Identification of thyroid hormone target genes in human cells

Abstract

Human skin fibroblasts are readily accessible cells for propagation in culture without transformation that can serve for direct pathophysiology studies in subjects with inherited diseases. We therefore examined by quantitative fluorescent cDNA microarray analysis the effect of thyroid hormone (TH) on the expression of over 15,000 genes in fibroblasts of two normal individuals. Fibroblasts from two subjects with resistance to thyroid hormone (RTH) due to mutations in the TH receptor (TR) β gene were used to confirm the specificity of the hormonal effect by the ability to discriminate between normal cells and cells with a defect in TH action. Microarray analysis identified 148 genes induced by ≥1.4-fold and 5 genes repressed to ≤0.7 after 24h treatment with 2 nM T3. Taking into account duplicate genes, these represented 91 upregulated and 5 downregulated genes. Confirmation by real-time PCR was obtained in 8 of 10 induced and 2 of 3 repressed genes. Further evidence for T3 specific induction was provided by progressive changes in mRNA concentrations with graded doses of TH, which were absent in fibroblasts from the patients with RTH. We could, thus, identify several new TH responsive genes, both positively (AKR1C1–3, PFKP, RAB3B, COLVIA3, ENO1) and negatively (FGF7, ADH1B) regulated. These genes, as well as three known to be TH regulated in other species and found in this study also in human cells (GLUT1, SLC16A3 and BTEB1), have a variety of regulatory functions in development and metabolism. TH seems to induce these genes by initiating either genomic or nongenomic mechanisms. TH-mediated downregulation of FGF7 and ADH1B surprisingly persisted in fibroblasts from patients with RTH. This first systematic study of TH-mediated gene expression in normal human cells identifies several new TH-responsive genes and demonstrates that skin fibroblasts are suitable for the study of TH action in health and in disease.