Alzheimer’s Disease Immunotherapy Mechanisms And Serum Levels Of Inflammatory Cytokines

Background and TargetAlzheimer’s disease （AD）is the most common type of dementia. β-amyloidpeptide (Aβ) is a major pathogenic factor of AD. In recent two decades,Aβ-specific immunotherapy is a crucial point in the research of prevention andtreatment of AD. In AD transgenic mice, both active immunization and passiveimmunization can effectively remove the Aβ plaques in the brain andcerebrospinal fluid (CSF), increasing Aβ levels in serum and improvingcognitive function. Active immunization of primates found no pathologicaleffects，such as meningoencephalitis. In the phase II clinical trial of the activeimmunization drug AN1792，6%of AD patients had meningoencephalitis.Passive immunotherapy drugs bapineuzum which is against human AβN-terminal sequence of a humanized monoclonal antibody, and Solanezumagainst Aβ intermediate sequence did not show significant efficacy and did notimprove cognitive function of mild to moderate AD patients compared with theplacebo group in the phase Ⅲ clinical trial. To date, the clinical trials haveshowed that immunotherapy fails to function..The mechanism of immunotherapy failure is unclear, we deduce that apossible mechanism may be the “dust-raising” effect, that is, antibody passthrough the blood-brain barrier (BBB) and combine with Aβ plaques in the brain,which promote the process of disaggregation of plaques into soluble Aβ, andfurther the aggregation of the oligomers. Though this process promote theclearance of the Aβ plaques, it increase the level of Aβ oligomers and increasethe neurotoxicity. Some academics believe that neuroinflammation is involvedin the pathology of AD, and that simply clear Aβ plaques in the brain would not improve cognitive function. Some researches find that in AD transgenic miceand AD patients, the peripheral glial cells around Aβ plaques are overactivatedand inflammatory cytokines are highly expressed, while the expression ofanti-inflammatory is reduced. In the progress of AD, glial cells can bestimulated by Aβ, and release a large number of inflammatory cytokines, whichincrease the synthesis of Aβ. This implys that the AD pathology is closelyrelated with the release of inflammatory cytokines. There are a number ofclinical researches about inflammatory cytokines in peripheral blood, but theresults are uncertain. We try to discuss two aspects:1. to observe anti-Aβantibody whether or not can promote the formation of Aβ oligomers afterdisaggregating Aβ fibers,.2. to test the serum levels of IL-6, IL-1β, TNF-α,IFN-γ in AD patients, and to explore the relationships between inflammatorycytokines and cognitive function.Materials and Methodsincubate Aβ monomers into fibers, then6E10(anti-Aβ N terminal),4G8(anti-Aβ middle),8G7(anti-Aβ C terminal)were co-incubated with Aβ fibersto promote disaggregation. After incubation, Thioflavine T fluorescence wasutilized to test Aβ fibers disaggregation; Fibers and products were observedunder TEM; Western Blot was used to detect the Aβ oligomer in the supernatantafter centrifugation. Thereafter, neurotoxicity of the samples to SH-SY5Y cellswere tested using MTT assay and axon growth assay. In vivo experiments: localinjection of disaggregated products into the cerebral cortex of SD rat wasconducted to observe their neurotoxic effects.110AD patients from our outpatient and inpatient department during theperiod of2010to2013were compared with120age, cognitive function andsex-matched normal controls from our physical examination center. Collectclinical data and test blood, serum concentrations of inflammatory cytokines byenzyme-linked immunosorbent assay (ELISA). Compare inflammatory cytokineslevels of the two groups; partial correlation analysis was used to analyse therelationship of serum concentrations of inflammatory cytokines with cognitivefunction (MMSE) and dementia severity (CDR); not-Logistic regression method for multivariate analysis of factors resulting in AD.Result1.Comparing with the4G8+Aβ fibers,8G7+Aβ fibers, IgG+Aβ fibersand PBS+Aβ fibers,6E10+Aβ fibers can disaggregate Aβ fibers intooligomers; and the oligomers can inhibit the outgrowth of the axon and promotethe cell death;4G8,8G7hardly disaggregate Aβ fibers.2.(1) IL-6, IL-1β levels In AD patients are significantly higher than thosein normal controls.(2) Serum level of IL-1β is negatively correlated withMMSE score （r=-0.184，p=0.007）in all subjects.(3) Serum levels of the IL-6,IL-1β, TNF-α and IFN-γ levels are not correlated with severity of AD (CDRscores).Conclusion1.Anti-Aβ N terminal antibody promotes Aβ fibers disaggregation whileproducing more neurotoxic Aβ oligomers.2.Serum inflammatory cytokines levels are correlated with the cognitivefunction, but not with disease severity in AD patients.