CAJ | 학술논문

目的对APP /tau/ PS1阿尔茨海默病(AD)转基因小鼠基因型进行鉴定,并研究在其病程进展中tau蛋白的病理变化. 方法根据PS1、APP、tau基因序列设计特异性引物,PCR扩增对三转基凶AD小鼠的基因型进行鉴定;免疫组化法对2、4、8个月龄小鼠大脑海马区tau蛋白表达进行检测;Western blot检测随着月龄增长小鼠海马组织的tau蛋白及其不同位点磷酸化水平. 结果三转基凶小鼠基凶组PCR扩增获得片段分别为960 bp、530 bp和400 bp,与APP、PS1、tau的预期条带大小相符;免疫组化检测发现,8月龄小鼠海马CA3区tau蛋白表达的阳性区域增加;Western blot检测发现,三转基因小鼠海马区tau蛋白及其ps262、ps404、ps202位点磷酸化水平均较正常野生型小鼠升高(P＜0.01),且随月龄增长,8、12个月龄小鼠tau蛋白表达较2个月龄均增高(P＜0.01);Ps404及Ps202位点的磷酸化水平自2个月龄开始就高于野生型(P＜0.01),8个月龄时表达较2个月龄再次出现升高(P＜0.01),而Ps262位点的磷酸化水平直到12个月龄才较野生型出现显著性变化(P＜0.01). 结论三转基凶小鼠能够稳定的遗传并表达APP/tau/PS,此三转基因动物能很好地模拟AD病tau蛋白的病理特征;tau蛋白及各位点磷酸化水平增高的时间并不一致,8个月龄可能是一个关键的时间点,这将为今后应用此动物模型进行的的病理及靶向治疗药物研究提供科研参考.
목적 대APP /tau/ PS1아이자해묵병(AD)전기인소서기인형진행감정,병연구재기병정진전중tau단백적병리변화. 방법 근거PS1、APP、tau기인서렬설계특이성인물,PCR확증대삼전기흉AD소서적기인형진행감정;면역조화법대2、4、8개월령소서대뇌해마구tau단백표체진행검측;Western blot검측수착월령증장소서해마조직적tau단백급기불동위점린산화수평. 결과 삼전기흉소서기흉조PCR확증획득편단분별위960 bp、530 bp화400 bp,여APP、PS1、tau적예기조대대소상부;면역조화검측발현,8월령소서해마CA3구tau단백표체적양성구역증가;Western blot검측발현,삼전기인소서해마구tau단백급기ps262、ps404、ps202위점린산화수평균교정상야생형소서승고(P＜0.01),차수월령증장,8、12개월령소서tau단백표체교2개월령균증고(P＜0.01);Ps404급Ps202위점적린산화수평자2개월령개시취고우야생형(P＜0.01),8개월령시표체교2개월령재차출현승고(P＜0.01),이Ps262위점적린산화수평직도12개월령재교야생형출현현저성변화(P＜0.01). 결론 삼전기흉소서능구은정적유전병표체APP/tau/PS,차삼전기인동물능흔호지모의AD병tau단백적병리특정;tau단백급각위점린산화수평증고적시간병불일치,8개월령가능시일개관건적시간점,저장위금후응용차동물모형진행적적병리급파향치료약물연구제공과연삼고.
Objective To identify the genotype of the APP/tau/PS1 triple transgenic Alzheimer's disease (AD) mice,and investigate the pathological changes of tau protein in the pathogenic process.Methods Using specific primers of PS1,APP,tau gene,the genotypes of the triple transgenic AD mice were identified.Expression of tau protein in hippocampal tissue of mouse model aged 2,4,8 month was detected by immunohistochemistry.The expression of tau and its hyperphosphorylation in different sites in the hippocampal tissue and different month old mice was detected by Western blotting.Results PCR amplification fragment of 960 bp,530 bp and 400 bp of transgenic mouse genome were the expected size of APP,PS1,tau,respectively.Expression of tau in hippocampal CA3 region was increased obviously in the 8 month old mice.Compared with the normal wild-type mice,the expressions of tau and phosphorylation of pS262,pS404 and pS202 were increased significantly in hippocampus tissue of the transgenic mice (P＜0.01).Expression of tau were significantly higher in 8-and 12-monthsold mice than in 2 months-old mice (P ＜ 0.01).Phosphorylation level of pS404 and Ps202 was significantly increased since 2-months-old in transgenic mouse compared to the wild type mouse (P＜0.01),and in 8-monthold mice,there was also a significant increase as compared to that in 2 month-old mice (P＜0.01).As to the phosphorylation level of pSs262,the significant increase did not appear until 12 months old in transgenic mouse as compared to the wild type mouse (P＜0.01).Conclusions The triple transgenic mice can stably express the APP/tau/PS1 gene.The transgenic animals can be a useful model with the pathological features of tau of AD.The phosphorylation level of tau in different site increases in different time,which will provide useful research reference in Alzheimer's disease pathology and medication research.