Induction of extracellular matrix gene expression in normal human keratinocytes by transforming growth factor β is altered by cellular differentiation

Abstract Changes in epithelial substrata have been related to the cellular capacity for proliferation and to changes in cellular behavior. The effect of TGFβ1 on the expression of the basement membrane genes, fibronectin, laminin B1, and collagen α1(IV), was examined. Northern analysis revealed that treatment of normal human epidermal keratinocytes with 100 p M TGFβ1 increased the expression of each extracellular matrix (ECM) gene within 4 h of treatment. Maximal induction was reached within 24 h after treatment. The induction of ECM mRNA expression was dose dependent and was observed at doses as low as 1–3 p M TGFβ1. Incremental doses of TGFβ1 also increased cellular levels of fibronectin protein in undifferentiated keratinocytes and resulted in increased secretion of fibronectin. Squamous-differentiated cultures of keratinocytes expressed lower levels of the extracellular matrix RNAs than did undifferentiated cells. Treatment of these differentiated cells with TGFβ1 induced the expression of fibronectin mRNA to levels seen in TGFβ-treated, undifferentiated keratinocytes but only marginally increased the expression of collagen α1(IV) and laminin B1 mRNA. The increased fibronectin mRNA expression in the differentiated keratinocytes was also reflected by increased accumulation of cellular and secreted fibronectin protein. The inclusion of cycloheximide in the protocol indicated that TGFβ induction of collagen α1(IV) mRNA was signaled by proteins already present in the cells but that TGFβ required the synthesis of a protein(s) to fully induce expression of fibronectin and laminin B1 mRNA. The differential regulation of these genes in differentiated cells may be important to TGFβ action in regulating reepithelialization.