Discussion Of Relationship Between Protein 4.1R And Myasthenia Gravis

BackgroundMyasthenia gravis is an autoimmune disease of the nervous system. It has currently the most specific antigen determined in the AID. Its pathogenesis is not yet clear, and it has less effective treatment methods. At present, oral medications, injections and immunosuppressive treatment thymectomy are more common, Our previous use of proteomics from myasthenia gravis(MG) patients with abnormal thymus thymus tissue screened differential protein 4.1R. 4.1R(EPB41) is a member of the protein 4.1 family, which is the cytoskeletal proteins expressed in the cell membrane, cytoplasm and nucleus. It is involved in the regulation of cell proliferation and division, and its abnormal expression affects cell polarity and cell migration and so on. And other membrane cytoskeletal proteins, muscle contraction and other related proteins are interacting, which can cause the occurrence of such as anemia, muscle contraction disorders and other diseases. It can interacte with ion exchange proteins and PCMA1 b to transport ion and be adjusted calcium absorption features. Protein 4.1R is studied in a variety of tumor diseases, and found that the protein 4.1R is a negative regulator of molecular in tumor. Protein 4.1R leads to abnormal activation of T cell and has a negative regulatory role. And abnormal activation of T cell may lead to the occurrence of AID. 4.1R associated with the occurrence of many diseases, but it currently has fewer studies in AID and less in MG. The main topic in myasthenia gravis two typical clinical symptoms "autoimmune" and "weakness" as the main line, to explore protein 4.1R in the mechanism of MG. It will provide further understanding and help for MG pathogenesis and treatment. PurposeWe aim to explore the mechanism of 4.1R in myasthenia gravis using the protein 4.1R biological function of the DC cells, in the perspective of autoimmune. Besides, we use the expression and significance of 4.1R in diaphragmatic weakness models to reflect the mechanism of 4.1R in MG in the perspective of weakness. MethodsRecombine the full-length CDS of 4.1R gene and p CMV-C-Flag or p EGFP-N2 vector. The overexpression plasmid 4.1R-Flag and 4.1R-GFP were structred, and then they were transfected to DC cells, And which were added to G418 after drug screening, to obtain a stable strain DC-FLAG(control NC), DC-4.1R-FLAG(DC-4.1R-Flag), DC-GFP(control NC), DC-4.1R-GFP(DC-4.1R-Flag), the empty Virus and lentiviral packaging sh-4.1R were transfected to, to obtain a stable strain DC- RNAi-Control(NC) and DC-sh-4.1R. By RT-PCR and WB, DC-4.1R-Flag and DC-sh-4.1R were detected in the expression levels of m RNA and protein. And the cell surface molecules and costimulatory molecules were detected by flow cytometry. Using cell scratches and Transwell chamber experiments to study the migration of DC-4.1R-Flag and DC-sh-4.1R. Then, using RT-PCR, Western blot and immunohistochemistry to detect changes of 4.1R in diaphragmatic weakness model. Using Western blot and cell immunohistochemical methods to verify the changes of muscle nicotinic acetylcholine receptors after 4.1R gene knockdown in C2C12 cells. Results1. The overexpression plasmids of 4.1R-Flag and 4.1R-EGFP were successfully constructed.2. The stable strains of DC-Flag(control), DC-4.1R-Flag; DC-EGFP(control), DC-4.1R-EGFP, DC-RNAi-Control(control) and DC-sh-4.1R were successfully constructed.3. The volume of DC-4.1R-Flag stable strains get wider and its synapses get longer than the control group. Conversely, The volume of DC-4.1R-Flag stable strains get smaller and its synapses get more and thiner than the control group.4. Compared with the control group, the data of flow cytometry detection suggested that the expression of MHCII and CD80/86 was decreased in DC-4.1R-Flag cells(P< 0.05), while the expression of CD80/86 was increased in DC-sh-4.1R cells(P< 0.05), but the MHCII was no significant difference(P>0.05)5. According to the data of cell scratches and transwell experiment, the migration of DC-4.1 R-Flag cells was enhanced, while the migration of DC-sh-4.1 R cells was weakened(P<0.05).6. The expression of 4.1R was increased in the diaphragmatic weakness model in the both levels of m RNA and protein, that was proved by RT-PCR, WB and immunohistochemistry experiments.7. We found that protein 4.1R can adjust the change of desmin and myosin by detecting the expression of protein 4.1R and various muslce contranactile protein in diaphragmatic weakness model and C2C12 cells.8. And the expression of Ach R was up-regulated in C2C12-sh-4.1R cells by immunohistochemistry experiments, they have negative correlation.Conclusions1. 4.1R can affect the morphology, migration and antigen presentation capabilities of the DC cells.2. Protein 4.1R was associated with the occurrence of Gravis. And acetylcholine receptor expression was negatively correlated with protein 4.1R in C2C12 cells, So 4.1R was correlated with the occurrrence of MG, neither from the perspective of autoimmune or muscle weakness. And it is an important molecule of MG.