Biomedical subjects

J R Wall

Publications and source records attributed to J R Wall.

At least 37 records · Page 2Linked to original sources

PubMed1998

Community-based training after acquired brain injury: preliminary findings.

Persons entering or returning to the workforce after brain injury often require assistance to identify, locate, secure and maintain employment. This is especially true for workers from economically disadvantaged environments who may have limited pre-injury work experience. Supported employment, the use of natural supports, and community-based approaches are helpful for the process of identifying, securing and maintaining work. However, reports are limited as to how these approaches are helpful for economically disadvantaged persons, especially in identifying meaningful work. An approach is described here which combines work adjustment training and supported employment; which is helpful in identifying consumer interest, and in improving position attainment, job retention and work satisfaction. A sample of persons (n = 38) with brain injury were contacted after completing the programme. A total of 59% of these persons remained employed at follow-up, compared to 32% at time of injury. Additional outcome data are presented to describe the usefulness of this approach.

Adult↗

PubMed1998

Natural killer cell activity in patients with Graves' disease and Hashimoto's thyroiditis.

Natural killer (NK) cell activity of peripheral blood lymphocytes (PBL) against k562 human tumor cell targets was studied in patients with Graves' disease and Hashimoto's thyroiditis. NK activity was measured in a standard 4-hour 51chromium (Cr) release assay. Cytotoxicity was expressed as lytic units (LU)/10(6) PBL. Significantly decreased NK cell activity was demonstrated in both groups of patients, with mean (+/- SE) lytic units of 10.3 (+/- 9.1) and 13.3 (+/- 10.3) for patients with Graves' disease and Hashimoto's thyroiditis, respectively, compared with 36.0 (+/- 26.3) for age- and sex-matched normal subjects. When patients with Graves' disease were analyzed according to their thyroid status; NK activity was significantly depressed in (1) hyperthyroid patients before treatment; (2) hyperthyroid patients receiving antithyroid therapy; and (3) euthyroid patients receiving antithyroid therapy, compared with normal subjects. Graves' disease patients who were hypothyroid after radioactive iodine therapy or thyroidectomy had normal NK activity. No significant differences between hyperthyroid and euthyroid patients or between hypothyroid patients and normal subjects were demonstrated. NK activity in patients with Graves' disease did not correlate with serum levels of thyroxine, the presence or severity of ophthalmopathy, or titers of serum thyroid antibodies. In patients with Hashimoto's thyroiditis there was no correlation between NK activity and goiter size, titers of antithyroid antibodies, or thyroid status. These findings suggest that depression of NK activity in both disorders is secondary to abnormalities of thyroid hormone secretion, although an effect of the underlying autoimmune reactions has not been excluded.

Adolescent↗

PubMed1998

Thyroid-associated ophthalmopathy in black South African patients with Graves' disease: relationship to antiflavoprotein antibodies.

Thyroid-associated ophthalmopathy (TAO) is a progressive eye disorder associated with Graves' hyperthyroidism, which is generally considered to have an autoimmune etiology. Eye muscle membrane proteins of 64 kd are good markers of ophthalmopathy in patients with thyroid autoimmunity. The 64-kd protein is now shown from a partial sequence to be the flavoprotein subunit (Fp) of mitochondrial succinate dehydrogenase. Hyperthyroidism due to Graves' disease is increasing in incidence among urban black female Africans, possibly because of exposure to environmental risk factors such as increased dietary iodine ingestion and stress. Ophthalmopathy is frequently observed in this clinical context, but its association with serum autoantibodies reactive with Fp has not been examined. We studied 19 black South African patients with Graves' disease during the course of prolonged antithyroid drug administration, of whom 10 had congestive ophthalmopathy, but no clinical evidence for eye muscle damage at the onset. Anti-Fp antibodies were detected in 2 of these patients, as well as in 2 of the 9 patients who did not have overt eye disease. Additionally, the antibodies became positive in 3 patients with ophthalmopathy in whom tests were negative initially, remained positive in 1 patient throughout the study period and became negative in 1 patient with positive tests initially. Ophthalmopathy did not develop in any of the 9 patients who lacked this complication on presentation. The reasons why we failed to demonstrate a close relationship between anti-Fp antibodies and the eye muscle component of ophthalmopathy are unclear although one possibility is that ocular myopathy is an uncommon manifestation in African thyrotoxic patients compared with those of Caucasian origin. The relationship between anti-Fp antibodies and eye muscle inflammation in patients with thyroid autoimmunity of different ethnic origins and environmental settings, needs to be addressed in a large prospective study.

