The Novel Anti-Aβ3-10 Monoclonal Antibody 7B8 Improves The Blood-brain Barrier And Cognitive Impairment Of APP/PS1 Mice By Reducing Senile Plaque Deposition And Inflammatory Response

Objective: Alzheimer’s disease(AD)is the most common type of dementia,manifested clinically by progressive memory,cognition,executive and other functional disorders.The main pathological features are the deposition of amyloid beta(Aβ)outside the cell to form senile plaque(SP)and the nerve fiber tangles,neuronal loss and neuroinflammation formed by excessive phosphorylation of intracellular tau protein.There are many hypotheses for the pathogenesis of AD,among which the "amyloid cascade hypothesis" is highly recognized.It is hypothesized that the formation of senile plaque by Aβ deposition is a key link in the onset and development of AD,which in turn causes pathological changes such as tau protein phosphorylation,neuronal loss and neuroinflammatory response.Although passive immunization for the cause can reduce the senile plaques of the brain parenchyma,it cannot effectively remove the senile plaques of the blood vessel wall,and may even aggravate cerebral amyloid angiopathy(CAA),cerebral edema and microhemorrhage.In clinical trials of the monoclonal antibody aducanumab,although the high-dose(10 mg/kg)group can significantly improve cognitive function in AD patients,amyloid-related imaging abnormalities(ARIA)may occur,including angiogenic edema(ARIA-E)and micro-hemorrhage(ARIA-H).It may be that high-dose antibodies over activate glial cells during passive immunity,aggravate the inflammatory response,damage the blood-brain barrier and vascular wall structure,and lead to extravasation of blood.The occurrence of ARIA is mainly related to the dose of antibodies,and once ARIA appears,it may be necessary to reduce the dose of antibodies,limiting the effective use of antibodies in the brain.In order to explore the clearance effect and possible side effects of novel anti-Aβ3-10 monoclonal antibody 7B8 on age spots,better understand the relationship between ARIA and passive immunotherapy of AD,and explore the effect of antibodies on age spots and the effect of antibodies on the blood-brain barrier and inflammatory response through the action of monoclonal antibody 7B8 on APP/PS1 double-transgenic mice.Methods: 1)The synthetic anti-Aβ3-10 monoclonal antibody 7B8 was passively immunotherapy with APP/PS1 double-transgenic mice by intraperitoneal injection,injected at a dose of 10mg/kg from the age of 6 months,and a total of 8 immunizations were carried out once a week(7B8 group);Another group of APP/PS1 doubletransgenic mice of the same month of age were injected with the same dose of IgG antibody as a negative control(IgG group)in the same way;C57BL/6J rats of the same month of age were in the wild group(WT group);2)the cognitive function changes of mice were detected by behavior,and the changes of age plaques in the wall of pia mater vessels were detected by fluorescence microscopy and two-photon microscopy;3)After cardiac saline perfusion,the brain tissue was dissected and removed,the left brain was fixed with 4% paraformaldehyde for paraffin sectioning,and the right brain made brain tissue homogenate and separated vascular tissue;4)Detection of brain parenchyma and blood vessel wall Aβ deposition and inflammation-related glial cells activation by immunohistochemical staining,Prussian blue staining to detect microbleeding in brain tissue,Masson and EVG staining to detect vascular collagen and elastic fibers,Western blot to detect the expression of blood-brain barrier and inflammatory signaling pathway proteins,immunofluorescence staining to detect the expression of age spots,vascular wall smooth muscle cells,endothelial cells and astrocytes.It was compared with wild-type background mice(WT group)of the same month of age.Results: 1)The results of immunohistochemistry and two-photon microscopy showed that the Aβ deposition in the vascular wall(pia mater,brain parenchyma)and brain parenchyma(hippocampus,cortex)of the 7B8 group and WT group was significantly lower than that in the IgG group(P<0.05);2)The results of HE staining and WB showed that the neuronal damage in the 7B8 group and WT group was significantly lower than that in the IgG group(P<0.05);3)The behavioral test results showed that the 7B8 group,The score of the fossa in the WT group was significantly higher than that in the IgG group(P<0.05),and in the new object recognition experiment,the resolution index of the 7B8 group and WT group was significantly higher than that of the IgG group(P<0.05);4)Prussian blue staining showed that there was no significant difference in perivascular and brain parenchymal microhemorrhage between the 78 B group and the transgenic mice in the IgG group(P>0.05);5)immunofluorescence and histochemical staining results showed that Aβ40 deposition of leptomeninges vascular wall of 7B8 group The damage of smooth actin and elastic fibers was significantly lower than that of IgG group(P<0.05);6)Masson staining: there was no significant difference between the perivascular space between IgG group and 7B8 group(P>0.05);7)immunofluorescence staining results showed that 7B8 group,The fluorescence intensity of endothelial cells in WT group was significantly higher than that in IgG group(P<0.05);8)WB results showed that the expression level of blood vessel wall RAGE in the 7B8 group was significantly lower than that in the IgG group(P<0.05),and the expression level of blood vessel wall LRP-1 in the 7B8 group was significantly higher than that in the IgG group(P<0.05);9)WT results showed that the brain tissue tight junction protein of the 7B8 group and the WT group(ZO-1,CLDN-5,Occludin)expression was significantly higher than IgG(P<0.05);10)immunohistochemistry and WB results showed that the activation degree of microglia and astrocytes in WT group and 7B8 group was significantly lower than that in IgG group(P<0.05);11)The results of immunohistochemistry showed that the levels of inflammatory factors IL-6 and cortical TNF-α in the 7B8 group were significantly lower than those in the IgG group(P<0.05),and the overall levels of TNF-α and IL-1β were not significantly increased compared with those in the IgG group.12)Immunofluorescence and WB results showed that the IgG group was more active than the 7B8 group,and the expression of vascular wall adhesion factors(ICAM-1,VCAM)in the 7B8 group was significantly lower than that in the IgG group(P<0.05);13)WT results showed that the expression levels of HMGB-1,RAGE and NF-k B proteins in the 7B8 group and WT group were significantly lower than those in the IgG group(P<0.05).Conclusion: 1)Antibody 7B8 passive immunization APP/PS1 transgenic mice can improve cognitive function impairment;2)Antibody 7B8 passive immunization APP/PS1 transgenic mice can effectively remove brain tissue and vascular wall Aβdeposition,and does not increase perivascular space and intracerebral microhemorrhage;3)Antibody 7B8 passive immune APP/PS1 transgenic mouse can protect the structure of neurons,blood vessel wall and BBB;4)After passive immunization of APP/PS1 transgenic mice with antibody 7B8,it can reduce glial cell activation and inflammatory response by inhibiting the HMGB-1/RAGE/NF-k B signaling pathway.