Aspects of proteinase-activated receptor-2: A sensor of enzymatic activity at the cell surface

Detta är en avhandling från Div. of molecular neurobiology, Sölvegatan 17, SE-223 62 Lund, Sweden

Sammanfattning: Information about extracellular proteolytic activity is transduced to the cell by a transmembrane G-protein coupled receptor subfamily. These receptors are activated by proteolytic cleavage in the extracellular amino-terminal domain and are thus termed proteinase-activated receptors (PARs). The second member of this subfamily, PAR-2, is activated by trypsin-like proteinases, however the physiological activator of PAR-2 is still undefined. The work described in the present thesis was initiated with the cloning of a mouse PAR-2 cDNA and comparisons with genomic sequences showed that the PAR-2 gene consists of a minimum of two separate exons. In cultured endothelial cells, PAR-2 activation was shown to cause an increase in intracellular calcium levels and the release of prostaglandins. PAR-2 activation also caused a rapid and transient formation of tissue factor mRNA in the endothelial cells which was accompanied by an increased tissue factor activity, leading to a shortened coagulation time in a standard clotting assay. In order to identify and study potential activators of PAR-2, a method was developed to measure proteolytic activation of PARs. PAR-2 was tagged with insulin C-peptide that upon receptor cleavage is released and then quantified. It was shown that extrapancreatic trypsin-2 is a potent activator of PAR-2. The presence of PAR-2 was studied in human endometrium during the menstrual cycle using in situ hybridization, immunohistochemistry and Northern blot. PAR-2 was expressed in the wall of small blood vessels during the entire cycle and in the glandular epithelial cells during only the secretory phase indicating heterologous regulation of the PAR-2 gene expression in the different cell types and suggesting a role for PAR-2 in implantation and/or menstruation.

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