characterize certain in vitro functional properties of the LC lines and clones. Methods selected are based on the hypothesis that relevant growth factors for LC are elaborated by KC. This hypothesis is supported by the knowledge that LC demonstrate mitotic activities in situ and that KC secrete a wide array of cytokines and growth factors. mRNAs detected in epidermal tissues (and in EC suspensions) included IL-1alpha, IL-1Beta, IL-3, IL-6, IL-7, IL-8, TNFALPHA, TNFBETA, GM-CSF, macrophage inflammatory protein-1alpha (MIP)-1alpha) and interferon-gamma (gammaIFN).
Specific Aim #1 is to identify and characterize soluble factor(s) produced by KC to promote the growth of LC. Knowledge gained in Aim 1 will be used to establish long-term LC lines. In addition, strategies other than the use of growth factors may also be employed. These include cell immortalization and transformation, accomplished with chemical carcinogens, viral transformation, gamma- radiation, or by the introduction of exogenous genes for growth factors and/or their receptors, or by transfection with viral DNAs that encode particular oncogenes or transcription factors.
Specific Aim #2 is to establish stable, long-term cell lines from mouse epidermal LC using currently available methodologies and materials.
In specific Aim #3 we will characterize the phenotypic and functional properties of the cell lines established in Aim #2. It should be noted that only the in vitro aspects of the Aim will be conducted in Vienna. No funds to support in vivo animal studies have been requested.
| Lenz, P; Elbe, A; Stingl, G et al. (1996) MHC class I expression on dendritic cells is sufficient to sensitize for transplantation immunity. J Invest Dermatol 107:844-8 |
