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Alzheimer's disease (AD) is a leading cause of dementia in the elderly. Mitogen-activated protein kinase phosphatase 1 (MKP-1) plays a neuroprotective role in AD. However, the molecular mechanisms underlying the effects of MKP-1 on AD have not been extensively studied. MicroRNAs (miRNAs) regulate gene expression at the post-transcriptional level, thereby repressing mRNA translation. Here, we reported that the microRNA-429-3p (miR-429-3p) was significantly increased in the brain of APP23/PS45 AD model mice and N2AAPP AD model cells. We further found that miR-429-3p could downregulate MKP-1 expression by directly binding to its 3'-untranslated region (3' UTR). Inhibition of miR-429-3p by its antagomir (A-miR-429) restored the expression of MKP-1 to a control level and consequently reduced the amyloidogenic processing of APP and Aβ accumulation. More importantly, intranasal administration of A-miR-429 successfully ameliorated the deficits of hippocampal CA1 long-term potentiation and spatial learning and memory in AD model mice by suppressing extracellular signal-regulated kinase (ERK1/2)-mediated GluA1 hyperphosphorylation at Ser831 site, thereby increasing the surface expression of GluA1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). Together, these results demonstrate that inhibiting miR-429-3p to upregulate MKP-1 effectively improves cognitive and synaptic functions in AD model mice, suggesting that miR-429/MKP-1 pathway may be a novel therapeutic target for AD treatment.
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Objective To investigate the role of protein O-linked mannose β1,2-N-acetylglucosaminyltransferase 1 (POMGnT1) in the pathogenic process of Alzheimer's disease (AD).Methods The expression of POMGnT1 gene was examined in AD model cells (N2a/amyloid-precursor protein (APP) 695swe,n=3) and N2a/wt cells (n=3) using real-time PCR and Western blotting.This expression was also examined in AD model mice (APP/PS1,n=3) and wild-type mice (n=3) using immunofluorescence staining.Amyloid β-protein (Aβ) were examined in POMGnT1-gene-knockout mice (n=3) and wild-type mice (n=3) using immunochemistRy.Results The expression of POMGnT1 gene decreased in mRNA and protein levels in N2a/APP695swe cells compared to N2a/wt cells (mRNA:0.80±0.02 vs 1.00,t=10.52,P＜0.01;protein:0.50±0.02 vs 1.31±0.04,t=18.64,P＜0.01).Immunofluorescence results showed the reduced expression of POMGnT1 protein in neurons of APP/PS1 mice.Immunochemistry results showed more Aβ deposits in POMGnT1-gene-knockout mice (2 months old:0.358±0.014 vs 0.048±0.001,t=22.58,P＜0.01;1 year old:0.266±0.004 vs 0.229±0.003,t=7.771,P=0.002).Conclusion These findings suggest POMGnT1 gene may play an important role in the pathogenic process of AD.
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Starting from the characteristics of nervous system,breaking the traditional subject-centered teaching model,combining the nervous system-related basic and clinical courses,Chongqing Medical University has established a nervous-module organ-system integration curriculum since 2010.This new teaching model is introduced and practiced in 5-year outstanding medical class of Chongqing Medical University.The new integrated curriculum guides students to learn knowledge from points to surface,helps them to combine knowledge vertically and horizontally,and simplifies the duplicate teaching content,so as to promote students to develop coherent and innovative thinking mode.Integrated curriculum pattern is an inevitable trend in medical education reform.
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Objective To examine the changes of Aβ expression and its related metabolic enzymes in the brains of AD and T2DM mice, so as to explore the possible mechanism of type 2 diabetes mellitus combined with AD.Methods Five-month-old APP/PS1 double transgenic mice, ob/ob mice and the wild-type control mice were employed in this study.Immunohistochemical (IHC) staining, Elisa and Western blot were used to detect SP, Aβ and its related metabolic enzymes.Results A certain number of SPs were observed in the cerebral cortex and hip-pocampus of APP/PS1 mice;SPs were occasionally observed in the cortex of ob/ob mice, while no SP appeared in wild-type mice.Aβ40 and Aβ42 levels were significantly increased in APP/PS1 and ob/ob mice brains as compared with controls (P<0.05), thought both Aβ40 and Aβ42 levels in AD mice were significantly higher than those of ob/ob mice (P<0.05).APP expression level was highest in APP/PS1 mice among 3 groups, and its expressed higher in ob/ob mice than that of control mice (P<0.05).BACE1 expression was notably increased in APP/PS1 and ob/ob mice as compared with control(P<0.05), however, it expressed higher in APP/PS1 mice than ob/ob mice (P<0.05).The expression of Aβ degradation enzyme IDE was reduced in APP/PS1 and ob/ob mice(P<0.05), while lowest in ob/ob mice.Conclusions Overexpression of Aβ may be one of main reasons for T2DM combined AD.
