SEROLOGICAL REACTIVITY OF RECOMBINANT 1D AUTOANTIGEN AND ITS EXPRESSION IN HUMAN THYROID AND EYE MUSCLE-TISSUE - A POSSIBLE AUTOANTIGENIC LINK IN GRAVES PATIENTS

Thyroid-associated ophthalmopathy (TAO) is a potentially severe autoim mune disease, in and around the orbit, usually accompanied by Graves' disease. It was the goal of this study to develop a serological indica tor for TAO and to characterize its expression in human thyroid and ey e muscle tissue. Thus, we have recloned the full-length 1D-complementa ry DNA and assessed its expression levels in 90 healthy and diseased h uman thyroids. Only Graves' patients suffering from TAO (n = 29) displ ayed a significant, 2.1-fold increase of 1D expression levels (P = 0.0 29), compared with normal controls (n = 9), as assessed using the Mann -Whitney U-test for paired, nonnormally distributed samples. In contra st, a decrease of 1D expression (to 40% of control normal values) was confined to thyroid autonomy (n = 19, P = 0.032). In all other disease d human thyroids, including Graves' thyroids from patients not sufferi ng from clinically overt TAO (n = 9), 1D expression levels were not di fferent from the healthy controls. 1D gene expression was demonstrated in both healthy (n = 10) and diseased (n = 10) eye muscle tissues. Fu rthermore, a recombinant protein derived from baculovirus-infected Sf9 insect cells was purified under both nondenaturing and denaturing con ditions. While under nondenaturing conditions, the molecular mass of r ecombinant 1D was determined to be 85 kDa; denaturing isolation yielde d the expected 64-kDa protein. Autoantibodies against denatured 1D pro tein were not detectable in sera of diseased or healthy subjects. Immu noreactivity against the 85-kDa, nondenatured protein, evaluated in a panel of 222 different human sera, showed that 82% of Graves' patients suffering from TAO had autoantibodies against recombinant 1D, whereas only 5% of the healthy controls were positive for antibodies against 1D. Taken together, our results demonstrate a high disease sensitivity and specificity of recombinant, nondenatured 1D, to distinguish Grave s' disease with or without TAO from other forms of thyroid and/or eye disease. Prospective studies will have to show whether autoantibodies against 1D can also be used as a prognosticator of TAO.