R. Åhlberg

Acetylcholine receptor-reactive T cells in myasthenia gravis: Evidence for the involvement of different subpopulations of T helper cells

Patients with myasthenia gravis have a high prevalence of acetylcholine receptor-specific T lymphocytes in the peripheral blood. Our earlier study shows that these T lymphocytes are stimulated to secrete interferon (IFN)-gamma and interleukin (IL)-2 in response to the autoantigen. Such stimulated T cells may be subdivided into different subsets according to the pattern of cytokine production. In the present study we have investigated the subpopulations of cells by analyzing their IL-4, IFN-gamma and IL-2 secretion pattern. Autoantigen-stimulated IL-4 secretion was found in 55% of the patients, IFN-gamma secretion in 86% and IL-2 secretion in 72%. T lymphocytes from all patients who responded with increased IL-2 secretion also showed increased IFN-gamma secretion. Stimulated IL-4 secretion was detected both in the presence and absence of stimulated IFN-gamma secretion. Depletion of monocytes/macrophages from peripheral blood mononuclear cell preparation and treatment of the cells with a mouse anti-human HLA-DR antibody abolished the secretion of IFN-gamma and IL-4. There were positive correlations between the numbers of IFN-gamma- and IL-2-secreting T cells and the numbers of B cells secreting antibodies against the acetylcholine receptor. Our results show that acetylcholine receptor-stimulated T lymphocytes secrete IL-4, IFN-gamma and/or IL-2. This T cell response is major histocompatibility complex (MHC) class II-restricted and monocyte/macrophage-dependent. Our study indicates that both Th1/Th2 or Th0 subpopulations of the T lymphocytes are involved in the autoimmune response in myasthenia gravis.

Treatment of myasthenia gravis with anti–CD4 antibody: Improvement correlates to decreased T–cell autoreactivity

We treated a patient with severe myasthenia gravis with a chimeric (murine/human) anti-CD4 monoclonal antibody (cM-T412) for 7 days and followed the therapeutic effect by standardized muscle function tests, single-fiber electromyography, and immunologic examinations of disease-specific B- and T-cell functions. Clinical and electrophysiologic improvement began within 4 days, lasted for 3 months, and was maximal between days 16 and 58. The CD4+ lymphocytes decreased to a minimum of 80 cells per μl of peripheral blood, recovered slowly during the first year of follow-up, and did not correlate with changes in disease severity. T-cell stimulation by human acetylcholine receptor was abolished by the treatment but became detectable at the time of worsening of symptoms. The concentration of acetylcholine receptor antibodies in serum was not decreased by the treatment. The results suggest that anti-CD4 antibody administration could be effective in the treatment of severe myasthenia gravis and indicate that acetylcholine receptor-specific T lymphocytes might contribute to the disturbed neuromuscular transmission in the disease.

Defective T lymphocyte function in nonthymectomized patients with myasthenia gravis

In vitro functional properties of peripheral blood mononuclear cells were evaluated in 29 patients with myasthenia gravis and in 11 healthy controls. Spontaneous cell proliferation was higher in patients than in controls. The production of interleukin-2 and interferon-gamma and the proliferative response to different mitogens were reduced in the patients. A positive correlation was found between the production of interleukin-2 and interferon-gamma. These defects in T cell function were the most pronounced in nonthymectomized patients. Patients with severe disease had a higher percentage of cells bearing the interleukin-2 receptor and a higher spontaneous production of tumor necrosis factor alpha in cell culture than in patients with mild disease. There was no difference between patients and controls in the level of soluble interleukin-2 receptor in cell culture supernatants or in sera. The results indicate a partially suppressed T cell function in myasthenia gravis. This defect was less pronounced in patients studied after thymectomy.

The autoimmune T and B cell repertoires in monozygotic twins discordant for myasthenia gravis

Two pairs of monozygotic twins, discordant for myasthenia gravis (MG) for more than 30 years, were studied regarding T cell and antibody reactivity against disease related autoantigens, the acetylcholine receptor, one idiotypic and one anti-idiotypic human monoclonal antibody. The healthy and myasthenic twins had very similar autoantibody repertoires. IgG fractions from both healthy and myasthenic twins had the same capacity to decrease the free acetylcholine receptor content in mice after passive transfer. In comparison with their myasthenic sisters, the healthy twins had lower T cell responses against the acetylcholine receptor.

