Effect of soluble amyloid precursor protein α on nerve cell apoptosis and neurological function in subarachnoid hemorrhage rats / 中华老年心脑血管病杂志

Objective To study the effect of soluble amyloid precursor protein α (sAPPα) on nerve cell apoptosis and neurological function in subarachnoid hemorrhage (SAH) rats.Methods Sixty male SD rats were randomly divided into control group (n=20),SAH+saline group (n=20) and SAH+sAPPα group (n=20).A SAH model was established by injecting autologous blood into cistern magna in rats.After a SAH model was established for SAH + saline group and SAH + sAPPα group by injecting saline and sAPPα respectively into the cistern magna of rats,the apoptotic cells were detected by immunofluorescene with TUNEL staining and the neurological function was scored in 10 rats from each group on day 3 after injection of sAPPα and saline.Results The number of apoptotic cells in brain tissue was significantly greater in SAH+saline group than in control group (P＜0.05) and was significantly smaller in SAH+sAPPα group than in SAH+ saline group (P＜0.05).The neurological function score was 26.7±0.5,13.9±0.7 and 23.0±0.8 respectively in control group,SAH + saline group and SAH + sAPPα group.Conclusion sAPPα alleviates secondary damage of neurological function by inhibiting the apoptosis of nerve cells in rats after SAH and can thus improve their neurological function.

Effects of Shouwu-Yizhi capsule on learning and memory and the expressions of presenilin 1,β-amyloid precursor protein mRNAs in the hippocampus following cerebral ischemia reperfusion in rats / 国际中医中药杂志

Objective To investigate the effects of Shouwu-Yizhi capsule on learning and memory, and the expressions of presenilin 1(PS1),β-amyloid precursor protein (APP) mRNAs in the hippocampus following cerebral ischemia reperfusion in rats. Methods Sprague–Dawley rats were randomly divided into a Shouwu-Yizhi group, a piracetam group, a model group, and a sham operation group with 20 rats in each group. Focal cerebral ischemia reperfusion model was induced by middle cerebral artery occlusion for 2 hours. Seven days after ischemia reperfusion, the rats in the Shouwu-Yizhi and piracetam groups were administered intragastrically Shouwu-Yizhi solution (52 mg/ml) and piracetam solution (28 mg/ml) for 28 days, both in dose of 1 ml/(100 g?d) for 28 days;and the rats in the model and sham operation groups were given intragastrically equivalent volumes of normal saline. Learning and memory were tested using the Morris water maze, and the expressions of PS1 and APP mRNAs were measured using real-time quantitative fluorescent PCR. Results Water maze test showed that the escape latency (12.98±0.70s vs. 9.43±0.78s) was significantly shorter, and the frequency of crossing platform (5.08±0.39 vs. 7.62±0.43) significantly lower in the model groupthan those in the sham operation group; the escape latency (9.77±0.58s vs. 12.98±0.70s) was significantly shorter and the frequency of crossing platform (7.40±0.44 vs. 5.08±0.39) significantly lower in the Shouwu-Yizhi group than those in the model group(all P<0.01). The expressions of PS1 (0.99±0.01 vs. 1.08± 0.03)and APP (1.06±0.03 vs. 1.12±0.04) mRNAs in the hippocampus in the Shouwu-Yizhi group were significantly decreased than those in the model group (P<0.05 or 0.01). Conclusion Shouwu-Yizhi capsule may inhibit the expressions of PS1 and APP mRNAs in the hippocampus, and improve the learning and memory following cerebral ischemia reperfusion in rats.