Adult↗

PubMed1998

Role of the eye muscles in thyroid eye disease: identification of the principal autoantigens.

Thyroid-associated ophthalmopathy (TAO) is a progressive orbital disorder associated with Graves' hyperthyroidism and, less often, Hashimoto's thyroiditis in which autoantibodies react with orbital antigens and lead to exophthalmos and eye muscle inflammation. Eye muscle (EM) membrane proteins initially reported as 55 and 64 kd are the best markers of ophthalmopathy. The "64-kd protein" is now shown to be the flavoprotein subunit of mitochondrial succinate dehydrogenase and to have a correct molecular weight of 67 kd. We have cloned a fragment of a novel eye muscle protein, which we call G2s, and sequenced 1.4 kb of the full length cDNA. G2s does not share any significant homologies with other reported proteins. The 5.9 kb G2s mRNA, that corresponds to a protein of approximately 220 kd, is expressed in EM, other skeletal muscle and thyroid, but not in other tissues tested. We have also cloned and sequenced a 63-kd eye muscle protein identified as the calcium binding protein calsequestrin. Antibodies against calsequestrin were found in 40% of patients with active ophthalmopathy, but in 0% of normal subjects. Finally, we have sequenced a 19 amino acid fragment of a 55-kd porcine eye muscle membrane protein that exactly matched porcine and human sarcalumenin, a 160-kd glycoprotein localized in the lumen of the longitudinal sarcoplasmic reticulum of the skeletal muscle fiber where it binds calcium. A 53-kd glycoprotein fragment of the molecule corresponds to the 55-kd protein. In a preliminary study, serum antibodies against purified sarcalumenin were detected in 40% of patients with active TAO of less than 1 year duration, but in no controls tested. We porpose that the primary autoantigen in TAO is G2s, which would also explain the association of ophthalmopathy with thyroid autoimmunity, and that antibodies against the intracellular proteins flavoprotein, calsequestrin, and sarcalumenin are secondary markers of an immune-mediated reaction in eye muscle in patients with thyroid autoimmunity.

Autoantigens↗

PubMed1998

T cell interactions with extracellular matrix proteins in patients with thyroid-associated ophthalmopathy.

Although thyroid-associated ophthalmopathy (TAO) is now generally accepted as an autoimmune inflammatory disorder of the extraocular muscles and the orbital connective tissue, its aetiopathogenesis remains poorly understood. Recent data indicate that impaired interactions between T cells and extracellular matrix (ECM) proteins may play an important role in development and maintaining of an inflammatory process. We report here results of the study focusing on interactions between T lymphocytes and collagen-I (Coll-I), collagen-IV (Coll-IV), fibronectin (FN), laminin (LM) in patients with TAO. Using a standard peripheral blood mononuclear cells (PBMC) proliferation assay, we observed a markedly enhanced T cell response to Coll-I in patients with active TAO (mean SI=4.5). The proliferatory response to Coll-I was significantly greater (Wilcoxon test; p < 0.001) than in normal subjects (mean SI=1.88), patients with stable TAO (mean SI=2.05) and patients with thyroid autoimmune diseases (AITD) without ophthalmopathy (mean SI=2.49). PBMC stimulation by Coll-I is likely to be antigen-dependent requiring engagement of the T cell receptor with collagen peptides, rather than mediated via integrins. The percentage of circulating CD29+ (beta1 integrin chain) T cells was not increased in patients with active TAO. Additionally in the assay of costimulation of CD3-mediated proliferation, we found that peripheral blood T cells from patients with TAO and AITD were costimulated only by FN. On the other hand a markedly enhanced costimulation of CD3-mediated proliferative responses by Coll-I, Coll-IV, FN and LM were observed in a retrobulbar T cell line. We conclude that abnormalities in T cell interactions with ECM proteins, especially Coll-I may play a role in the pathogenesis of TAO.

