2 research outputs found
A Genomic Point Mutation in the Extracellular Domain of the Thyrotropin Receptor in Patients with Graves’ Ophthalmopathy
Orbital and pretibial fibroblasts are targets of autoimmune attack in Graves' ophthalmopathy (GO) and pretibial dermopathy (PTD). The fibroblast autoantigen involved in these peripheral manifestations of Graves' disease and the reason for the association of GO and PTD with hyperthyroidism are unknown. RNA encoding the full-length extracellular domain of the TSH receptor has been demonstrated in orbital and dermal fibroblasts from patients with GO and normal subjects, suggesting a possible antigenic link between fibroblasts and thyrocytes. RNA was isolated from cultured orbital, pretibial, and abdominal fibroblasts obtained from patients with severe GO (n = 22) and normal subjects (n = 5). RNA was reverse transcribed, and the resulting cDNA was amplified by the polymerase chain reaction, using primers spanning overlapping regions of the entire extracellular domain of the TSH receptor. Nucleotide sequence analysis showed an A for C substitution in the first position of codon 52 in 2 of the patients, both of whom had GO, PTD, and acropachy. Genomic DNA isolated from the 2 affected patients, and not from an additional 12 normal subjects, revealed the codon 52 mutation by direct sequencing and AciI restriction enzyme digestions. In conclusion, we have demonstrated the presence of a genomic point mutation, leading to a threonine for proline amino acid shift in the predicted peptide, in the extracellular domain of the TSH receptor in two patients with severe GO, PTD, acropachy, and high thyroid-stimulating immunoglobulin levels. RNA encoding this mutant product was demonstrated in the fibroblasts of these patients. We suggest that the TSH receptor may be an important fibroblast autoantigen in GO and PTD, and that this mutant form of the receptor may have unique immunogenic properties
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Signaling “cross-talk” between TGF-β1 and ECM signals in chondrocytic cells
The objective of this investigation was to clarify how the integrin pathway modulates downstream effectors of the TGF-β1 pathway in chondrocytic cell signaling. The levels of Smad2 and Smad3 phosphorylation upon TGF-β1 or α2β1 integrin (Type II collagen) stimulation were analyzed by Western blotting techniques. Cellular response was determined by quantitation of procollagen gene expression.
Stimulation of cells with TGF-β1 and Type II collagen led to rapid phosphorylation of Smad2 and 3 with phosphorylation peaking between 15 min and 1 h. Combined stimulation led to a synergistic increase in the phosphorylation of Smad2 and Smad3. Type II collagen gene expression paralleled Smad phosphorylation.
Type II collagen modulates the TGF signaling cascade involving Smad2 and Smad3 leading to an increase in Type II collagen transcription. Therefore, we conclude that TGF-β1 and integrin stimuli interact prior to Smad2 and 3 phosphorylation in the cytoplasm of chondrocytic cells and regulates the expression of ECM components in chondrocytes