Use of a cDNA microarray to determine molecular mechanisms involved in grey platelet syndrome

The grey platelet syndrome (GPS) is a bleeding disorder of unknown aetiology with phenotypic and genetic heterogeneity. Affected patients exhibit macrothrombocytopenia, decreased alpha-granule content and, sometimes, myelofibrosis. We used microarray technology to investigate changes in gene expression that might reveal mechanisms involved in GPS. The expression of 4900 unique genes and expressed sequence tags was evaluated in fibroblasts from a GPS patient; normal fibroblasts provided the reference standard. Genes that were differentially regulated in the GPS cells were categorized into gene clusters based upon similarity/differences of expression differences. The results showed that genes with functional similarities clustered together. This analysis revealed significant upregulation of selected biological processes involving the production of cytoskeleton proteins, including fibronectin 1, thrombospondins 1 and 2, and collagen VI alpha. These genes appear to play a role in the pathogenesis of GPS. Indeed, Northern blot analyses confirmed that fibronectin, thrombospondin and matrix metalloprotease-2 were overexpressed in GPS fibroblasts compared with normal fibroblasts. Moreover, immunohistochemistry studies revealed robust fibronectin staining in GPS fibroblasts compared with normal ones. Our findings support the feasibility of using cDNA microarray techniques to detect distinctive and informative differences in gene expression patterns relevant to GPS, and suggest that the molecular basis for myelofibrosis in GPS involves upregulation of cytoskeleton proteins.