| Objective:1. To explore rat model preparation of experimental autoimmune myasthenia gravis(EAMG) with rat-derived 97-116(Rα97-116) peptide of the acetylcholine receptor(AChR)α-subunit and provide a simple and available animal models for basic researches of MG..2. To study effects of nasal tolerance with Rα97-116(V108A) on EAMG and its immunity function.3. To investigate the expression of Fas and Bcl-2 in thymus of EAMG rats after nasal immune tolerance and its mechanism.Methods:We divided Female Lewis rats(6-8 weeks) into three groups (EAMG group, control group and tolerance group). The EAMG rats model were prepared by mult-spot subcutaneous injection with mixture of rat-derived AChRα97-116(Rα97-116) and complete Freund's adjuvant(CFA) in EAMG group and tolerance group and boosted on day 30 and 60 with the same peptide in incomplete Freund's adjuvant(IFA),The control group rats were immunized with phosphate buffer saline(PBS). Clinical manifestation was evaluated by measurement of body weight and Lennon clinical score. EAMG was further confirmed by 3,5 Hz repetitive nerve stimulation (RNS) for positive decremental respnse and ELISA assay for serum AChR-Ab titers.At the 8th day after immunized with Rα97-116 for three times, The rats of EAMG group and tolerance group were immunized respectively with the peptide of alanine replacement of valine at position 108 (V108A) corresponding to Rα97-116 sequence? Rα97-116(V108A) and PBS buffer solution for 10 days by route of nasal mucous(30μg/kg.d). Then their body weight and Lennon score were measured again,serum anti-AChR antibody were tested by ELISA. the apoptosis thymocytes in rat thymus of three groups were counted by TUNEL methods and the expression of Fas and Bcl-2 in the thymus were studied by immunohistochemical(IHC).Results:1. EAMG occurred in 71.7% of Lewis rats in the EAMG group ,which was confirmed by the Lennon clinical score>1 grade, the positive decremental response D5 >10% of 3,5 Hz RNS and the ratio >2.1 of the serum AChR-Ab titers to the control group. The achievement ratio of EAMG model was 71.7%.2. The 20th day after receiving nasal tolerance with Rα97-116(V108A) , the body weights of rats in tolerance group were increased significantly compared with those of control group(P﹤0.05).3. The 10th after receiving nasal tolerance with Rα97-116(V108A), the mean Lennon score of rats in tolerance group decreased obviously compared with control group(P﹤0.05).4. Compared with control group , the amount of serum anti-AChR IgG in tolerance group decreased significantly on the 16th day after nasal tolerance with Rα97-116(V108A) (P﹤0.05).5. Numbers of apoptosis thymocytes in EAMG group was less than those of control group and golerance group respectively (P<0.01).6. Contrary to the expression of Bcl-2 ,The expression of Fas in the thymus of EAMG group rats was lower than that of control and tolerance group. Conclusions:1. Immunization with the synthetic Rα97-116 peptide can induce EAMG in Lewis rats .The achievement ratio of EAMG model was 71.7% . which will provide a potential animal models for basic researches of MG..2. Nasal mucous tolerance with Rα97-116(V108A)could ameliorate muscular weakness of EAMG rats. Which mechanism of therapy might be related to passway of apoptosis. The apoptosis of auto-react lymphocyte maybe participated In the mechanism of nasal immune tolerance.
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