ABSTRACT
Myasthenia gravis (MG) is considered as an autoimmune disease mainly mediated by antibodies against acetylcholine receptor. In recent years, other targets related to MG have been the subject of interest. Our previous research found that protein P25 was lower in muscles of MG patients using two-dimensional electrophoresis. In present study, anti-serum to P25 was prepared, immunohistochemistry and ATPase staining revealed that P25 was a muscle specific cytosolic protein and was mainly distributed in type I muscle fibers. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and precise molecular weight derived from mass spectrometer identified P25 as carbonic anhydrase III (CA III). Some members of CA family are related to autoimmune diseases and CA III is recently reported to be involved in rheumatoid arthritis. The results of immunoblot in this report showed that the level of CA III is specifically insufficient in the skeletal muscle of MG patients. The possible roles that CA III play in MG need further elucidation.
Subject(s)
Carbonic Anhydrase III/deficiency , Muscle, Skeletal/enzymology , Myasthenia Gravis/enzymology , Adolescent , Adult , Antibodies, Monoclonal/immunology , Carbonic Anhydrase III/immunology , Cytoplasm/enzymology , Female , Humans , Immune Sera/immunology , Male , Middle Aged , Muscle Cells/enzymology , Muscle Fibers, Fast-Twitch/enzymology , Muscle Fibers, Slow-Twitch/enzymology , Pectoralis Muscles/enzymology , Peptide Mapping , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Young AdultABSTRACT
OBJECTIVES: To confirm the specific decrease of P25 protein, a novel myasthenia gravis (MG) associated protein, in myasthenia gravis (MG) patients and to study the tissue and species specificity of its expression. METHODS: Samples of great pectoral muscle were taken from 28 MG patients, 17 males and 11 females, 10 of which being complicated with thymoma and 18 with hyperplasia of thymus, during thymectomy, and 24 wounded persons as normal controls, 14 males and 10 females, during thoracic surgery. Ten samples of skeletal muscle were taken from 10 patients with other muscular disorders (OMD) during skeletal muscle biopsy. Six samples of normal skeletal muscle, smooth muscle, brain, lung, kidney, and skin were taken from persons who died of accident. Two samples of normal cardiac muscle and thymus gland were taken from a patient undergoing cardiac valve replacement. Eight samples of animal skeletal muscle were taken from the thighs of pig, cattle, dog, rabbit, rat, mouse, chicken, and frog. Immunohistochemistry was used to examine the expression of P25 protein in the samples from the MG patients and the normal controls. Western blotting was used to detect the expression of P25 in the samples from MG patients, normal controls, OMD patients, 8 samples of different human tissues, and skeletal muscle samples from 8 different animals. RESULTS: Immunohistochemistry showed that the staining intensity of P25 protein in skeletal muscle of MG patients was much lower than that of the normal controls. Western blotting showed that the relative density value of P25 protein in MG patients was 1.04 +/- 0.18, significantly lower than those in the normal controls and OM patients (1.27 +/- 0.21 and 1.21 +/- 0.15 respectively. P < 0.01 and P < 0.05). No statistically significant difference in expression of P25 protein was found among different clinical and pathological types of MG patients. Among the 8 human tissues P25 protein was expressed only in skeletal muscle and among the 8 animal samples of skeletal muscle P25 protein expression was found only in swine skeletal muscle. CONCLUSION: The expression of P25 protein is significantly decreased in skeletal muscle of MG patients. The expression of P25 protein is tissue- and species specific.
