Report, FWF Project P16188, Immunosuppression by 4-amino tetrahydrobiopterin,
Ernst R. Werner, Division of Biological Chemistry, Biocenter,Medical University of Innsbruck
Our research focusses on the metabolic roles of pteridines. This class of substances comprises important vitamins such as folic acid and riboflavin, which are formed by bacteria and plants. Mammals have, however, the capacity to synthesize another pteridine, namely tetrahydrobiopterin. This compound is used as cofactor for specific hydroxylation reactions in our body, which are required for the biosynthesis of neurotransmitters, metabolism of essential amino acids and ether lipids, and the biosynthesis of nitric oxide, which in turn is important for neurotransmission, for the regulation of blood pressure and the defence against pathogens and tumours.
In the project 16188 we investigated pharmacological actions of derivatives of tetrahydrobiopterin, which we had initially developed as tools to investigate the biochemistry of stimulation of nitric oxide synthases by tetrahydrobiopterin. One of these tetrahydrobiopterin analogues, 4-amino tetrahydrobiopterin, is an inhibitor of nitric oxide synthases and shows immunosuppressive action in animal models of septic shock and of allograft rejection.
Our experimental approach was to study the action of tetrahydrobiopterin compounds on cultivated cells. We chose cells that constitute the immune systems host defence line. In a murine macrophage like cell line, we could demonstrate that tetrahydropterins down regulate gene expression and trigger programmed cells death. We succeeded in characterising the biochemical events leading to these effects: Tetrahydropterins in the medium convert oxygen to hydrogen peroxide. This triggers the activation of caspase 3, which in turn triggers programmed cell death and cleaves the P65 protein of the NF-kappa B transcription factor, which in turn causes the shutdown of gene expression.
We then studied T-cells and dendritic cells, both key cellular constituents of the immune system that cooperate in the regulation of the immune response. We isolated these cells from mice with and without a functional gene for inducible nitric oxide synthase. In contrast to the macrophage cell line, these cells are resistant to the actions of hydrogen peroxide. We found that 4-amino tetrahydrobiopterin specifically attenuated the expression of a protein on the surface of dendritic cells, that is essential for its interaction with T-cells. This effect occurred irrespective of the presence or absence of a functional gene for inducible nitric oxide synthase.
This novel immunosuppressive mechanism discovered in the project 16188 could possibly explain the immunosuppressive action of 4-amino tetrahydrobiopterin in animals, and may lead to the development of a new class of immunosuppressants.
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