Deposition Of Exogenous Aβ₄₂ In Hippocampus Of HIAPP Transgenic Mice And Its Mechanism

Objective Alzheimer’s disease（AD）is a common neurodegenerative disease characterized by the memory impairment,aging dependent and progressive aggravation.The main pathological feature of AD is characterized byβamyloid deposition which consists of the monomer,oligomer or fibrillar amyloid.These amyloid deposits have strong neurotoxic effects and are the main components of senile plaques.Diabetes mellitus is characterized by aberration of glucose metabolism and utilization,which can be divided into type 1 and type 2 diabetes mellitus.Type 2diabetes mellitus（T2DM）accounts for as much as 90 percent of diabetes.Islet amyloid polypeptide（IAPP,amylin）deposition in pancreatic islet is the pathological features of T2DM.The secretion of islet amyloid peptides is harmless under steady-state conditions.However,when its production or processing is out of balance,cytotoxicity oligomers can be formed.These amyloid proteins can cause dysfunction of isletβcells.At the same time,the peripheral IAPP can enter the central nervous system（CNS）through the blood brain barrier（BBB）.There are IAPP receptors in the CNS,so peripheral IAPP plays multiple roles in the CNS.Both AD and T2DM are multifactorial diseases,which are characterized by local deposition of amyloid protein,although there are great differences in phenotype.Although the sources of IAPP and Aβare independent,their fibrillar sequences are highly homologous.Although the incidence of AD in the T2DM population is much higher than in the normal population,the link between the two diseases in the pathogenesis is not clear.Therefore,oligomer Aβ₄₂2 was stereotaxically injected into the dentate gyrus（DG）of hIAPP^+/-mice to study the interaction between hIAPP and Aβ₄₂2 in the brain and its possible mechanism.Methods1.Evaluation of peripheral islet function and glucose utilization in hippocampal neurons of 2-month-old wild type/hIAPP^+/-transgenic mice:Fasting blood glucose level,insulin tolerance test（ITT）and glucose tolerance test（GTT）were detected to evaluate islet function.Four hours after injection of 2-NBDG into the hippocampus,brains were perfused,removed and sliced to evaluate of neuronal glucose utilization levels in the hippocampus.2.Injection of oligomer Aβ₄₂2 into the bilateral hippocampal DG region of mice and evaluation of Aβ₄₂2 and Amylin expression after injection:The synthetic oligomer Aβ₄₂2 was stereotaxically injected into the bilateral hippocampus DG region of wild type/hIAPP^+/-transgenic mice at the age of 2 months,and the control group was injected with vehicle control.After injection of oligomer Aβ₄₂2 for 8 hours,24 hours,3days,7 days and 15 days,the expression of Aβ₄₂2 in the hippocampus of mice was detected by immunohistochemistry.The expressions of Aβ₄₂2 and Amylin in hippocampus and olfactory bulb of mice were detected by immunofluorescence after2 and 6 months of injection.3.Behavioral test:2 months and 6 months after injection of oligomer Aβ₄₂,the spontaneous activity of mice was evaluated by opening experiment,and the social recognition ability of mice was evaluated by social experiment.Olfactory preference test was used to evaluate the olfactory function and olfactory-dependent memory function of mice,and object recognition was used to evaluate the spatial memory function of mice.4.The possible mechanisms of brain damage after injection of oligomer Aβ₄₂2 into wild type/hIAPP^+/-transgenic mice:The expression of mitochondrial function related proteins in hippocampus was detected by Western Blot.The expression of synaptic markers in hippocampus and olfactory was detected by Western Blot.The number of neural stem cells in hippocampus and SVZ was detected by immunofluorescence.Results1.Compared with wild type mice,hIAPP^+/-transgenic mice had lower insulin sensitivity and higher blood glucose,showing the characteristics of T2DM.The level of glucose utilization in hippocampus of hIAPP^+/-transgenic mice was lower than that of wild type,especially in DG region and CA1 region.2.The exogenous oligomer Aβ₄₂2 was not completely removed in the brain,and the aggregation of Aβ₄₂2 was produced with the prolongation of injection time.Exogenous Aβ₄₂2 and Amylin could interact with each other in the brain,Aβ₄₂promoted the deposition of Amylin,and the increase of Amylin in brain also promoted the deposition effect of Aβ₄₂.3.Injection of exogenous oligomer Aβ₄₂2 did not affect the motor ability of wild and transgenic mice,but affected social recognition ability of mice,olfactory function and olfactory-dependent memory as well as spatial memory.The effects of Aβ₄₂2 were more significant in the hIAPP^+/-transgenic mice.4.Aβ₄₂2 and Amylin can cause changes in the expression mitochondrial function related molecules in hippocampus,degeneration and death of neurons in hippocampal DG region,loss of neural stem cells in hippocampal DG region and SVZ region,and synaptic damage in hippocampal and olfactory region.These changes were more significant when Aβ₄₂were injected into hIAPP^+/-transgenic mice.Conclusion1.In hIAPP^+/-transgenic mice,insulin sensitivity decreased and the level of glucose utilization in hippocampal neurons decreased.2.When Aβ₄₂2 was injected into the hippocampus of hIAPP transgenic mice,Aβ₄₂2 and Amylin were co-located in the hippocampus.Both Aβ₄₂2 and Amylin deposition increased in the hippocampus.3.In hIAPP^+/-transgenic mice,the impairment of cognitive and memory function was more significant after injection of Aβ₄₂2 into hippocampus.4.In hIAPP^+/-transgenic mice,injection of Aβ₄₂2 into hippocampus can cause more obvious changes in the expression of mitochondrial function related molecules in the hippocampus.5.In hIAPP^+/-transgenic mice,injection of Aβ₄₂2 into the hippocampus resulted in more significant synaptic reduction in the hippocampus and olfactory,a decrease in the number of neural stem cells in the hippocampus DG and SVZ region.