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Objective:To study the spinal cord injury, spinal cord transection and persistent placeholder damage on the influence of secondary neural cell apoptosis in rats.Methods: Select 60 healthy male Wistar rats, numbered after using the random number table method is divided into A (18,spinal cord contusion),B (18,spinal cord transection),C (18,continuous placeholder),D (6,control),E (6,the control group only) groups of five,were observed at the 1,4,7 D after 5 group of rats nerve cell apoptosis index, spinal cord tissue Bcl-2,the expression of Bax,caspase 3 protein.Results:A,B,C three groups of rats after building 1 d are gray and white matter positive markers, and the gray matter and white matter of three groups of rats nerve cell apoptosis index differences statistically significant ( P<0.05);4 d,7 d after building gray matter and white matter of three groups of rats tend to place increased ap-optotic cells in the spinal cord index ( P<0.05);in building 1,4,7 d group C after rat spinal cord grey matter and white matter of apoptotic cell index was significantly higher than that of group A and group B, group B were significantly higher in group A and the differences were statistically significant (P<0.05).1,4,7 d after building A,B,C,D,E five group rats the Bcl-2,Bax,caspase-3 protein expression differences were statistically significant (P<0.05),1,4,7 d after building A,B,C the Bcl-2 of three groups of rats, Bax,caspase-3 protein expression was significantly higher than that of group D and group E ( P<0.05).Conclusion: Secondary rats after spinal cord injury of nerve cells apoptosis,apoptosis time,severity,and damage type and severity.
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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.
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This paper proposed the idea of building the applied anatomical experiment and research platform for surgical postgraduates with professional degree,establishing double-tutorial system and applying applied anatomical teaching in basis course learning,clinical skill training and research capacity cultivating after analyzing the reasons of poor applied anatomical background of surgical postgraduates with professional degree.These ideas were intended to improve the cultivation quality.
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ObjectiveTo investigate the distribution and expression of y-secretase subunit (APH-1)in the central nervous system (CNS) of APP/PS1 double transgenic Alzheimer's disease (AD) adult mouse model,and to detect the expression difference of APH-1 in developmental brain between AD model mouse and wild-type littermates in order to further clarify the relationship between APH-1 and AD. MethodsOffspring bred by APP/PS1 double transgenic AD mice were genotyped.Immunohistochemical staining was used to detect APH-1 distribution and expression in the CNS of adult APP/PS1 double transgenic AD mouse model,in the brain of AD model mouse and its wild-type littermates on postnatal day 1,7,21 and 120.Results APH-1 was widely expressed in almost all regions of the CNS,especially in the cerebral cortex,hippocampus,olfactory bulb,hypothalamus,ventral striatum,caudate putamen,raphe magnus nucleus,cerebellum,brainstem and spinal cord of the adult APP/PS1 double transgenic mice.APH-1 expression was higher in the cortex of both AD and wild type mouse on postnatal day 1 than on postnatal day 7 and 21 with increased level of APH-1 protein in adult mouse brain.APH-1 expression in the brain of AD mice was higher than in its wild type littermates at any stage(P＜0.05).Conclusions Distribution of APH-1 is ubiquitous and region-dependent in the CNS.The different distribution and expression between APP/PS1 double transgenic mouse model and its wild type littermate indicate that APH-1 may be related to AD.
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Objective To investigate whether valproic acid (VPA) affect spatial learning memory and senile plaques in the APP/PS1 double transgenic AD mouse model of different gender. MethodsTwenty 3-month old APP/PS1 double transgenic AD mice,male and female mouse evenly,were randomly divided into VPA-treated and saline-treated groups ( 10 for each group). 30 mg· kg-1 · d-1 of VPA and the same amount of saline were peritoneally injected into mice for 4 weeks. Morris water maze was conducted to check the effect of VPA on the capability of spatial learning and memory of AD mouse model. Immunohistochemical staining was used to examine the effect of VPA on the morphological changes in the brains of mice. ResultsVisible platform test showed that VPA-treated and saline-treated mice had similar escape latency (P＞0.05) and path length (P＞0.05) ,the swimming speed between male and female mice had no difference (P＞0.05). Hidden platform test showed that VPA treated mice had a significantly shorter latency (P＜0.01) and path length (P＜0.01) to reach the platform compared with saline-treated mice. Meanwhile, both in VPA-treated and control groups, the male mice had a shorter correlation escape latency and path-length than female mice had(P＜0.05 ). Immuohistochemical staining showed that the number (11.23±3.78) of senile plaques (SP) in the cerebral cortex and hippocampus of VPA-treated male mice were notably decreased than that(28.17 ±3.46) in the control group ( t= 14.67, P＜0.01 ),furthermore,the number of SP in the cerebral cortex and hippocampus of VPA-treated male mice was significantly reduced,as compared with which (20.36 ±4.21)in the VPA-treated female mice(P＜0.05). ConclusionVPA can significantly lower formation of SP, and remarkably improve the capability of spatial learning and memory of APP/PS1 transgenic mice,which have gender difference.