Effect of 3-n-butylphthalide on expression of matrix metalloproteinase andβ-APP in rats / 中华老年心脑血管病杂志

Objective To study the effect of 3-n-butylphthalide on expression of MMP-2 ,MMP-9 andβ-APP in rats following chronic cerebral hypoperfusion at molecular level .Methods Seventy-two male Wistar rats were randomly divided into sham operation group ,ischemic group ,and bu-tylphthalide group (24 in each group) .Rats in ischemic group and butylphthalide group were di-vided into postischemic 7 ,14 ,30 ,60 d subgroups (6 in each group) .Expressions of β-APP ,MMP-2 , MMP-9 in each group were detected by Western blot .Results The β-APP expression level was significantly higher in postischemic 14 d ,30 d ,60 d subgroups than in postischemic 7 d subgroup (P<0 .05 ,P<0 .01) ,and in postischemic 30 d ,60 d subgroups than in postischemic 14 d sub-group (P< 0 .01) .The β-APP expression level differed greatly in 30 d and 60 d butylphthalide subgroups (P<0 .01) .The MMP-2 expression level was significantly higher in postischemic 60 d subgroup than in postischemic 30 d subgroup (P<0 .05 ,P<0 .01) .Theβ-APP and MMP-2 ex-pression levels were significantly higher in ischemic group and butylphthalide group than in sham operation group (P<0 .05 ,P<0 .01) ,and significantly lower in butylphthalide group than in is-chemic group (P< 0 .01) .Conclusion MMP-2 and β-APP tend to change similarly and are in-volved in blood brain barrier destruction and amyloid deposition .Butylphthalide intervenes in am-yloid deposition by downregulating the expression of MMP-2 andβ-APP .

Changes of Cytokine Expression in the Hippocampus of Aβ1-42-Induced Alzheimer’s Disease Rat Model / 天津医药

Objective To explore changes of expression of pro-and anti-inflammatory cytokines in the hippocam-pus of Aβ1-42-induced Alzheimer’s disease (AD) rat model. Methods Twenty-four SD rats were divided into control group, PBS group (PBS was injected into CA1 area of hippocampus) and AD model group (Aβ1-42 was injected into CA1 area of hip-pocampus). The escape latency was evaluated by Morris water maze in three groups. Nissl staining was used to detect the le-sions of hippocampal CA1 neurons. Levels of amyloid precursor protein (APP) and protein phosphatase 2A (PP2A) in hippo-campus were measured by Western blot analysis. Real-time PCR was employed to examine the expressions of pro-inflamma-tory cytokines, including interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ), and the mRNA expressions of anti-inflammatory cytokines, including IL-4, IL-10 and transforming growth factor-β(TGF-β). Re-sults Rats subjected to Aβ1-42 injection in bilateral hippocampus led to a ability reduction of learning and memory, a loss of neurons in hippocampus and an increase in the expression of APP, and a decrease in PP2A expression in the hippocampus. In AD hippocampus, The mRNA expressions of the pro-inflammatory mediator, IL-1β, TNF-αand IFN-γ, were significant-ly up-regulated, but the expressions of the anti-inflammatory cytokines, IL-4, IL-10 and TGF-β, were markedly down-reg-ulated in AD group compared with those of control and PBS groups. Conclusion The pro-inflammatory/anti-inflammatory imbalance induced neuro-inflammation in AD rats, which was involved in pathogenesis of AD.

Effect of catalpol on senile plaques and spatial learning and memory ability in amyloid-β protein precursor/presenilin 1 double transgenic mice / 中华神经科杂志

Objective To investigate whether catalpol affects senile plaque formation and spatial learning and memory ability in the amyloid-β protein precursor/presenilin 1 (APP/PSI) double transgenic mice.Methods Three month-old APP/PS1 double transgenic mice were randomly divided into catalpoltreated and saline-treated groups (n =10),with C57 mice of the same age and genetic background as normal control group (n =10).The catalpol (in a dose of 5 mg · kg-1 · d-1) and the same amount of saline were peritoneally injected into Alzheimer' s disease (AD) model mice for 3 weeks.Immunohistochemical staining was performed to examine senile plaques in the brain of AD model mice,and Morris water maze was used to assess the spatial learning and memory abilities of the mice.Results Compared with the saline-treated AD model mice (6.0 ±0.6),the number of senile plaques of catalpol treated AD mice significantly decreased (2.3± 0.7; t =3.500,P =0.025); Mice in each groups had similar latency and path length to reach platform in visible platform test; In hidden platform test,catalpol-treated mice had a significant lesser latency and path length compared with saline-treated mice,furthermore,catalpol-treated mice had much more platform-crossing times (6.4 ± 0.8) than saline-treated mice (2.9 ± 0.4 ; t =5.592,P =0.001).Conclusion Catalpol can significantly decrease the senile plaque formation and improve the spatial learning and memory abilities of APP/PS1 double transgenic mice.