Adolescent↗

PubMed1997

Antibodies against striated muscle, connective tissue and nuclear antigens in patients with thyroid-associated ophthalmopathy: should Graves' disease be considered a collagen disorder?

The identity and subcellular localization of the principal extraocular muscle (EOM) antigens and prevalences of the corresponding serum autoantibodies in thyroid-associated ophthalmopathy (TAO) need to be clarified. We have used porcine eye muscle tissue, which expresses all autoantigens identified in human tissue, as substrate in an indirect immunofluorescence assay. Several different patterns of antibody binding to EOM tissue antigens were observed with sera from patients with TAO namely, membrane, cytoplasmic, interstitial (endomysial) and nuclear. Overall, sera from 75% of patients with TAO contained one or more antibodies reactive with EOM, compared to 32% of patients with Graves' hyperthyroidism, 38% with Hashimoto's thyroiditis, and 16% of normals. All sera which reacted with EOM membrane or cytoplasmic antigens also reacted with the same antigen(s) in other skeletal muscle, but not in the other tissues tested. Sera from 31% of patients with TAO, but only 7% of those with Hashimoto's thyroiditis, and no patient with Graves' hyperthyroidism without evident ophthalmopathy, contained antinuclear antibodies (ANA). The most common nuclear fluorescence pattern was the finely speckled type typically associated with anti-Sm or anti-RNP antibodies. Significant positive correlations in patients with TAO were found between (i) EOM dysfunction and ANA (ii) eye disease of < 1 yr duration and EOM membrane-reactive antibodies and (iii) eye disease of < 1 yr duration and interstitial (endomysial) tissue-reactive antibodies. Although patients with Graves' disease do not usually exhibit other signs or immunologic features of a generalized collagen disorder, the finding of high prevalences of ANA and anti-striated muscle antibodies and, less often, anti-connective tissue antibodies in patients with ophthalmopathy, is consistent with it being a collagen-like disorder of the striated muscle, connective tissue and the thyroid. The reason why the inflammatory process is mainly limited to these tissues is unclear although cross reaction of ANA with tissue specific proteins or increased expression of muscle and connective tissue antigens in the orbit and skin, are possibilities.

Adult↗

PubMed1997

Hodgkin's disease treated with neck radiation is associated with increased antibody-dependent cellular cytotoxicity against human extraocular muscle cells.

Patients with Hodgkin's disease have higher a prevalence of thyroid function abnormalities and, perhaps, orbitopathy than the general population, but the pathophysiology of this association and its relationship to Hodgkin's disease treatment remain unclear. We analyzed the frequency of thyroid function abnormalities, autoantibodies against thyroid antigens, and autoimmunity against extraocular muscle cell antigens by Western blot analyses and antibody-dependent cellular cytotoxicity (ADCC) assays in patients with Hodgkin's disease (n = 20) and controls (n = 10). Hodgkin's disease patients were subdivided into those treated with thyroidal external beam radiation therapy (XRT, n = 15) or chemotherapy (MOPP/ABVD, n = 5). The ADCC assay against extraocular muscle cells was increased in patients with Hodgkin's disease (5.5% vs. <1.0%, p = .026) when compared with controls. In addition, Hodgkin's disease patients treated with XRT (with or without chemotherapy) had significantly higher ADCC tests than controls (9.7% vs. <1.0%, p = .010), In contrast, ADCC assays were not different between Hodgkin's disease patients treated with chemotherapy alone and controls (<1.0% vs. <1.0%, p = .53). Hodgkin's patients treated with XRT had higher ADCC assays than those treated with chemotherapy alone (p = .087), although this difference did not achieve statistical significance. Serum measurements of antithyroid peroxidase (TPO) antibodies, antithyroglobulin (Tg) antibodies, thyroid binding inhibitory immunoglobulins (TBII), and thyroid stimulating immunoglobulin (TSI) were similar in all groups. Antibodies against the 64 kDa orbital antigen were detected in 1 patient and 1 control subject. Excluding patients already treated with L-thyroxine for hypothyroidism (n = 5), free T3, but not free T4, was lower in the Hodgkin's disease group than in controls (2.2 pg/mL vs. 2.7 pg/mL, p = .008). Thyrotropin (TSH) concentrations were not statistically different between these groups. In summary, these data show: (1) ADCC against human orbital muscle cells is increased in patients with Hodgkin's disease compared with controls: (2) these differences were noted among Hodgkin's disease patients treated with thyroidal XRT, with or without chemotherapy, and not among those patients treated with chemotherapy alone; and (3) no statistically significant differences in the frequency of thyroid autoantibodies were found. These data suggest that patients with Hodgkin's disease display altered antibody-dependent immune function toward extraocular muscle cells that may possibly be related to by XRT. Larger, prospective studies assessing thyroid and orbital-related immunologic abnormalities in Hodgkin's disease are warranted.

