| Background and objectAlzheimer’s disease (AD) is the most prevalent neurodegenerative disease causing dementia in the elderly, whose typical clinical symptoms include the progressive and irreversible memory loss and cognitive decline. It has been well documented that the pathological manifestations include extracellular amyloid-β protein (Aβ) deposition, intracellular tau deposition as neurofibrillary tangles as well as neuronal and synaptic loss with region-specific brain atrophy. However, there is no effective method to cure AD.To date, the etiology of AD is not clear. However, extracellular Aβ deposition is considered to play a pivotal role in the progress of AD pathogenesis. There is a close correlation between Aβ deposition and neuronal loss as well as impaired integrity of brain extracellular matrix (ECM). Augmented evidences demonstrate that there are two neurogenic regions in the adult central nervous system:the subventricular zone (SVZ) and the subgranular zone (SGZ) of dentate gyrus (DG) in the hippocampus. And endoneurogenesis exits thoughout the whole life. However, the proliferation and/or neuronal differentiation of neural stem cells (NSCs) are impaired in human AD or AD animal models because of inflammatory microenvironment in the brain. More and more evidences suggest that endoneurogenesis play an important role in hippocampal-dependent learning and memory. Increasing evidences demonstrate that strategies to augment the endoneurogensis or neural stem cell replacement for neuronal loss maybe the promising method to improve the impaired cognition and prevent the pathophysiological progress of AD animal model. But the ratio of neuron differentiation was low after transplantation of NSCs into AD animal model. In addition, most cells became glial cells after transplantation. Therefore, it is an emergency to find a safe and effective chemical owning the properties of promoting the proliferation and neuronal differentiation of endogenous neural stem cells as well as restoring the integrity of impaired ECM, which maybe a promising strategy to prevent or treat AD.Under physical conditions, the self assembling peptide amphiphile (PA) with an IKVAV (isoleucine-lysine-valine-alanine-valine) epitope from laminin (IKVAV-PA) can be triggered into three dimensional nanostructures, which can amplify the IKVAV epitope density relative to a lamini monolayer to103times. IKVAV-PA can be resembled into the collagen fibrils, one of ECM, in vivo to replace the impaired or lost ECM. It has been demonstrated that IKVAV-PA can be resembled into three-dimensional cell networks in vitro and promote the proliferation as well as rapidly neuronal differentiation of NSCs. It has been proposed that KVAV-PA promote the functional improvement of injured spinal cord via inhibiting glial scar formation and promoting axon alongation. In addition, IKVAV-PA can be metabolized into single amino acid as the nutrition for cell growth after2weeks of injection in vivo. Could IKVAV-PA have the effect on the cognition and the pathophysiological progress of AD animal model? Could IKVAV-PA improve the impaired the structure of ECM and promote the proliferation and neuronal differentiation of NSCs in AD?Therefore, in the current study, we used male heterozygous APP/PS1double transgenic (Tg) mice as AD animal model. We try to investigate whether IKVAV-PA has the effect on the impaired cognition and the pathophysiological progress of APP/PS1mice as well as potential mechanism after2weeks and4weeks of stereotaxic injection into the bilateral hippocampus.Methods1. Male APP/PS1aged6months are used as AD animal model and randomly divided into2groups:one group receiving4μl1%IKVAV-PA as IKVAV-PA-treated group; the other group receiving4μl NS (0.9%saline) as vehicle-treated group. Male C57/BL6(wild type, WT) aged6months are used as negative control group and receive4μl NS.2. Morris water maze (MWM) is performed to test the ability of learning and spatial memory after1week and3weeks of IKVAV-PA injection.3. After MWM, ELISA is performed to test the levels of soluble Aβ1-42and Aβ1-40in the cortex and hippocampus of APP/PS1Tg mice. Thioflavin S staining is performed to measure the size of Aβ plaques in the cortex and hippocampus of APP/PS1Tg mice. Immunohistochemistry is for labeling Aβ1-42, Brdu-positive cells, DCX-positive cells as well as Brdu and DCX double-labeled cells in the cortex and hippocampus of APP/PS1Tg mice. Western Blot is performed to measure the protein levels of APP, p-Tau, p-ERK1/2and ERK1/2in the cortex and hippocampus of APP/PS1Tg mice. Real time PCR is for testing the gene expression of BACE1, IDE and NEP in the cortex and hippocampus of APP/PS1Tg mice.Results1. The effect of IKVAV-PA on the cognition and the pathophysiological progress of APP/PS1Tg mice as well as the potential mechanisms after4weeks of injection.1.1The performance in MWM of IKVAV-PA-treated APP/PS1Tg mice were significantly improved compared to vehicle-treated APP/PS1Tg mice.During the last3days of spatial acquistion, IKVAV-PA-treated Tg mice and vehicle-treated WT mice exhibited significantly decreased the escape latencies and pathway to locate the hidden platform compared to vehicle-treated Tg mice. More importantly, during the spatial acquisition trial, both