Effects Of Monoclonal Antibody 7B8 On Aggregation Kinetics Of Aβ42

Objective: The main clinical features of Alzheimer’s disease(AD)include memory loss,cognitive decline,language impairment,and dementia.The two major pathologic changes in the disease are amyloid-β(Aβ)found in plaques and hyperphosphorylated tau protein found in neurofibrillary tangles(NFTs).The two main forms of Aβ are Aβ40 and Aβ42,of which Aβ42 tends to form aggregates because it is more difficult to dissolve.In addition,recent studies have found that Aβ oligomers may be the main toxic substances in the pathogenesis of Alzheimer’s disease.Therefore,inhibition of the self-assembly process of Aβ42 has become one of the main potential therapeutic strategies against AD,and designing in a reasonable manner that can selectively inhibit monomer aggregation to form oligomers is a critical therapeutic approach.Previous studies have confirmed that active and passive immunotherapy targeting Aβ3-10 can improve the cognitive function of transgenic mice.On this basis,this study further investigated the effect of monoclonal antibody 7B8 targeting Aβ3-10 on the aggregation process of Aβ42,and analyzed and summarized the mechanism of action of monoclonal antibody 7B8.Methods: 1)ELISA demonstrated the binding properties of monoclonal antibody 7B8 to Aβ42 monomer and fibrils.2)Immunohistochemical analysis was performed on the brain tissue of double-transgenic AD mice,and the mechanism of action of monoclonal antibody 7B8 and 6E10 was compared and analyzed.3)The reaction trend of Aβ42 monomer was analyzed by Th T fluorescence and the appropriate reaction concentration of monomers was determined.4)The data were fitted to the models of primary,secondary and extended nucleation,and it was found that monoclonal antibody 7B8 had the most obvious effect on secondary nucleation.It was speculated that monoclonal antibody 7B8 could reduce the contact between Aβ42 monomer and fiber surface,thus inhibiting the formation of oligomers.5)Aggregation kinetics was used to analyze secondary nucleation,and the effects of different concentrations of antibodies and different concentrations of prefabricated fibrils on the aggregation process of Aβ42monomer were detected.6)Aggregation kinetics was used to analyze the monomer elongation process,and the influence of antibodies on the aggregation process of Aβ42 was detected when different concentrations of antibodies,different concentrations of prefabricated fibrils and prefabricated fibrils prepared from antibodies were added to Aβ42 monomer.7)The effect of monoclonal antibody 7B8 on monomer elongation was analyzed by aggregation kinetics.8)The influence of 7B8 on microscopic aggregation steps is further summarized.Results: 1)The ELISA results showed that monoclonal antibody 7B8 could bind to Aβ42 monomer and fibrils,but the binding to monomer was weak(P<0.05).2)Immunohistochemical analysis showed that 7B8 and 6E10 had similar binding properties in mice,and could bind to senile plaques.3)According to the trend of monomer reaction,the specific processes involved in aggregation in the experiment were determined and analyzed.The microscopic reaction steps that could be observed mainly included primary nucleation,secondary nucleation and elongation.4)The data were fitted with the models of primary and secondary nucleation and elongation,which showed that monoclonal antibody 7B8 had the most obvious effect on secondary nucleation.Therefore,it can be said that in this step,antibodies inhibit the secondary nucleation process,reduce the contact between monomer and the surfaces of existing fibrils,and thus inhibit the formation of oligomers.5)Once again,the antibody inhibited secondary nucleation by slowing the rate of reaction and reducing the number of fibers detected in the reaction plateau when the prefabricated fibril were added at a concentration of 2%.6)When 30% of the prefabricated fibrils were added,the antibody still had some inhibitory effect on the aggregation reaction,but it was not obvious.7)The prefabricated fibrils synthesized with antibodies and the prefabricated fibers synthesized without antibodies were added to the reaction system respectively.The aggregation trends of Aβ42 under different conditions were compared,which further indicated that 7B8 had an inhibitory effect on secondary nucleation.8)Monoclonal antibody 7B8 inhibited both primary and secondary nucleation during oligomer formation.Conclusion: In this study,it was preliminarily verified that monoclonal antibody 7B8 has binding effect on Aβ42 monomer and fibrils by ELISA and immunohistochemistry,and the binding effect on fib rils is more obvious.The effect of monoclonal antibody 7B8 on the aggregation kinetics of Aβ42monomer was analyzed by Th T fluorescence method.After fitting the fluorescence values of the reaction to the reaction model,the results showed that the monoclonal antibody 7B8 had more obvious effects on the primary and secondary nucleation,that is,inhibiting the main step of oligomer formation,while the monoclonal antibody 7B8 had less effect on the fib rils fragmentation and elongation.