Adult↗

PubMed1997

The study of visual evoked potentials in patients with thyroid-associated ophthalmopathy identifies asymptomatic optic nerve involvement.

In the present study we have recorded visual evoked cortical potentials (VECP) in 88 patients affected by autoimmune thyroid disease and thyroid-associated ophthalmopathy (TAO) without clinical signs of optic neuropathy. At the time of ophthalmological examination, 37 of these patients were hyperthyroid, 41 were euthyroid, and 8 were hypothyroid; 2 were not assessed. Twenty-nine normal subjects served as controls. We performed pattern reversal visual stimulation and recorded the amplitude and latency of the cortical electric response at 100 ms (P100 wave). There were no differences in the mean P100 amplitude of TAO patients and normal subjects. The mean P100 latency in patients was 105.6 +/- 0.5 ms, significantly higher than that in normal subjects (102.0 +/- 0.5 ms; P < 0.00003). Latency in euthyroid patients did not differ from that in either hypo- or hyperthyroid patients. The VECP test was positive (latency, > or = 110.0 ms) in 21 (23.8%) TAO patients. In patients with proptosis greater than 21 mm, latency was 106.7 +/- 0.7 ms, significantly higher than that in patients with normal Hertel measurements (104.3 +/- 0.6 ms; P < 0.01). Latency was not increased in patients with acute inflammatory signs compared to those with inactive eye disease and in patients with altered extrinsic motility. In patients with an abnormal visual field study, the mean latency was 110.3 +/- 1.5 ms, significantly higher than that in patients with a normal visual field (104.7 +/- 0.4; by t test, P < 0.000003). In conclusion, we observed a prolongation of the latency of the evoked cortical response in patients with TAO without subjective visual complaints and without optic nerve compression. We believe that the study of VECP in TAO is complementary to the study of the visual field in identifying early optic nerve dysfunction in the absence of decreased visual acuity.

Adolescent↗

PubMed1996

Tissue distribution and quantitation of a gene expressing a 64-kDa antigen associated with thyroid-associated ophthalmopathy.

We have developed a competitive reverse transcriptase-polymerase chain reaction (RT-PCR) assay for the quantitation of 1D mRNA, which encodes a 64-kDa protein associated with thyroid-associated ophthalmopathy, in preparations of total RNA from a variety of human tissues. This competitive RT-PCR assay is based on coamplification of competing 1D cDNA internal control template (pGEM-1D') and target cDNA template. 1D-specific mRNA was quantified in 10 human tissues. The level of 1D transcript expression in these tissues decreased in the following order: thyroid (5 pg/mg total RNA) > eye muscle (3.2 pg/mg) > skeletal muscle (2.4 pg/mg) > ovary (2 pg/mg) > cerebellum (0.4 pg/mg), kidney (0.33 pg/mg), pancreas (0.27 pg/mg), spleen (0.22 pg/mg), and thymus (0.19 pg/microgram) > retina (0.016 pg/mg). Graves' disease may be a multisystem autoimmune disorder of the connective tissue and skeletal muscle (and thyroid) that is mainly localized in the orbit and skin. One mechanism for this localization may be increased expression of target autoantigens, of which the 64-kDa protein 1D is a candidate, in the involved tissues. The higher expression of the 1D molecule in thyroid and eye supports this hypothesis.