Mutation analysis of presenilin-1 gene in Alzheimer' s disease patients and the effects of its mutation on expression of presenilin-1 and amyloid precursor protein / 中华神经科杂志

Objective To analyze the resenilin-1 (PS-1) gene mutations in Alzheimer' s disease (AD) patients and investigate the influence of the initiation codon mutation on the mRNA expression of PS-1 and amyloid precursor protein (APP) genes and the expression of PS-1 proteins.Methods (1) All 111 AD patients were enrolled by the Department of Neurology,Second Affiliated Hospital,College of Medicine,Zhejiang University from July 2004 to June 2010.Mutations in the 13 exons and flanking regions of PS-1 gene were examined by direct sequencing.(2) cDNAs encoding full-length wild-type and mutant (c.1A ＞G) PS-1 were subcloned into enhanced green fluorescent protein.Levels of the mRNA expression of PS-1 and APP genes and PS-1 proteins expression in the transfected cells were detected by quantitative real-time PCR and Western blot,respectively.Results A new heterozygous initiation codon mutation changing from ATG to GTG in one individual was identified.Compared to the control groups,the mRNA expression of the mutant PS-1 gene in HEK293 and N2a was significantly lower than the normal PS-1 gene(116.8 ± 3.9 vs 49.5 ±3.3,t =13.27,P ＜0.01 ;69.0 ± 1.9 vs 29.5 ± 1.3,t =17.20,P ＜0.01) and the APP gene was not obviously altered.The proteins were detected by Western blot analysis in HEK293 cells but not in N2a cells.Conclusions Since we only identified one novel heterozygous initiation codon mutation (from ATG to GTG),mutations in PS-1 are likely to be rare in AD patients.Initiation codon mutation would reduce the expression of PS-1 proteins.Inactivation of some of the PS-1 proteins could be insufficient to lead to AD and could be more likelv to act as a risk factor.

Microglial P2X7 receptor expression is accompanied by neuronal damage in the cerebral cortex of the APPswe/PS1dE9 mouse model of Alzheimer's disease

The possibility that P2X7 receptor (P2X7R) expression in microglia would mediate neuronal damage via reactive oxygen species (ROS) production was examined in the APPswe/PS1dE9 mouse model of Alzheimer's disease (AD). P2X7R was predominantly expressed in CD11b-immunopositive microglia from 3 months of age before Abeta plaque formation. In addition, gp91phox, a catalytic subunit of NADPH oxidase, and ethidium fluorescence were detected in P2X7R-positive microglial cells of animals at 6 months of age, indicating that P2X7R-positive microglia could produce ROS. Postsynaptic density 95-positive dendrites showed significant damage in regions positive for P2X7R in the cerebral cortex of 6 month-old mice. Taken together, up-regulation of P2X7R activation and ROS production in microglia are parallel with Abeta increase and correlate with synaptotoxicity in AD.

