肾脏病理.pptVIP

a1,3,5(Col-IV)免疫荧光染色及其应用复旦大学附属儿科医院病理科Col-IV胶原分子在体内的分布IV胶原合成或降解障碍Col-IV型胶原的分子结构与分布TypeIVcollagenchainsform3formsofheterotrimericunitsinbasementmembranes:a1a1a2,a3a4a5anda5a5a6Col-IV型胶原的分子结构与分布3’3’3’COL4A6COL4A1COL4A4COL4A3COL4A2COL4A55’5’5’5’5’5’130pb450pb3’3’3’Xq222q3513q34SchematicrepresentationofthedistributionoftypeIVcollagengeneonchromosomes13,2andXTypeIVcollagenfamilycomprisessixdistinctchains[a1(IV)toa6(IV)]encodedbysixgenes(COL4A1–COL4A6)TypeIVcollagenchainsform3formsofheterotrimericunitsinbasementmembranes:a1a1a2,a3a4a5anda5a5a6a3a4a5typeIVcollagennetworkpredominatesinGBMɑ1-ɑ1-ɑ2--所有器官的BMɑ3-ɑ4-ɑ5--分布于GBMLBM眼耳ɑ5-ɑ5-ɑ6–皮肤血管BMɑ1、ɑ2链—TBM、肾球囊壁BM、小球MeS和GBMɑ3、ɑ4、ɑ5链—GBM、DTBMɑ6链—肾球囊壁BM、CTBMa1,3,5(Col-IV)在肾组织表达a1,3,5(IV)荧光染色方法荧光素：1、异硫氢酸荧光素（黄绿色荧光）fluoresceinisothiocyanate,FITC,495nm2、罗丹明（红色荧光）LissaminerhodamineB,RB200,3、R—藻红蛋白R-pheoerythin,R-PE直接法：α1，α3（ＩＶ）免疫荧光技术切片滴加一抗抗置于37℃恒温箱孵育1h70%甘油（甘油：PBS=7:3）封片。免疫荧光技术间接法：α5(IV)　　a5加修复液10mins（0.1MKCL+0.1MHCL等比例混合）　　滴加一抗孵育过夜，PBS洗约5mins　　滴加二抗置于37℃恒温箱孵育1h　　70%甘油（甘油：PBS=7:3）封片。α1α3α5ɑ1—TBM、球囊壁BM、MeR、GBM；ɑ3,ɑ5链—GBM.DTBMα3Euro-DiagnosticCo的AlportSKit的染色结果评判NormalAlportdominantAlportrecessivekidneyskinkidneyskinkidneyskinMAB1++++++MAB3+-----MAB5+++--++◆用于遗传性肾小球基膜病的诊断InheritedGBMdiseasesInheritedpodoyctediseasesHematuriaProteinuriaa3b1-integrina-actinin-4agb3-integrinuPARPaxilinGlepp1PodocalyxinNherf2EzrinActincytoskeletona-actinin-4SynaptopodinVinculinTalinILKCD151ab-dystroglycanUtrophinVitronectinAgrinLamininCollagenIV(a3,a4,a5)Slit-DiaphragmcomplexGBMGBM的主要成分是IV型胶原.层连蛋白.蛋白多糖和巢蛋白ɑ1ɑ2ɑ3ɑ4ɑ5ɑ6GHI注：正常人:G～I中ɑ2(红)；ɑ5（绿）DisordersduetomutationsingenesencodingGBMproteinsAlportsyndromeX-linkedX-linked+diffuseleiomyomatosisAutosomalrecessiveAutosomaldominantThinbasementmembranenephropathyHANAC(Hereditaryangiopathynephropathyaneurysmmusclecramps)PiersonsyndromeLocusCOL4A5COL4A5+COL4A6COL4A3orCOL4A4COL4A3orCOL4A4COL4A3orCOL4A4OtherunidentifiedlociCOL4A1LAMB2DisordersduetodysregulationofgenesencodingGBMproteinsNail-patellasyndromeLMX1B遗传性肾小球基膜病TYPE