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Aim To explore mechanism of epigallocatechin-3-gallate (EGCG) on alleviation of hippocampal neuronal autophagy in APP/PSI transgenic mice. Methods 8-month old APP/PSI transgenic mice were randomly divided into three groups;model group (Tg), EGCG low dose group (Tg/EGCG-L), high dose group (Tg/EGCG-H). C57BL/6J mice were utilized as control. Learning and memory were detected by Morris water maze test. The hippocampal ULK1, P62, LC3 I I / LC3 I,mT0R and Aß M2 expressions were detected by Western blot, immunohistochemical staining and ELISA. Results Compared with NT mice, Tg mice showed a marked prolongation of the escape latency in MWM test (P <0. 05). Decreased ULK1 expression and increased P62, LC3 II/LC3 I and A ßM 2 were detected (P < 0. 05). EGCG-treated group showed marked improvement of all these abnormal changes (P < 0. 05). Conclusions EGCG treatment is able to improve cognitive function, which may be attributed to ameliorated autophagic networks dysfunction and reduced Aß plaques in the the hippocampi of APP/PS1 transgenic mice.
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Objective To investigate the expression and relationship of canonical transient receptor potential channel-3 (TRPC3) and brain-derived neurotrophic factor (BDNF) in the hippocampus of a rat model of Alzheimer's disease (AD). Methods SD rats were randomly divided into PBS, AD, and AD+BDNF experimental groups. AD models were generated by intracerebroventricular injection ofβ-amyloid protein (Aβ1-42). BDNF was injected via the lateral ventricle catheter after 14 days. The Morris water maze test was used to assess the spatial learning and memory ability of the rats. The expression of TRPC3 and BDNF mRNA and protein in the hippocampus were detected by RT-PCR and Western blotting, respectively. Results The Morris water maze test showed that the escape latencies of the fifth day in the AD group were longer than those in the PBS group (P ＜ 0. 05). The escape latencies in the AD+BDNF group were shorter than those in the AD group (P ＜ 0. 05). RT-PCR and Western blotting results showed that the expression of both TRPC3 and BDNF were reduced in the AD group compared with the PBS group (P ＜ 0. 05). TRPC3 expression was increased in the AD+BDNF group compared with the AD group (P ＜ 0. 05). Conclusion The expression of BDNF and TRPC3 is decreased in the hippocampus of AD rats. An exogenous BDNF injection appears to improve the spatial learning and memory of AD rats that are impaired by a Aβ1-42 injection, possibly via TRPC3 upregulation, and may play a protective role in neurons.
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Objective To investigate the distribution of divalent metal transporter 1 (DMT1) in the cerebellum of APP/PS1 transgenic mouse. Methods Immunohistochemistry, immunofluorescence, and confocal laser scanning microscopy were used to analyze the relationship between DMTl and amyloid beta (Aβ) and their distribution in senile plaques. Western blotting was used to analyze DMT1 protein level in the APP/PS1 transgenic mouse cerebellum. Results DMTl and Aβ were mainly located in the amyloid plaques, which were predominately located in the molecular layer of the cerebellar cortex of the transgenic mouse. Only a few plaques could be seen in the Purkinje cell layer and granular layer. Confocal laser microscopy revealed the DMTl and Aβ were co-localized in senile plaques. Conclusion The abundant expression of DMTl protein suggests that DMTl and the divalent metal ions that it transports might be involved in the formation of Aβ senile plaques and other pathological processes in the cerebellum in Alzheimer' s disease.
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Amyloid beta-peptides ( Aβ) is the key pathological feature of Alzheimer’s Disease (AD). Various factors contrib-ute to the accumulations of Aβ in the brains of patients. Among them, blood-brain barrier ( BBB) plays a crucial role in trans-porting Aβ between the brain and the bloodstream while this transfer function is mediated by the receptor of Aβon BBB. The abnormality of Aβ transport and related receptor expression can be detected in the brains of patients with AD, resulting in an un-usual increase in Aβlevels unusually increased . This review e-laborates the structure and function of BBB, the transport of Aβand the expression and transport mechanism of the related recep-tor, as well as summarizes the possible clearance strategy of Aβacross the BBB.