Identification of a rare amyloid precursor protein gene mutation in a Chinese family with early-onset familial Alzheimer's disease / 中华神经科杂志

Objective To analyze the phenotype and genatics in a Chinese family with early-onset familial Alzheimer's disease(EOFAD). Methods Peripheral blood were collected in available members in the family and genomic DNA was extracted. PCR-sequencing of exon 16 and exon 17 of the amyloid precursor protein(APP) gene, presenilin 1 (PSEN1), and presenilin 2 (PSEN2) was performed. Results At age 40, two EOFAD patients (siblings) in the family developed an insidious onset of difficulties in memory. One ( Ⅱ3 in the pedigree) showed blinking. The other ( Ⅱ 5 ) showed irritability and bradykinesia.Progressive diffuse coritcal atrophy in bilateral temporal cortex was observed. Moderate diffuse cerebral dysfunction was observed in Ⅱ3 by the electroencephalogram study and neuropsychological assessments.Sequencing revealed that both patients were heterozygous for a mutation c. 2343 G ＞ A in exon 17 of APP,causing the amino acid substitution Val715Met. Four members ( Ⅱ1, Ⅱ 3, Ⅱ 5 and Ⅲ1 ) were homozygous for ApoE ε4 allele. Ⅱ9 was ε2/ε4. Conclusions This study identified a mutation, Val715Met in the APP gene in Chinese patients with EOFAD. We suggest screening for APP gene mutations in Chinese patients with EOFAD.

Toxic effect of carboxyl-terminal peptide of β-amyloid precursor protein (APPC31) on Neuro2a cells / 中华神经科杂志

Objective To investigate the toxic effect of the carboxyl-terminal peptide of β-amyloid precursor protein (APPC31) on Neuro2a cells as well as its role in the toxic process in Neuro2a cells induced by Aβ42 in vitro.Methods The plasmid vector and the APPC31 construct were transiently transfected into Neuro2a cells respectively by lipofectamine 2000.The viability of the cells was measured by the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay at 48 h after transfection,and their morphocytology was observed by 4', 6-diamidino-2-phenylindole (DAPI) nucleus staining.Afterword different constructs including vector, WTAPP695, APP( D664A), the amino-terminal peptide of β-amyloid precursor protein (APP△C31) and APPC31 were transiently transfected into Neuro2a cells respectively via the same method.At 24 h after transfection Aβ42 was added into the culture medium of Neuro2a cells with the desired concentration for another 24 h for cell studies.The viabihty and morphocytology of the cells were measured by using the MTT assay and DAPI nucleus staining, respectively.Results When incubated in the absence of Aβ42, the viability of cells transfected with vector and APPC31construct were 0.81 ±0.10 and 0.88 ±0.12 respectively, and accordingly there was no significant difference between the these two groups (t = - 0.78, P = 0.48 ); meanwhile no obvious cell nuclear morphological changes of apoptosis or death occurred.However when incubated in the presence of Aβ42, the viability of cells transfected with vector, WTAPP695, APP( D664A), APP△C31 and APPC31 constructs were 0.82 ±0.01, 0.78 ±0.03, 0.55 ±0.04, 0.81 ±0.04, 0.78 ±0.02 and 0.54 ±0.02 respectively.The viability of cells transfected with WTAPP construct and APPC31 construct decreased significantly ( F = 47.53, P ＜0.05) compared with the control group, meanwhile cells displayed condensed nuclear and even nuclear fragmentation.Conclusions In vitro, over-expression of a certain level of APPC31 in Neuro2a cells fails in causing cell death, but this short peptide enhances cytotoxicity induced by Aβ42 in Neuro2a cells.Thus,these results provide the experimental basis for the further explication of the pathogenesis of Alzheimer's disease.

Transgenic mice of Alzheimer's disease: developing and preliminary identification of transgenic mice harboring mutant amyloid precursor protein gene fused with two types of fluorescent protein genes / 中华神经科杂志

Objective To develop transgenic mice harboring the fusion gene of mutant amyloid precursor protein and two types of fluorescent protein for the future study on Alzheimer's disease.Methods The fusion gene CFP-54 bp-YFP-C99 was introduced into mice by mieroinjection.The presence of CFP-54 bp-YFP-C99 was confirmed by PCR in the founders.Results CFP-54 bp-YFP-C99 gene was injected into pronucleus of 2202 zygotes and 1806 injected eggs were implanted into 56 foster mothers, 13 of which were pregnant.There were 13 foster mothers who borne 52 offspring and 32 of them survived.Recipient mouse pregnancy rate was 23.2% (13/56) and the integration rate was 3.9% (2/52).Conclusion CFP-54 bp-YFP-C99 transgenic mice is obtained, but the transgenic efficiency is low.