Animals↗

PubMed1996

Antibody-dependent cell-mediated cytotoxicity against orbital target cells in thyroid-associated ophthalmopathy and related disorders; close relationship between serum cytotoxic antibodies and parameters of eye muscle dysfunction.

We have carried out tests for antibody-dependent cell-mediated cytotoxicity (ADCC) against extra ocular muscle (EOM), Müller's muscle, orbital fibroblasts and skeletal muscle in patients with thyroid-associated ophthalmopathy (TAO) and related eye disorders. Cytotoxicity was measured as lactate dehydrogenase (LDH) release and results expressed as % cytotoxicity. Tests were positive, with EOM cells, in 65% of patients with TAO, 75% with ocular myopathy, a variant of TAO in which periorbital inflammation is minimal, 50% with euthyroid Graves' disease defined as ophthalmopathy associated with subclinical thyroiditis and in 50% of patients with stable lid lag and retraction but no other signs of progressive ophthalmopathy, but in only 13% of patients with Graves' hyperthyroidism without ophthalmopathy, 10% with Hashimoto's thyroiditis and 14% of patients with other thyroid disorders. Tests were positive, with Müller's muscle cells, in 40% of patients with TAO, 25% with ocular myopathy, 40% with euthyroid Graves' disease, 44% with lid lag, 19% with Graves'hyperthyroidism, 50% with Hashimoto's thyroiditis and in 37.5% of patients with other thyroid disorders. When skeletal muscle cells were used as target, tests were positive in 13% of patients with TAO, 31% with lid lag, 25% with Graves' hyperthyroidism and in 29% of patients with Hashimoto's thyroiditis, but in no patient with euthyroid Graves' disease or other thyroid disorders. Tests were negative in all patients and normals tested when EOM-derived fibroblasts were used as targets in ADCC. A significant positive correlation between % cytotoxicity against EOM cells and the severity of the eye muscle dysfunction expressed as an eye muscle index, was observed in patients with TAO. There was a significant negative correlation between the duration of eye disease and % cytotoxicity against EOM cells, suggesting higher titers of cytotoxic antibodies in the early stages of TAO. There was no correlation between % cytotoxicity and serum level of anti-TSH receptor antibodies, measured in a radioreceptor assay. These findings suggest that autoimmunity against Müller's muscle may play a role in the pathogenesis of persistent lid lag and retraction. The nature of the EOM and Müller's muscle autoantigens recognized by cytotoxic antibodies in the serum of patients with TAO and related eye disorders is unknown.

Adult↗

PubMed1996

Failure to demonstrate cell-mediated immunity to orbital tissue antigens and epitopic fragments of a 64 kDa protein in the majority of patients with thyroid-associated ophthalmopathy.

We have studied a possible role of T cell sensitization to eye muscle antigens in patients with thyroid-associated ophthalmology (TAO). Peripheral blood mononuclear cell (PBMC) proliferation in response to crude porcine orbital tissue antigens, partially purified porcine eye muscle membrane proteins and predicted epitopic fragments of the recombinant 64 kDa protein 1D, was determined in patients with TAO and thyroid autoimmunity without eye disease. When membrane and cytosol fractions were used as antigen PBMC from 43% of patients with TAO but only 12.5% of normal subjects were responsive to a crude orbital connective tissue membrane fraction, although this difference was not significant. We were unable to demonstrate specific recognition of partially purified eye muscle membrane fractions; although most of the fractions tested were occasionally recognized by T cells from patients with ophthalmopathy, this was also the case for patients with autoimmune thyroid disease without ophthalmopathy and normal subjects. We did not clearly identify epitopic sequences within the 1D protein, most of the predicted peptides tested being recognized not only by T cells from a small proportion of patients with TAO, but also by those from some patients with autoimmune thyroid disease without ophthalmopathy and normal subjects. It is noteworthy however that approximately 22% of TAO patients, but no normal subjects, were positive to one or more of three peptides, suggesting that reactivity to the 1D protein may play a role in the pathogenesis of the eye disorder in some patients with TAO. The inconsistent and generally low T cell responses to crude and purified antigens noted in a few patients with TAO could be explained by low numbers of specifically sensitized lymphocytes in peripheral blood.