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Aim To investigate the protective effect of Panax Notoginseng Saponins ( PNS ) on Aβ25-35-in-duced apoptosis in PC12 cells and molecular mecha-nism. Methods The cell viability of PC12 cells was detected by MTT assay. The levels of LDH leakage, ROS,MDA,Caspase-3 activity and antioxidant enzyme activities of SOD, CAT, and GSH-Px activity were de-termined by respective kits. The apoptosis of cells was decteced by Western blot. Results PNS could signifi-cantly inhibit the decrease of cell viability and LDH leakage, reduce the production of MDA and ROS( P<0. 01), increase the activity of SOD,CAT and GSH-Px ( P <0. 01 ) , and the mitochondrial membrane poten-tial, inhibit the activation of caspase-3 ( P <0. 01 ) in PC12 cells which were induced by Aβ25-35 . PNS could incrase nuclear Nrf2 and up-regulate HO-1 . The neu-roprotective of PNS could be inhibited by HO-1 inhibi-tor ZnPP. Conclusion PNS may inhibit Aβ25-35-in-duced oxidative stress and apoptosis in PC12 cells by activating Nrf2/HO-1 signaling pathway.
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Aim To investigate the effects of neurotro-phin-3 ( NT-3 ) gene overexpression on the differentia-tion into cholinergic neuron of neural stem cells ( NSCs) in vitro and its underlying mechanism. Meth-ods Brain-derived NSCs from newborn mice were iso-lated and cultured in vitro and determined by immuno-fluorescence. The NSCs were divided into three groups: NSCs, GFP-NSCs and NT-3-NSCs groups. The expression of NT-3 was detected by immunofluo-rescence and ELISA. Then, the ability of NSCs on dif-ferentiation into cholinergic neuron was detected by im-munofluorescence and RT-PCR, and the Acetylcholine Assay Kit was used for acetylcholine ( ACh) , and the expression of Hes1 , Mash1 and Ngn1 mRNA was de-termined by RT-PCR. Results The neurosphere dis-played Nestin and Sox 2-postive by immunofluores-cence, suggesting that the cultured cells were NSCs. The proportion of ChAT immunopositive cells was sig-nificantly higher in the NT-3-NSCs group than that in the other two groups ( P <0. 01 ) . Ach secretion in NT-3-NSCs was significantly elevated compared with the other two groups ( P <0. 01 ) . NSCs transfected with NT-3 increased the levels of Mash1 and Ngn1 mR-NA, and decreased the level of Hes1 mRNA ( P <0. 05 ) . Conclusion NT-3 can significantly promote the in vitro differentiation of NSCs into cholinergic neu-rons via probablly inhibiting Notch signaling pathway.
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Aim To explore the protective effects of lithium chloride ( LiCl ) on neurous injuries and phos-phorylation of tau protein at serine262 induced by okada-ic acid( OA) . Methods The neuroblastoma SK-N-SH cells were differentiated by all-trans-retinoic acid ( AT-RA) . The differentiated SK-N-SH cells were treated with OA to establish the Alzheimer′s disease cellular model. SK-N-SH cells′ viability and proliferation were measured by SRB test. Giemsa staining was used to observe cell morphology. The neurite length of SK-N-SH cells was measured by Image-Proplus software. Syn-aptophysin and phosphorylated tau protein at serine262 expression levels were tested by Western blot. Results The SK-N-SH cells which were treated with 10 μmol ·L-1 ATRA for 7 days displayed mature neuronal fea-tures. The synaptic length of SK-N-SH cells became longer. And the levels of serine262 phospho-tau was sig-nificantly elevated. 20~100 nmol·L-1 OA effectively inhibited the viability of differentiated SK-N-SH cells in a concentration-dependent manner and in a time-de-pendent manner. The OA treatment induced obvious synaptic atrophy in differentiated SK-N-SH cells. And the phosphorylation level of tau protein serine262 also greatly increased. The pretreatment with 10 mmol · L-1 LiCl significantly ameliorated the synaptic atrophy, the decrease of synaptophysin expression and the in-crease of tau phosphorylation at serine262 induced by OA in differentiated SK-N-SH cells. Conclusion LiCl could effectively inhibit OA-induced synaptic atro-phy in differentiated SK-N-SH cells, and it could also result in the increase of synaptophysin expression and the decrease of the phosphorylation of tau protein at serine262 .