Identification of PSEN1 and APP Gene Mutations in Korean Patients with Early-Onset Alzheimer's Disease

Although mutations in three genes, amyloid precursor protein (APP), presenilin 1 (PSEN1), and presenilin 2 (PSEN2), have been identified as genetic causes of earlyonset Alzheimer s disease (EOAD), there has been a single report on a PSEN1 mutation in Koreans. In the present study, we performed a genetic analysis of six Korean patients with EOAD. Direct sequencing analysis of the APP, PSEN1 and PSEN2 genes revealed two different mutations of the PSEN1 gene (G206S and M233T) and one mutation of the APP gene (V715M) in three patients with age-atonset of 34, 35, and 42 yr, respectively. In addition, two patients with age-at-onset of 55 and 62 yr, respectively, were homozygous for APOE epsilon 4 allele. One woman had no genetic alterations. These findings suggest that PSEN1 and APP gene mutations may not be uncommon in Korean patients with EOAD and that genetic analysis should be provided to EOAD patients not only for the identification of their genetic causes but also for the appropriate genetic counseling.

Caveolin-1 upregulation in senescent neurons alters amyloid precursor protein processing

Lipid rafts provide a platform for regulating cellular functions and participate in the pathogenesis of several diseases. However, the role of caveolin-1 in this process has not been elucidated definitely in neuron. Thus, this study was performed to examine whether caveolin-1 can regulate amyloid precursor protein (APP) processing in neuronal cells and to identify the molecular mechanisms involved in this regulation. Caveolin-1 is up-regulated in all parts of old rat brain, namely hippocampus, cerebral cortex and in elderly human cerebral cortex. Moreover, detergent-insoluble glycolipid (DIG) fractions indicated that caveolin-1 was co-localized with APP in caveolae-like structures. In DIG fractions, bAPP secretion was up-regulated by caveolin-1 over-expression, which was modulated via protein kinase C (PKC) in neuroblastoma cells. From these results we conclude that caveolin-1 is selectively expressed in senescent neurons and that it induces the processing of APP by beta-secretase via PKC downregulation.

Experimental studies on the expression of amyloid ?-protein precursor in early stage of diffuse axonal injury in rats / 解放军医学杂志

s Objective To observe the changes in the expression of ?-amyloid precursor protein (?-APP) in selected brain areas in early stage of diffuse axonal injury (DAI) in rats. Methods DAI was induced in SD rats by a self-made rotating injury device. 16 rats in injury group were sacrificed by decapitation at 3 hours and 6 hours post-trauma, while the other 8 rats in sham injury group were put to death after 24 hours. The samples of the rat brain were stained with ?-APP for immunohistochemistry in order to detect the early stage pathological changes in subcortex white matter, hippocampus, corpus callosum and brain stem at different time points. Results The ?-APP expression was positive at 3 hours and was particularly strong at 6 hours after injury. The results of semi-quantitative analysis showed a significant differentiation of 3 hours and 6 hours in the immuno-positive expression of brain stem, corpus callosum and hippocampus. Histological results demonstrated that DAI occurred in all rats of injury group. Conclusion ?-APP is a kind of sensitive and useful immunochemical marker for early DAI diagnosis, and it indicates axonal lesion or breakage.