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Objectives To investigate whether degradation of anterior pharynx decfective-1(Aph-1) goes through proteasomal pathway or lysosomal pathway.Methods Various methods such as cell culture,Western blotting,pulse-chase metabolic labeling technique,double immunofluoresecnt staining,combined with proteasomal and lysosomal inhibition were used to check Aph-1 expression level in stable Aph-1-transfected or non-transfected neuronal(SH-SY5Y)cell line.Results Using Western blotting,treating the neuronal cells with proteasome specific inhibitors significantly increased the expression of both endogenous and exogenous Aph-1.The effect of the proteasome inhibitors on Aph-1 expression was dose-and time-dependent Lysosomal pathway was not involved in Aph-1 degradation. Pulse-chase metabolic labeling experiment showed that the turnover of newly-synthesized radiolabeled Aph-1 protein was blocked by Lactacystin.Double immunofluorescent staining revealed colocalization of Aph-1 and ubiquitin in the same cells.Conclusion Degradation of Aph-1 protein is mediated by proteasomal pathway in neuronal cells,and is not related to lysosomal pathway.Aph-1 protein is ubiquitinated before degradation.
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Objective To explore the role of transcription factor activator protein 2?(AP-2?) on the expression of tyrosine hydroxylase(TH) in noradrenergic neurons of locus ceruleus(LC) after traumatic brain injury.Methods Oinety rats were!randomly divided into control group and injury group that comprised 5 subgroups.There were 15 rats in each group.The rat model of traumatic brain injury was established with Feeney's methods.Brain stems were dissected from decapitated heads 1,3,6,24 and 72 h after traumatic injury and freeze-mounted for cryo-sectioning.TH and AP-2? expressions in noradrenergic neurons of LC were analyzed with double immunofluorescence.Results The number and fluorescence intensity of TH and AP-2?-positive neurons in LC at various intervals after injury significantly increased as compared with control group(P
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Objective To investigate the changes of dopamine-?-hydroxylase(DBH) and activator protein 2-?(AP-2?) expression in spinal cord under the condition of stress or pain stimulation,so as to explore the mechanism for changes of noradrenergic(NA) neurons in the spinal cord of rat pain model.Methods Immunohistochemical staining,double immunofluorescent staining,Western blotting and computing-image analysis system were used to detect the changes of DBH/AP-2? expression in the spinal cord of formalin-induced rat pain model.Results A small number of DBH-positive neurons were sparsely distributed in the ventral horn of the normal spinal cord,while in the formalin-treated group,much more darkly-stained DBH-positive neurons appeared primarily in the ventral horn,intermediate zone,and the dorsal horn,which reached the highest level on day 3 after formalin-injection.The grey value and number of DBH-positive neurons on day 7 after injection began to decrease,but still higher than that in the control group.Compared with control group,the number of noradrenergic neurons in spinal cord of formalin-treated rat was increased significantly,which was also confirmed by Western blotting.Double immunofluorescent staining showed that DBH and AP-2? co-existed in the cells of the spinal cord.The changes of AP-2? expression were similarly to that of DBH in the spinal cord of rat pain model.Conclusion Our results indicated that some non-noradrenergic neurons with different chemical properties might convert into noradrenergic neurons under pain stimulation;noradrenaline may be involved in the formalin-induced pain and behavior regulation;As one of transcription factors,AP-2? may promote the DBH synthesis.
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AIM: To explore the possibility that proteasome is involved in nicastrin(NCT) degradation and NCT is ubiquitinated before degradation.METHODS: Following the generation of NCT stable cell lines,the methods of Western blotting,pulse-chase metabolic labeling technique,double immunofluorescent staining,combined with proteasomal inhibition were used to investigate the NCT expression in NCT stable cell line.RESULTS: Treatment of the cells with proteasomal inhibitors significantly increased both endogenous NCT(produced by the cell itself) and exogenous NCT(produced by the gene transfection) in SH-SY5Y cells.The effect of specific proteasomal inhibitor lactacystin on NCT expression was in time-and dose-dependent manners.Pulse-chase metabolic labeling experiment showed that the turnover of newly-synthesized radio-labeled nicastrin protein was blocked by lactacystin.The results of double immunofluorescent staining showed that NCT and ubiquitin were co-located in the cells.CONCLUSION: The proteasome is involved in the degradation of NCT in neuronal cells,and NCT is ubiquitinated before degradation.