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Aim Tongluoxingnao effervescent tablets ( TLXNET) ,based on the ancient formula of QiongGui Tang, can improve cognitive dysfunction in different AD models. This research is aimed to study the effects of TLXNET on the Aβmetabolism and explore the anti-AD mechanism in SH-SY5 Y-APP and SH-SY5 Y-C99 cells. Methods Cells were incubated for 48h in dif-ferent concentrations of medicated serum ( containing 0% ~40% of TLXNET in the serum ) . Firstly, the non-toxic concentration was measured by MTT assay, the activity of cells was detected by LDH methods, and ELISA was used to measure the levels of Aβ1-40 and Aβ1-42 . Then, the gene and protein expressions of APP were detected via RT-PCR and Western blot of which the expressions of shearing fragments such as sAPPα and sAPPβ were investigated by Western blot, the same as the expressions of IDE and NEP. Addi-tionally, the level of mRNA, protein and activity of BACE1 were measured by qRT-PCR, Western blot and fluorescence detection kits respectively. Eventually, MTT assay was performed to detect the cell viability after the SH - SY 5 Y cells were treated with various concentrations of medicated serum and Aβ25-35 for 48h. Results There was no significant toxicity of TLXNET medicated serum in SH-SY5 Y-APP and SH-SY5 Y-C99 cells ( P >0 . 05 ) . TLXNET could signifi-cantly inhibit Aβ secretion in SH-SY5 Y-APP cells ( P0. 05). Additionally, TLXNET could still notably inhibit the expression of sAPPβ pro-tein in a dose-dependent way, with statistical signifi-cance ( P <0. 05 ) . Meanwhile, the level of mRNA, protein and activity of BACE1 were also significantly decreased by TLXNET (P <0. 01). Moreover, the medicated serum of TLXNET had a protective effect on SH-SY5 Y apoptosis induced by Aβ25-35 ( P <0 . 01 ) . Conclusion TLXNET could obviously inhibit β-secretase enzyme, and has an antagonistic effect a-gainst Aβneurotoxicity, which suggests that the inhibi-tion of β-secretase enzyme and antagonism against Aβneurotoxicity are the main anti-AD mechanism of TLX-NET .
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Objective To observe the protective effect of the traditional Chinese medicine prescription, Jiu Nao Yi Zhi water extract, on human neuroblastoma SH-SY5Y cell line, its effect on expression of insulin signal transduction pathway, and to explore the related mechanisms.Methods SH-SY5Y cells cultured in vitro, were divided into control group, Jiu Nao Yi Zhi No.1 prescription group and No.3 prescription group.The doses were 0.0625 mg/mL, 0.125 mg/mL, 0.25 mg/mL, 0.5 mg/mL and 1 mg/mL.The thiazolyl blue ( MTT) metabolic rate of each group was determined.The dose of 0.125 mg/mL was chosen for cell immunofluorescence analysis, and to observe the expression of insulin receptor substrates-1 ( IRS-1 ) , cAMP response element binding protein ( CREB ) , and the factors of insulin signal transduction pathway.Results Compared with the control group, MTT metabolic rates of the Jiu Nao Yi Zhi groups were significantly increased (P<0.05), and the cell morphology was much better in those groups, cell body more plump, well-adherent and neurite extensions were observed.The expressions of IRS-1 and CREB were higher than that in the control group.Conclusions The traditional Chinese medicine prescription Jiu Nao Yi Zhi water extract can protect neurons by promoting nerve cell growth, and improving the expression of IRS-1 and CREB, the factors of insulin signal transduction pathway.