Alzheimer disease associated phenotypes in PDAP_(SW) transgenic mice / 中华神经科杂志

Objective In order to study the role of APP SW mutation in the aetiology of Alzheimer′s disease(AD), the phenotypes of the PDAP SW transgenic mice were investigated. Methods Body weight, coat color and reproducibility of transgenic mice were observed;The pathological changes in the brain of the transgenic mice were examined using immunohistochemistry; Behavioral changes of transgenic mice were examined by Y-maze. Result At 3 month′s age, the mean body weight of the transgenic mice was (31.0?3.7) g, that of non-transgenic was (34.0?2.9) g;while the mean body weight of F1 transgenic mice was (32.0?3.3) g, of negative mice was (31.0?4.2) g. There were no significant difference between these two groups. There were three transgenic mice with abnormal coat and one male was infertile. The transgenic mice had amyloid deposit in their brains. In the Y-maze test, transgenic mice showed an increased number of arm entered (53?7 versus 37?4) and an impaired spontaneous alternation. The frequency of alternation was 48.2% in the transgenic mice and 76.4% in the non-transgenic mice,respectively.Conclusion The PDAP SW transgenic mice had typical pathological and behavioral changes similar to that of AD and could be used as an animal model for studying AD.

Change of ?-amyloid precursor protein processing in platelet of Alzheimer's disease patients / 中华神经科杂志

Objective To investigate the characteristic of ?-amyloid precursor protein (A?) processing in activated platelet in AD.Methods Thirty-six sporadic AD patients and 30 control subjects were included in this study.Blood was collected from the subjects to separate platelets.After treated by thrombin,the soluble amyloid precursor protein (APP) level in the snpernatants of platelets from 36 were analyzed by means of western blot with a specific antibody recognizing soluble APP.Meanwhile A? level was measured by radioimmunoassay.Results After treated with thrombin,the level of soluble APP in the supernatants of platelets in patients with AD decreased by 31.0% (P

The possible mechanism of insulin reducing amyloid beta peptide in the brain cortex of diabetic rats / 中华神经科杂志

Objective To investigate the reduction of amyloid beta peptide (A?) production induced by insulin in the cortex of diabetic rat. Methods 15 male SD rats were randomly divided into control group, diabetes group, and diabetes plus insulin group. The diabetic rats were induced by streptozotocin. The activity of glycogen synthase kinase-3 (GSK-3) was measured by 32P liquid scintillography for incorporated radioactivity in control group, DM group, DM+insulin group, the production of A? was determined by sandwich ELISA in each group, and the expression of APP was determined by Western-blot. Results In control group, the activity of GSK-3 (1.04?0.11), the production of A? (A?_ 40 (40.92?5.34) pg/?l, A?_ 42 (29.64?3.19) pg/?l)and the levels of full-length APP(1.05?0.08) was low, but in DM group, the activity of GSK-3 (2.02?0.12) and the production of A?(A?_ 40 (67.53?11.69) pg/?l, A?_ 42 (45.02?4.10) pg/?l) increased (P

Effect of ginsenoside Rg1 on the amyloid protein precursor and neprilysin expression induced by lipopolysaccharide in C6 cell line / 中国病理生理杂志

AIM: This study was designed to investigate the effect of ginsenoside Rg1 on the amyloid ?-protein precursor (APP) and neprilysin (NEP)expression induced by lipopolysaccharide (LPS) in C6 cell line in order to discover effectual Alzheimer's disease (AD)-treated drugs. METHODS: MTT colorimetric analysis was used to measure the survival rate of C6 cultured with ginsenoside Rg1 at different concentrations (2 5, 5, 10 and 20 ?mol?L -1) and LPS (100 mg?L -1). The expression of APP and NEP mRNA was measured by RT-PCR. RESULTS: LPS decreased the survival rate of C6, furthermore, the increase in APP expression and the decrease in NEP expression were observed. On the other hand, the above alteration induced by LPS was reversed by ginsenoside Rg1. CONCLUSION: This study demonstrates that LPS can cause cell damage, the increase in APP expression and the decrease in NEP expression. Ginsenoside Rg1 can exert a neuroprotective action, protect C6 cells against LPS-induced injury via inhibiting APP expression and increasing NEP expression.