The Detection Of Autoantibodies Against Acetylcholine Muscarinic 3 Receptor In Sjogren Syndrome

BackgroundSjogren syndrome (SS) is a chronic autoimmune disease which occurs mainly in the fourth and fifthdecades and characterized by histologic and functional alterations of exocrine glands with progressive loss of salivary and lacrimal gland secretion. Respiratory, digesting, renal and nervous system are as well involved in SS. There are many mechanisms of loss of exocrine glands such as destruction of the acinar cells by lymphocytes infiltration and the antibody-mediated destruction of the exocrine glands. It has been recently proposed that pathology underlying the glandular hypofunction contributes inhibitory autoantibodies especially the autoantibodies directed against mucarinic recptors. The aim of this work is to test the positive rate of the anti-M₃ receptor antibody in SS , the relationship between the antibody and other clinical manifestations and the salivary flow rate decreased by the affinity-purified antibody from SS patients in mice.ObjectivesTo detect anti-M₃ receptor antibodies in Sjogren' s Syndrome (SS) patients, SLE patients and the donating controls by methods of Enzyme linked immunosorbent assay and Western blotting. And to explore its association with clinical manifestation. At last detect the affinity-purified antibody' s pathogenic mechanism in animal experiments. Materials and MethodsSpecimens:70 pieces serum of Sjogren syndrome patients from the SICCAobject in PUMCH from 2005-2007. 49 pieces serum of patients of SLE in-patients in PUMCH at the same time. 76 pieces serum of healthy donators in PUMCH. And the three groups match in sex and age.Methods: With the muscarinic 3 receptor(from sigma) as the antigen ,to detect anti-muscarinic 3 receptor antibody in SS, SLE and donator controlsusing methods of ELISA. The positive cut-off is (?)+SD in donator controls.And test the positive rate of anti-muscarinic 3 receptor antibody in SS, SLE by means of western blotting. Analyze the differences of the antibody-positive patients and antibody-negative patients in SS in aspects of Schirmer test,BUT,PEE on the corneas,the conjunctival epithelial defects, whole saliva flow rate in 5 minutes,FI,RF,IgG,anti-SSA and anti-SSB.Affinity- purified of human polyclonal IgG antibodies (15mg/kg)by Protein A chromatography was passive transfered to mice and detect the differences of the salivary secrection in the groups of IVIG (15mg/kg)which is the negative controls, atropine(0.125mg/kg) which is the positive controls and the human IgG.Results1.Using ELISA the positive rate of anti-M₃ antibody in SS is 47. 10%(33/70), in SLE is 4.02%(2/49) and in donator controls is 14.47%(11/76).The rate in SS is much higher than in SLE(47.10% vs 4.02%,P<0.05). Compared with patients in donator control the antibody is more frequently found in patients with SS(47.10% vs 14.47%, P<0.05).2. Using Western the positive rate of anti-M₃ antibody in SS is 60.0%, in SLE is 12.2% and in normal controls is 15.79%. Compared with patients with SLE and donator controls, the anti-M₃ antibody in SS is more frequently found,P<0.05.3. The incidence of IgG,rheumatoid factor, anti-SSB and FI>3 are higher in positive group than in negative group using ELISA. And the differences are significant,P<0.05. But the differences between the two goups in Schirmer test,BUT,PEE on the corneas,the conjunctival epithelial defects, whole saliva flow rate in 5 minutes and anti-SSA are not significant.4. The data of passive transfer to mice indicates that the recipient mice of purified human IgG from anti-M₃ antibody positive SS(15mg/kg) develop glandular hypofunction which is similar to those recipient mice of atropine. But that is not similar to those recipient mice of IVIG.And the differences between them are significant.P<0.05Conclusions1. The positive rate of anti-M₃ antibody in SS is much higher than that in SLE and in normal controls.2. The IgG, RF, anti-SSB and FI in SS patients with positive anti-muscarinic 3 receptor antibody are much higher than in patients with negativeanti-muscarinic 3 receptor antibody.3. The data of WS mice experiments indicates that the anti-muscarinic 3 antibody may decrease the salivary secretion.