vehicle-treated WT mice and IKVAV-PA-treated Tg mice exhibited the significant spatial bias for target quadrant as opposed to three non-target quadrants, vehicle-treated Tg mice did not show such spatially biased performance. However, there is no significant difference on the escape latencies and pathway to locate the hidden platform between IKVAV-PA-treated Tg mice and vehicle-treated WT mice. In addition, compared to vehicle-treated Tg mice, vehicle-treated WT mice and IKVAV-PA-treated Tg mice have significantly more time in the target quadrant and significantly increased number of crossing the previous platform location during the probe trial. Although IKVAV-PA-treated Tg mice exhibit less time in the target quadrant and number of crossing the previous platform location, there is no significant difference between two groups. In addition, there is no significant difference in the swimming speed among three groups.1.2IKVAV-PA augmented the proliferation and neuronal differentiation of endogenous neural stem cells.The number of Brdu-labeled proliferating cells and DCX-labeled new neurons in the dentate gyrus of IKVAV-PA-treated Tg mice were significantly increased compared to those of vehicle-treated Tg mice. In addition, we found that the new neurons of IKVAV-PA-treated Tg mice were more well-organized than those of vehicle-treated Tg mice. We also observed that more Brdu-labeled cells in the dentate gyrus of IKVAV-PA-treated Tg mice were double-labeled with DCX.1.3IKVAV-PA had effect on the pathophysiological progress of APP/PS1Tg mice.As assessed by Thioflavin S staining, our data showed that the total areas of Aβ plaques in the cortex and hippocampus of IKVAV-PA-treated APP/PS1Tg mice were significantly reduced compared to those of vehicle-treated APP/PS1Tg mice. In addition, we observed that IKVAV-PA significantly reduced the soluble levels of Aβ1-40and Aβ1-42, as measured by ELISA, in the cortex and hippocampus.1.4IKVAV-PA activated the ERK1/2cell signal pathway and increased the expression of p-ERKl/2.Statistical analysis uncovered that IKVAV-PA had no effect on the total protein levels of ERK1/2in the cortex and hippocampus of APP/PS1Tg mice. However, the protein levels of p-ERK1/2in the cortex and hippocampus of IKVAV-PA-treated Tg mice were significantly improved compared to those of vehicle-treated Tg mice.2. The effect of IKVAV-PA on the cognition and the pathophysiological progress of APP/PS1Tg mice as well as the potential mechanisms after2weeks of injection.2.1IKVAV-PA significantly improved the performance in MWM.During the last day of spatial acquisition, IKVAV-PA-treated Tg mice exhibited significantly reduced escape latencies and pathway to locate the hidden platform compared to vehicle-treated Tg mice. However, there is no significant difference on the escape latencies and pathway to locate the hidden platform between IKVAV-PA-treated Tg mice and vehicle-treated WT mice. In addition, compared to vehicle-treated Tg mice, vehicle-treated WT mice and IKVAV-PA-treated Tg mice have significantly more time in the target quadrant and significantly increased number of crossing the previous platform location. Although IKVAV-PA-treated Tg mice exhibit less time in the target quadrant and number of crossing the previous platform location, there is no significant difference between two groups. In addition, there is no significant difference in the swimming speed among three groups.2.2IKVAV-PA prevented the pathophysiological progress in the brain of APP/PS1Tg mice.Statistical analysis suggested that the total area of Thioflavin S staining-positive Aβ in the hippocampus of IKVAV-PA-treated Tg mice was significantly reduced compared to those of vehicle-treated Tg mice. However, there was no significant difference in the total area of Thioflavin S staining-positive Aβ in the cortex of two groups. In addition, we found the same tendency on the levels of Aβ1-42, measured by immunohistochemistry staining. IKVAV-PA significantly reduced the levels of soluble Aβ1-40and Aβ1-42in the cortex and hippocampus of APP/PS1Tg mice.2.4IKVAV-PA altered the processing of APP in the brain of APP/PS1Tg mice.Statistical analysis shed light on that IKVAV-PA has no effect on the output of APP measured by Western Blot. The expressions of BACE1, IDE and NEP were tested by the real time PCR. The data showed that IKVAV-PA significantly decreased the gene expression of BACE1and significantly increased the gene expressions of IDE and NEP in the cortex and hippocampus of APP/PS1Tg mice. In addition, we found that IKVAV-PA had no effect on the output of p-Tau measured by Western Blot, in the cortex and hippocampus of APP/PS1Tg mice.Conclusion1. IKVAV-PA could attenuate the impaired cognition and the total areas of Aβ plaque and soluble Aβ1-40and Aβ1-42of AD animal model via augmenting the proliferation and neuronal differentiation of the endogenous neuronal stem cells because of activation of ERK1/2in the brain.2. IKVAV-PA could improve the impaired cognition of AD animal model and attenuate the total areas of Aβ plaque and soluble Aβ1-40and Aβ1-42via altering the processing of APP after2weeks of injection.3. IKVAV-PA, as one of biomaterials, maybe a promising agent to treat AD. |
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