Recombinant Human Acetylcholine Receptor Gamma Subunit Protein Induces Experimental Autoimmune Ocular Myasthenia Gravis In HLA-DQ8 Transgenic Mice

Objective:To establish a novel model of ocular experimental autoimmune myasthenia gravis (oEAMG) in HLA-DQ8 transgenic mice and study the immunologic pathogenesis of oEAMG .Methods:Eighteen HLA-DQ8 transgenic mice were randomly divided equally into control group, experimental group. The mice in experimental group were immunized with 20μg of E. coli plasmid expressing recombinant human acetycholine receptor gamma (AChRγ) subunit emulsified in CFA on day 0 and in IFA on days 30 and 60, while mice in control group were injected with PBS emulsified in CFA or IFA. Ocular symptom was evaluated by documentation of partial or full closure of one or both eyes and generalized muscle weakness was measured by grip strength machine. Sera were collected on days 15, 45 after the second immunization. The seurum anti-AChR antibody levels were tested by RIA and ELISA. The mRNA expression of mouse AChRα, fetalγandεsubunit proteins was quantitated in the extra-ocular muscles of each group by RT-PCR.Results:(1) Compared with the control group, the mice from experimental group exhibited partial or full closure of one or both eyelids, the scores of ocular symptom in experimental group were higher than that in control group (P<0．001) , and the oEAMG incidence between experimental(89%) and control group are significantly different (P<0．001) , (2) The 6th and 7th week after the 2nd immunization, experimental group mice had higher average grip strengths, as compare to control group mice(all P value <0．05) , ( 3 ) The serum anti-AChR antibodies levels were significantly higher in experimental group mice as compared to control group mice (p<0．01) , ELISA detected the serum anti-AChR IgM,IgG,Ig2b,Ig2c,IgG1 Antibodies levels were significantly higher (all P value <0．01) in experimental group mice as compared to control group mice,(4) Compared with the control group , the mRNA expression of AChR subunits(α,γ,ε) were increased in immunized mice (all P value <0．01) .Conclusions:Our findings suggest that oEAMG can be induced in HLA-DQ8 transgenic mice by immunizing with recombinant H-AChRγsubunit. Anti-AChR antibodies played a crucial role in the pathogenesis and were involved in the development of oEAMG in transgenic mice. The H-AChRγsubunit induced the autoimmune response to mouse AChR which led to destruction of AChR, and triggered the up-regulation of AChR genetic transcription, the mRNA expression of AChR subunits(α, γ,ε) were increased in AChR-immunized mice. This new model of oEAMG could be a valuable tool to study the pathogenesis of ocular MG (oMG) and generalized MG (gMG) in humans and further for clinical therapeutic analysis.