Adult↗

PubMed1996

Orbital tissue-derived T lymphocytes from patients with Graves' ophthalmopathy recognize autologous orbital antigens.

Lymphocytic and other mononuclear cell infiltration of the retro-bulbar space is observed in Graves' ophthalmopathy (GO). We investigated the antigenic character of orbital adipose/connective tissue and muscle from 21 euthyroid patients with severe GO after orbital surgery. Orbital tissue proteins were separated and recovered in soluble form by means of an electroelution technique. Twenty-two protein fractions, identified according to their molecular mass ranges, were used as antigens for orbital tissue-derived and peripheral blood T lymphocytes. Seventeen T cell lines from 6 patients were established from in vivo activated orbital T cells using interleukin-2 and anti-CD3 antibodies. T cell proliferation was measured as [3H] thymidine uptake. When screened for their reactivity to autologous adipose/connective tissue proteins, all T cell lines responded significantly to protein fractions 6-10 kDa [stimulation index (SI) = 32.9 +/- 9.8 (mean +/- SE)] and 19-26 kDa (17 +/- 5), but not to tuberculin, which was used as a control. Phenotypic analysis analysis of 10 orbital T cell lines indicated that 6 lines consisted predominantly of CD4+ cells. Incubation of a representative T cell line with allogeneic orbital protein fraction induced a very low response to protein fraction 19-26 kDa, but not to other fractions. Thyroid protein fraction 6-10 kDa also induced the proliferation of orbital T cell lines. Incubation of peripheral blood mononuclear cells with autologous orbital protein fractions gave similar results; positive responses to 6-10 and 19-26 kDa fractions were observed with orbital tissue from 12 of 14 patients (mean SI = 22 +/- 5.9 and 6.3 +/- 1.7, respectively), and positive responses were observed with orbital tissue from 3 of 4 patients to eye muscle fractions 6-10 and 19-26 kDa (13.8 +/- 6.9 and 6 +/- 2, respectively). When proteins from cultured orbital fibroblasts were used as antigens, autologous peripheral blood mononuclear cells from the 7 of the 9 patients tested responded to these 2 fractions (15.2 +/- 6.9 and 6.8 +/- 2.4, respectively), whereas a response to cultured orbital myoblasts was observed with the 19-26 kDa fraction only (SI = 8). Positive responses to abdominal adipose or muscle proteins, as controls, were not found. The demonstration of sensitized, orbital tissue-specific, T lymphocytes in the peripheral blood and orbit from patients with GO provides evidence for a role of cellular immunity in the pathogenesis of this eye disorder.

Adult↗

PubMed1995

Upper eyelid retraction in the absence of other evidence for progressive ophthalmopathy is associated with eye muscle autoantibodies.

We have studied 25 clinically euthyroid patients with eyelid lag and retraction referred to thyroid/eye clinic for clinical and orbital imaging evidence of extraocular eye muscle (EM) involvement, evidence of progressive ophthalmopathy and serum antibodies reactive with EM membrane antigens in immunoblotting. Fourteen patients had Graves' hyperthyroidism, 5 had Hashimoto's thyroiditis, and 6 had euthyroid Graves' disease. By carrying out orbital imaging we showed EM abnormalities in 10 of 23 patients (43%). Serum antibodies reactive with EM membrane antigens were detected in 96% of patients. Antibodies reactive with a 64-kDa antigen were detected in 66% of patients, while those reactive with 35-, 55-, and 95-kDa antigens were found in 21, 33, and 25% of patients, respectively. Antibody prevalences compared to normals were significantly different (P < 0.005) only for the 64-kDa protein. The prevalence and the degree of reactivity of 64-kDa antibodies were significantly different in patients with abnormal EM compared to those with normal EM at orbital imaging (P < 0.04 and P < 0.01, respectively). The results of this work suggest that in some patients inflammation of the eyelid muscles may be an isolated feature of ophthalmopathy and remains as the only sign of a "subclinical" eye disease in patients with thyroid autoimmunity.

