魚藤酮誘導(dǎo)的線粒體損傷在過表達(dá)人源ApoE4轉(zhuǎn)基因小鼠中的作用分析 目錄24965_WPSOffice_Level1摘要 Ⅰ30132_WPSOffice_Level2Abstract Ⅱ20712_WPSOffice_Level2前言 115742_WPSOffice_Level21材料與方法 230132_WPSOffice_Level11.1實(shí)驗(yàn)動(dòng)物分組及處理 220712_WPSOffice_Level11.2主要實(shí)驗(yàn)試劑 215742_WPSOffice_Level11.3主要試劑配制 219032_WPSOffice_Level11.4主要實(shí)驗(yàn)儀器 31128_WPSOffice_Level11.5實(shí)驗(yàn)方法 419032_WPSOffice_Level21.5.1染毒處理 41128_WPSOffice_Level21.5.2電跳臺(tái)實(shí)驗(yàn) 427405_WPSOffice_Level21.5.3組織冰凍切片樣本制作備 45260_WPSOffice_Level21.5.4線粒體ATP的檢測(cè) 427641_WPSOffice_Level21.5.5活性氧ROS的檢測(cè) 525961_WPSOffice_Level21.5.6免疫熒光實(shí)驗(yàn) 618339_WPSOffice_Level21.5.7Westernblot實(shí)驗(yàn) 63146_WPSOffice_Level21.5.8統(tǒng)計(jì)分析 727405_WPSOffice_Level12實(shí)驗(yàn)結(jié)果 85260_WPSOffice_Level12.1小鼠的一般健康狀況 827641_WPSOffice_Level12.2電跳臺(tái)實(shí)驗(yàn)結(jié)果 825961_WPSOffice_Level12.3線粒體ATP的檢測(cè)結(jié)果 918339_WPSOffice_Level12.4線粒體ROS的檢測(cè)結(jié)果 93146_WPSOffice_Level12.5魚藤酮ROT處理對(duì)小鼠炎癥反應(yīng)的作用 914212_WPSOffice_Level12.6魚藤酮處理對(duì)小鼠Aβ生成、清除及對(duì)突觸蛋白的影響 118239_WPSOffice_Level13討論 1312261_WPSOffice_Level14結(jié)論 1518297_WPSOffice_Level1參考文獻(xiàn) 1618297_WPSOffice_Level1致謝 18長治醫(yī)學(xué)院本科畢業(yè)論文（設(shè)計(jì)）ⅠTOC\o"1-4"\h\u摘要目的：探究經(jīng)口灌胃魚藤酮（ROT）誘導(dǎo)的線粒體損傷在過表達(dá)人源ApoE4轉(zhuǎn)基因小鼠中的作用，明確線粒體損傷能否加重ApoE4轉(zhuǎn)基因小鼠的認(rèn)知行為及病理變化。方法：選用7月齡ApoE4轉(zhuǎn)基因小鼠，并給予處理組（ApoE4+ROT)1mg/kg/d的魚藤酮灌胃，用玉米油做對(duì)照(ApoE4)，處理時(shí)間持續(xù)5周。運(yùn)用電跳臺(tái)行為學(xué)實(shí)驗(yàn)檢測(cè)小鼠認(rèn)知能力變化，用Westernblot實(shí)驗(yàn)檢測(cè)認(rèn)知相關(guān)蛋白的表達(dá)水平，免疫熒光分析ROT處理對(duì)ApoE4小鼠病理改變的作用。結(jié)果：電跳臺(tái)實(shí)驗(yàn)發(fā)現(xiàn)ROT處理對(duì)小鼠認(rèn)知能力無顯著性的改變。線粒體功能學(xué)檢測(cè)發(fā)現(xiàn)ROT處理顯著降低了小鼠線粒體ATP的表達(dá)水平，但對(duì)ROS的作用不顯著。Westernblot實(shí)驗(yàn)發(fā)現(xiàn)ROT處理顯著提升了BACE1的表達(dá)水平，同時(shí)顯著降低了NEP的表達(dá)，說明ROT處理加劇了Aβ的累積。同時(shí)一定程度降低突觸前蛋白和突觸后蛋白的表達(dá)，但作用不明顯。免疫熒光結(jié)果提示ROT處理對(duì)小鼠GFAP的表達(dá)無明顯作用，westernblot的結(jié)果也驗(yàn)證了這一點(diǎn)。結(jié)論：魚藤酮能夠誘導(dǎo)ApoE4小鼠線粒體功能障礙，顯著改變Abeta的生成和清除，但對(duì)小鼠的認(rèn)知能力無明顯作用。關(guān)鍵詞：阿爾茲海默??；魚藤酮；焦慮行為；抑郁行為ⅡAbstractObjective:Toinvestigatetheeffectofrotenone-inducedmitochondrialinjuryinducedbyoralgavageontheoverexpressionofhumanApoE4transgenicmice,andtoexplorewhethermitochondrialinjurycanworsenthebehaviorandpathologicalchangesofApoE4transgenicmice.Methods:ApoE4transgenicmiceat7monthsofagewereselected,androtenone(1mg/kg/d)ofthetreatmentgroup(ApoE4+ROT)wasgivenbygavage,andcornoilwasusedasthecontrol(ApoE4).After5weeksoftreatment,thememoryabilityofthemicewastestedbyelectricjumpingplatformbehaviorexperiment,andfrozensectionsofthehippocampusandcerebralcortexofthemiceweretakenforpathologicalexamination.Results:Thememoryofmicedidnotchangesignificantly.ThenumberofneuronsintheCA1,CA3andDGregionsofthetwogroupsdidnotdecreaseundera40×microscopeandtherewasnosignificantdifferencebetweenthetwogroups.ATPshowedadownwardtrend,butthedifferencewasnotstatisticallysignificant.TheROStestfoundthattheROScontentwasreducedtosomeextent.WesternblotexperimentsshowedthatROTtreatmentsignificantlyincreasedthelevelofBACE1andsignificantlydecreasedtheexpressionofNEP,indicatingthatROTtreatmentincreasedtheaccumulationofA.Atthesametime,theexpressionofpresynapticproteinandpostsynapticproteinwasdecreasedtosomeextent,buttheeffectwasnotobvious.Conclusions:insummary,rotenone-inducedmitochondrialinjuryhasacertaineffectontheoverexpressionofhumanApoE4transgenicmice,butnosignificanteffectoninflammatoryresponse.Keywords:Alzheimer'sdisease;Rotenone;Anxiousbehavior;Depressionbehavior長治醫(yī)學(xué)院本科生畢業(yè)論文（設(shè)計(jì)）參考文獻(xiàn)Alzheimer’sDiseaseInternational.WorldAlzheimerreport2015[EB/OL].[2015-09-23]..王璇;滿進(jìn)勁;李興義;陳娟.阿爾茨海默病遺傳學(xué)研究進(jìn)展[J].神經(jīng)損傷與功能重建,2019,v.14,33-35.SandoSB,MelquistS,CannonA,etal.APOEepsilon4lowersageatonsetandisahighriskfactorforAlzheimer'sdisease;acasecontrolstudyfromcentralNorway[J].BMCNeurol,2008,8:9.FanLiao,TonyJ.Zhang,HongJiang,etal.MurineversushumanapolipoproteinE4:differentialfacilitationofandco-localizationincerebralamyloidangiopathyandamyloidplaquesinAPPtransgenicmousemodels[J].ActaNeuropathologicaCommunications,2015,3(1):1-10.HuangY.RolesofapolipoproteinE4(ApoE4)inthepathogenesisofAlzheimer'sdisease:lessonsfromApoEmousemodels[J].BiochemSocTrans,2011,39(4):924-932.PENGKG,TAOY,ZHANGJ,etal.ResveratrolregulatesmitochondrialbiogenesisandFission/Fusiontoattenuaterotenone-inducedneurotoxicity[J].OxidMedCellLongev,2016,2016:6705621.DOI:10.1155/2016/6705621.鄧勇,逯軍.魚藤酮致線粒體氧化損傷的作用和機(jī)制[J].中南大學(xué)學(xué)報(bào):醫(yī)學(xué)版,2014,39(10):1093-1099.LenazG,GenovaML.Structureandorganizationofmitochondrialrespiratorycomplexes:anewunderstandingofanoldsubject[J].AntioxidRedoxSignal,2010,12(8):961-1007．MurphyMP.Howmitochondriaproducereactiveoxygenspecies[J].BiochemJ,2009,417(1):1-13.丁紅群,丁鎮(zhèn),熊御云,等.魚藤酮誘導(dǎo)線粒體輕度損傷細(xì)胞氧化應(yīng)激時(shí)硫氧還蛋白轉(zhuǎn)錄水平降低[J].細(xì)胞生物學(xué)雜志,2007,29(5):753-757.山媛;郭峰;宋海慶;賈建平;屈秋民.腦啡肽酶基因多態(tài)性與散發(fā)性阿爾茨海默病的關(guān)系研究[J].中風(fēng)與神經(jīng)疾病雜志,2015,v.32;No.208,23-26.NalivaevaNN,BelyaevND,ZhuravinIA,etal.TheAlzheimer’samyloid-degradingpeptidase,neprilysin:canwecontrolit[J].IntJAlzheimersDis,2012,383796.doi:10.1155/2012/383796.韋秋蘭，周常文.ADAM10基因缺失對(duì)成年小鼠大腦Aβ肽的產(chǎn)生和認(rèn)知功能的影響[D].福建：福建醫(yī)科大學(xué)，2013,1-7.樊文萍,張茜,馬建平,等.短期胰島素強(qiáng)化治療誘導(dǎo)初診2型糖尿病患者血糖長期良好控制的臨床研究[J].檢驗(yàn)醫(yī)學(xué)與臨床,2014,11(1):38-39.許保磊,王蓉,張茁.阿爾茨海默病與3型糖尿病[J].中華老年心腦血管病雜志,2012,14(12):1338-1340.張爽曦，龍建綱，劉健康.胰島素降解酶與阿爾茲海默病的發(fā)生[J].中國生物化學(xué)與分子生物學(xué)報(bào),2018,v.34,26-31.PanW,HanS,KangL,etal.EffectsofdihydrotestosteroneonsynapticplasticityofthehippocampusinmidcognitiveimpairmentmaleSAMP8mice[J].ExpTherMed,2016,12(3):1455-1463.XuJ,deWinterF,FarrokhiC,etal.Neuregulin1improves