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Alzheimer’ s disease ( AD) , the most prevalent type of neurodegenerative disease, characterized by extracellular β-amyloid ( Aβ) plaque deposition, intracellular neurofibrillary tangles, and the loss of diverse neurons. Neuron loss might be the main cause that induces irreversible decline of cognitive function in AD patients. At present, AD therapy only relieves symptoms instead of fundamentally affecting the major pathologi-cal characteristics of the disease. Recently, rapid advances in neurogenesis and stem-cell biology have provided a new and pro-spective potential for AD treatment. Stem-cell types in the treat-ment of neurodegenerative diseases include neural stem cells ( NSCs ) , embryonic stem cells ( ESCs ) , mesenchymal stem cells ( MSCs ) , and induced pluripotent stem cells ( iPSCs ) . Stem cell therapy shows anti-AD function by replacing the dam-aged and lost neurons, rebuilding the cellular loop, inhibiting amyloidogenesis, promoting neurotrophic factor release, and reg-ulating immune reaction. This review highlights the recent pro-gress, mechanisms, and preclinical evidence of exogenous trans-planted stem cell, and addresses the current major challenges of stem cell transplantation in clinic. In addition, this review also summarizes the research status of compound promoting endoge-nous neurogenesis in brief.
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Aim To investigate the effect of progester-one ( PROG) in protecting the neurons against impair-ment induced by the Aβ1-42 activated astrocytes, and the underlying molecular mechanism. Methods The astrocytes were divided into 5 groups: control, Aβ, and Aβplus PROG groups treated with 3 different con-centrations of progesterone for 24h. Then, Aβand pro-gesterone were removed, and neurons were co-cultured with the treated astrocytes. MTT assay was used to e-valuate the viability of cultured neurons; ELISA was used to detect the levels of IL-1βand TNF-αin culture media of astrocytes; immunofluorescence and Western blot were performed to detect the activation of NF-κB in astrocytes. Results PROG dose dependently pro-tected against Aβ1-42 activated astrocytes induced via-bility decrease in co-cultured neurons. Aβ induced release of IL-1β and TNF-α from astrocytes, and in-crease of NF-κB activity was abolished by progesterone treatment. Conclusion PROG protects the neurons through inhibiting the reactivity of astrocytes, and the underlying mechanism involves the NF-κB signal trans-duction.
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Objective To investigate the relationship between activation of gliacytes , mitogen-activated protein kinase (p38MAPK) and neuronal apoptosis after microinjecting aggregated Aβ25-35 into hippocampus.Methods The model was established by using stereotaxic technique to inject 10μg aggregated Aβ25-35 into dorsal hippocampus in rats .The rats were grouped as the control , vehicle and model groups .Immunohistochemistry and Western blotting were used for detection of activation of microglia(MG), atrocytes (AS) and expression of p-p38MAPK in the hippocampus.ELISA was used to evaluate the level of TNF-αand IL-1β.The survival neurons were observed by Nissl staining and the apoptotic neurons were identified by tunnel staining .Results Expression of ox-42, GFAP, p-p38MAPK were up-regulated in hippocampus, as well as TNF-α、IL-1β, which reached a highest value on the 7th day after injection of Aβ25-35.However, the number of neuron with Nissl positive decreased gradually , and the tunnel positive neurons increased highly and reached a peak value on the 7th day.There were significant differences between the control and vehicle group ( P <0.01). Conclusion Apoptosis of the neuron caused by Aβ25-35 injection may result from activation of gliacytes , p38 MAPK and increase of TNF-αand IL-1βlevel.