Adult↗

PubMed1995

Identification of antigenic epitopes of 1D antigen recognized by antibodies in the serum of patients with thyroid-associated ophthalmopathy.

Random fragments of full-length 1D cDNA encoding the 64-kDa antigen were cloned and expressed in Escherichia coli as fusion proteins. Antigenic peptides were detected by colony blotting with sera from patients with thyroid-associated ophthalmopathy (TAO) and corresponding cDNA inserts were sequenced. Four antigenic peptides were characterized and positioned with respect to the 1D amino acid sequence. One antigenic peptide (G-8) was found to be close to the C-terminal end of 1D, and three others (G-2, G-4, and G-5) were clustered at sites between amino acids 76 and 205. The immunoreactivities of a panel of sera were tested against full-length 1D and the four peptides. A panel consisting of sera from (1) 15 patients with TAO, (2) 9 patients with Graves' hyperthyroidism without ophthalmopathy, (3) 6 patients with Hashimoto's thyroiditis without ophthalmopathy, and (4) 12 normal subjects were tested against the four peptides and full-length 1D protein by Western blotting. Sera from patients with TAO also reacted more often with G-4 and/or G-5 peptides than did those from normal subjects [7 of 15 (47%) sera from TAO patients versus 1 of 12 (8%) sera from normal subjects; P < 0.05]. Based on positioning of peptides onto the full-length 1D sequence and the reaction pattern of the serum panel, six immunogenic epitopes were identified on the 1D molecule.

Adult↗

PubMed1995

Should Graves' disease be considered a collagen disorder of the thyroid, skeletal muscle and connective tissue?

Graves' disease comprises hyperthyroidism, ophthalmopathy, pretibial myxedema and acropachy, which occur separately or in various combinations. We have used the indirect immunofluorescence test to investigate reactivity of sera from patients with autoimmune thyroid disorders with and without ophthalmopathy, with porcine extra ocular muscle (EOM) and control tissue substrates. Sera from 75% of patients with Graves' hyperthyroidism (GH) and ophthalmopathy, which we call thyroid-associated opthalmopathy (TAO), contained one or more antibodies reactive with EOM compared to 32% of those with GH without the eye disorder, 41% of patients with Hashimoto's thyroiditis (HT), and 16% of normals. Antibodies reactive with an EOM connective tissue antigen(s), seen as fluorescence of the interstitium and endomysium, were found in sera from 10% of patients with TAO and 16% of those with GH, but not from any patient with HT or normal subject. Similar patterns of connective tissue reactivity were also found in lacrimal gland, skeletal muscle, kidney and salivary gland. Antinuclear antibodies were detected in sera from 31% of patients with TAO, but from only 8% with HT, in no patient with GH and in only 3% of normal subjects. The most common pattern was a fine speckled fluorescence, found in 45% of sera, consistent with reactivity against the Sm antigen or nuclear RNP. The finding of a high prevalence of ANA and, less often, anti-connective tissue antibodies in patients with thyroid autoimmunity and ophthalmopathy, is consistent with Graves' disease being a "collagen-like disorder". The reason why inflammation and resulting tissue damage is limited to the thyroid, connective tissue of the skin and orbit, skeletal muscle and, possibly, the lacrimal gland, is unclear. One possibility is cross reaction of ANA with tissue specific membrane proteins in these sites. The extent of immunologic abnormalities, and the resulting clinical features, in patients with Graves' disease may reflect the severity of a putative defect in immune regulation.

Antibodies, Antinuclear↗

PubMed1995

New assays for the measurement of serum antibodies reactive with eye muscle membrane antigens confirm their significance in thyroid-associated ophthalmopathy.

