Expression of tenascin-C splice variants by human skin cells

Tenascin-C is an extracellular matrix glycoprotein that is expressed in a s patially and temporally restricted pattern, Various functionally different tenascin-C isoforms can be expressed as a result of alternative splicing of the pre-mRNA. Previously we identified human epidermal keratinocytes as a source of tenascin-C in healing wounds, In this studs, we investigated whet her different tenascin-C transcripts are expressed by epidermal keratinocyt es and dermal fibroblasts, In addition, we compared expression of tenascin- C splice variants at the mRNA and protein levels in tissue samples of norma l and diseased skin. Northern blot analysis revealed two major tenascin-C m RNA transcripts of approximately 7500 and 5800 nucleotides in cultured epid ermal keratinocytes and fibroblasts, and in biopsies. Although both dermal fibroblasts and epidermal keratinocytes predominantly expressed the larger tenascin-C mRNA, epidermal keratinocytes expressed smaller transcripts at h igher levels than dermal fibroblasts. In keratinocytes the levels of the tw o mRNAs were differentially affected by inflammatory cytokines that increas ed tenascin-C expression in these cells, The addition of IFN gamma slightly increased the proportion of large transcripts, In contrast, TNF alpha favo ured expression of smaller tenascin-C transcripts, and IL-4 equally affecte d the expression of large and small tenascin-C mRNAs, To enable detection o f tenascin-C transcripts that are expressed at very low levels we amplified by polymerase chain reaction the fibronectin type III repeats whose expres sion is regulated by alternative splicing, In cDNA of cultured keratinocyte s and fibroblasts, and in skin biopsies, several tenascin-C transcripts cou ld be detected that corresponded to tenascin-C variants including different numbers of fibronectin type III repeats. Distribution of tenascin-C isofor ms at the protein level was studied immunohistochemically in healthy skin, wounds, psoriatic lesions and epidermal tumours and hyperplasia, No differe nces were observed in reactivity between an antibody that binds all tenasci n-C isoforms and antibodies that bind fibronectin type III repeats that can be spliced out from smaller tenascin-C isoforms, We conclude that the tena scin-C isoforms that are translated from transcripts that we identified at the mRNA level seem to be distributed similarly in the conditions investiga ted.