Preliminary Study Of Mitochondrial Injury On Patients With Myasthenia Gravis

Background Myasthenia gravis (MG) is a common autoimmune disease in neurological system, its characteristic is the transmission impairment in neuromuscular junction. Its pathogenesis is due to the acetylcholine receptor antibodies (Ach-Ab) produced by immune abnormalities,which may damage the acetylcholine receptor in the postsynaptic membrane. However, the previous studies found that about10-20%of MG patients have negative AChR-Ab, and the underlying pathogenesis is still unclear. Many scholars have found some other antibodies in the patients with negative AChR-Ab,among them,the muscle-specific tyrosine kinase antibody (MuSK-Ab) is the most significant. MuSK-Ab can be detected in4-49%of AChR-Ab (-) patients, and they share different clinical manifestations from AChR-Ab (+) MG patients. Eye-bulbar muscles weakness is always the first and major symptom, which is very similar to that of mitochondrial myopathy. Recent studies have found that obvious mitochondrial dysfunction is detected in the MG patients by muscle histochemical study, especially in the MuSK-Ab postitive patients. Therefore, we hypothesize that mitochondrial dysfunction is also involved in the pathogenesis of MG. This study aims to explore whether mitochondrial dysfunction exists in MG and whether it is related to the antibody type.Part One:Detection of antibodies in MG patients’sera and observation of mitochondrial dynamics in peripheral blood mononuclear cellObjectives To compare the clinical features of myasthenia gravis(MG)with different antibodies, and observe the mitochondrial dynamics. Methods The MuSK and AChR antibodies were detected in the sera of62MG patients by radioimmunoprecipitation assay (RIPA).The clinical features of AChR-Ab positive、MuSK-Ab positive and sera negative MG patients were compared. Peripheral blood mononuclear cells wer extracted from6patients, labelled by mitochondrial fluorescent probe, and were observed for mitochondrial dynamics by using laser confocal microscopy. Results There were51patients with positibe AChR-Ab(82.3%). and1case with MuSK-Ab (1.6%). This MuSK antibody positive case was AChR antibody negative, accounting for9.1%of the AChR-Ab negative patients (1/11). The MuSK-Ab positive patient is an elderly men, manifests as bulbar weakness with no no thymic abnormalities. There was no significant difference between the other two groups regarding sex and age at onset. The initial symptoms mostly manifests as external ophthalmoplegia, with thymic hyperplasia or thymoma. The level of AChR antibodies is related to the disease severity (P=0.045). Among the6patients with mitochondrial dynamics test,5of them indicated abnormal dynamics, including2cases with positive AChR antibody and3cases with negative antibodies. Conclusions MuSK antibodie is only detected in the patient with negative AChR-Ab. And its clinical features include bulbar invlovment without thymus abnormality. Some MG patients have abnormal mitochondrial dynamics in the peripheral blood mononuclear cells Part Two:Mitochondrial function in muscle of myasthenia gravis patientsObjective To investigate the mitochondrial function in the muscle tissue of MG patients. Methods This study included9cases of MG patients,9cases of CPEO patients and9cases of normal control. The frozen sections of muscle biopsy were stained by histochemical staining, and then observed under the microscope to determine the mitochondrial function.Mitochondria was isolated by differential centrifugation from the muscle samples of above patients. The activities of respiratory chain complexes I-V and citrate synthase were determined by colorimetric assay. The activity of CI/CS, CII/CS, CIII/CS, CIV/CS, CV/CS were compared, to determine the mitochondrial oxidative phosphorylation. Results By histochemical study, cytochrome C oxidase (COX) negative fibers were observed in MG and CPEO patients, suggesting that mitochondrial function is impaired. By mitochondrial respiratory chain complex activity analysis, the activity of omplex IV is found to be significantly reduced in MG patients. However, CPEO patients had predominantly low activity of respiratory chain complexes III and IV. Conclusions Mitochondrial function abnormalities could be found in the muscle tissue of MG patients. Conclusions:1. The MuSK antibody positive case was AChR antibody negative, accounting for9.1%of the AChR-Ab negative patients. The MuSK-Ab positive patient is manifests as bulbar weakness with no no thymic abnormalities. In the other two groups, the initial symptoms mostly manifests as external ophthalmoplegia, with thymic hyperplasia or thymoma.2. MG patients had mitochondrial dynamics abnormalilies in peripheral blood mononuclear cell, performing excessive fission, but had no correlation with the type of antibody3. The muscle tissue of MG patients exists mitochondrial function abnormalities, manifesting COX-negative fibers in histochemical staining and low mitochondrial respiratory chain complex IV (COX) activity.