Although sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting are widely used to detect serum antibodies in patients with autoimmune disorders, this procedure unfolds and denatures proteins and may alter antibody binding sites. We have used nondenaturing methods for the purification of a 64-kDa eye muscle (EM) membrane antigen associated with thyroid-associated ophthalmopathy (TAO). Pig EM membrane proteins were prepared from crude homogenates by high-speed centrifugation and solubilized by hand homogenization. The 64-kDa protein was further purified by isoelectric focusing performed in the absence of SDS, detergents, reducing agents, and urea. Sera from patients with active TAO of recent onset and thyroid autoimmunity without ophthalmopathy were tested for reactivity against purified native 64-kDa protein in immunoblotting. Tests were positive in 64% of patients with TAO, in 37.5% of those with Graves' hyperthyroidism without eye disease, in 11% of patients with Hashimoto's thyroiditis without eye disease, and in 13% of normal subjects. Many of the same sera were also tested for cytotoxic activity against human EM cells in an antibody-dependent cell-mediated cytotoxicity (ADCC) assay. ADCC tests were positive in 69% of patients with TAO but in no normal subject. The specificity and sensitivity of these two tests in TAO surpass those for all other published results for orbital tissue reactive autoantibodies. Although there was a tendency for a relationship between reactivity to the 64-kDa protein and cytotoxic activity against EM cells in ADCC there were many exceptions and overall the relationship between the two tests was not significant.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

PubMed1995

The ocular muscle cell is a target of the immune system in endocrine ophthalmopathy.

The exact pathogenic mechanism of thyroid-associated ophthalmopathy (TAO) remains unclear, and extensive studies on this disorder have resulted in often conflicting data. Well-known technical difficulties including the limited access to orbital tissues from patients with active and early disease, lack of an animal model and poor reproducibility of some of the immunological techniques used are in part responsible for this confusing situation. Despite this there is considerable evidence for eye muscle (EM) tissue involvement in the autoimmune reactions of TAO. Although the primary EM antigen(s) recognized by immunocompetent cells and autoantibodies has not been definitely identified, some good candidates, among them a membrane antigen of 64 kD which is also expressed in the thyroid, have been partially characterized. While it is unclear which component of the autoimmune reaction against EM-humoral or cell mediated-plays the more important role, autoantibodies seem to be responsible at least in part for the clinical features of the eye disorder. On the other hand, the orbital connective tissue (OCT) cells, especially the fibroblasts surrounding the EM fibers, seem to be extremely sensitive to stimulation by cytokines and other soluble proteins and immunoglobulins released in the course of an immune reaction in the muscle cells. Fibroblasts secrete large amounts of glycosaminoglycans and also participate in maintaining the autoimmune reaction. It seems likely that the EM is the main and primary target of the orbital autoimmune process whereas inflammation of the OCT is probably secondary.

Autoantibodies↗

PubMed1995

Human histocompatibility leukocyte antigen-DR and heat shock protein-70 expression in eye muscle tissue in thyroid-associated ophthalmopathy.

To investigate the expression and localization of human leukocyte antigen (HLA)-DR and heat shock protein-70 (HSP-70) in orbital tissue from patients with thyroid-associated ophthalmopathy (TAO), we carried out an immunohistochemical study using anti-HLA-DR and anti-HSP-70 monoclonal antibodies and a streptavidin-biotinperoxidase detection system. Eye muscle tissues were obtained at surgery from 38 patients with TAO and 8 control subjects. HLA-DR expression on eye muscle cells was demonstrated in orbital tissue from 2 of 3 untreated patients and 2 of 35 patients who had been treated with orbital irradiation or corticosteroids, in all of whom lymphocytic infiltration was also demonstrated. HLA-DR was not detected on eye muscle cells from 8 normal controls studied. HLA-DR was expressed on endothelial cells and interstitial cells from almost all patients with TAO and all 8 control subjects. HSP-70 was detected in eye muscle cells from 31 of the patients with TAO, including all 3 untreated patients, and 3 of the controls. Although the degree of HSP-70 expression did not correlate with the severity of the ophthalmopathy, significant expression of HSP-70 in eye muscle cells was more often demonstrated in patients with eye disease of short duration (83%) than in those with disease of longer duration (33%). These results support the notion that eye muscle fiber is an important target of the orbital autoimmune reactions that characterize TAO.

Adult↗