Levels of CD5+ B lymphocytes do not differ between patients with myasthenia gravis and healthy individuals

CD5+ B cells might be involved in autoimmunity mainly as autoantibody-producing cells. To investigate the possible role of these cells in myasthenia gravis, we studied the numbers of CD5+ B cells, CD5+ B cells, and CD19+ B cells as well as CD5+ T cells in the peripheral blood from 31 patients with myasthenia gravis and 31 healthy individuals. Both absolute percentages (percent of peripheral blood mononuclear cells) and relative percentages (percent of total CD19+ B cells) of CD5+ B cells were the same in patients as in controls. The numbers of CD5+ B cells and CD19+ B cells were the same in both groups, whereas CD5+ T cells were lower in the patients. There was no correlation between clinical stage, sex, thymectomy, or pathology of thymus and the levels of CD5+ B cells, CD5+ B cells, or CD19+ B cells. Patients treated with azathioprine had lower levels of CD5+ B cells than untreated patients and controls. Our results show that patients with myasthenia gravis have the same levels of CD5+ B cells as healthy individuals.

THE EFFECT OF THYMECTOMY ON AUTOREACTIVE T- AND B-LYMPHOCYTES IN MYASTHENIA GRAVIS

Eleven patients with myasthenia gravis were followed for three years after thymectomy. Acetylcholine receptor-specific T-cell stimulation was found in 8/11 patients before operation as compared to 2/11 three years after thymectomy. Changes of T-cell antireceptor-reactivity were commonly paralleled by changes in disease severity. The numbers of cells secreting IL-2 upon stimulation with human acetylcholine receptor correlated with those secreting IFN-gamma. T-cell reactivity against a monoclonal acetylcholine receptor antibody did not decrease after thymectomy. Such reactivity could reflect a beneficial immune response counteracting anti-receptor reactivity. The frequency of autoantibody-secreting cells remained unchanged, while the serum concentration of acetylcholine receptor antibodies started to decrease one year after thymectomy. All examined thymus-cell suspensions contained autoreactive T- and B-lymphocytes. There was a preferential enrichment of autoreactive lymphocytes in the thymus in a few patients with recent onset of disease.

T-cell epitopes on the human acetylcholine receptor α-subunit residues 10-84 in myasthenia gravis

In myasthenia gravis the production of anti‐acetylcholine receptor antibodies is modulated by acetylcholine receptor‐specific T cells. Most B‐and T‐cell epitopes are located on the α‐subunit of the receptor. In order to map the fine specificity of the antigen‐specific T cells in myasthenia gravis, T‐cell stimulation in response to 70 hexapeptides was studied in 24 patients and 24 healthy individuals. The hexapeptides overlapped with one amino acid and represented residues 10‐84 of the NH2‐terminal part of the α‐subunit of the receptor. The IFN‐γ secretion from single T cells was used to detect T‐cell stimulation.

T cell epitopes on the human acetylcholine receptor α-subunit residues 10–84 in myasthenia gravis

In myasthenia gravis the production of anti-acetylcholine receptor antibodies is modulated by acetylcholine receptor-specific T cells. Most B- and T-cell epitopes are located on the alpha-subunit of the receptor. In order to map the fine specificity of the antigen-specific T cells in myasthenia gravis, T-cell stimulation in response to 70 hexapeptides was studied in 24 patients and 24 healthy individuals. The hexapeptides overlapped with one amino acid and represented residues 10-84 of the NH2-terminal part of the alpha-subunit of the receptor. The IFN-gamma secretion from single T cells was used to detect T-cell stimulation. A significant difference in the T-cell response to several of the peptides was found between patients and healthy controls. The majority of the hexapeptides induced T-cell stimulation in at least one of the patients. Peptide-induced T-cell stimulation was evident in all but one of the patients. The results indicate that different epitopes and multiple T-cell clones are involved in the T-cell recognition of the acetylcholine receptor.