Characterization of nuclear thyroid hormone receptors of cultured skin fibroblasts from patients with resistance to thyroid hormone

Abstract

Nuclear thyroid hormone receptors of patients with the syndrome of resistance to thyroid hormone were investigated in cell lines from seven patients in four affected families and compared to results from six normals. Fibroblasts cultured from skin biopsies were used. When binding affinity and capacity for l-triiodothyronine (T 3) were examined by incubating whole cells or isolated nuclei, no significant differences were found. The amount of receptor released during the incubation of nuclei (9.3% to 19.0% of total nuclear receptors) was also within the normal range in these patients. When T 3 binding assays were performed on 0.3 mol/L KCl extracted receptor, a significant decrease in binding capacity (MBC) without a difference in binding affinity (Ka) was observed in four patients and a lower Ka with normal MBC was found in two patients. Recovery of receptors in saline extracts, from patients' fibroblasts showing a low MBC, was low in comparison to normals. Lability of salt extracted receptors at 38 °C was normal and salt extractability of T 3 occupied receptors, examined by incubation of [ 125I]-T 3 labeled nuclei with various concentrations of KCl, was only slightly decreased. This lower salt extractability of receptors was insufficient to account for the low MBC obtained by Scatchard analysis of T 3 binding to nuclear extracts. Gel filtration and density gradient sedimentation of salt-extracted receptors showed Stokes radius of 34 Å, and sedimentation coefficient of 3.4 S in all patients and normals. From these values, molecular weight of 49,000 and total frictional ratio ( f fo ) of 1.4 were calculated for nuclear receptors from patients and normals, suggesting a somewhat asymmetrical shape of receptors. Density gradient sedimentation patterns of micrococcal nuclease digested receptors, in which receptors were associated with 6.5, 12.5, and 17 S chromatin components, was also indistinguishable from normals. We conclude that in most patients with generalized thyroid hormone resistance, abnormalities of receptor function are evident after salt extraction of fibroblast nuclear receptors, but that the receptors have normal shape and thermolability. These abnormalities were consistent within but heterogeneous between sibships, suggesting heterogeneity of hereditary abnormalities in nuclear receptors in this disorder.