SAMP8小鼠海马内CD147与NCT的表达及其相互关系

The expression and relationship between CD147 and nicastrin in the hippocampus of SAMP8 mouse

目的:运用免疫荧光组织化学和免疫共沉淀技术(Co IP),探讨快速老化小鼠(SAMP8)海马组织中细胞外基质金属蛋白酶诱导因子(CD147)和单过性跨膜蛋白(NCT)间的表达情况,为寻找阿尔茨海默病(Alzheimer disease,AD)相关的治疗策略和方向提供理论依据。方法:3月龄雄性SAMP8小鼠6只。3只小鼠用于免疫荧光组织化学实验,经4%多聚甲醛灌注、取脑、石蜡包埋后,冠状面切片进行组织化学染色,观察CD147与NCT在海马区是否存在共定位的关系。3只用于Co IP实验,提取海马组织蛋白,与CD147抗体孵育后加入琼脂糖微珠A/G孵育过夜,离心后上清用于Western Blot实验检测CD147与NCT间的相互作用关系。结果:SAMP8海马区,CD147和NCT的阳性表达部位主要分布于CA1、CA3、CA4区的锥体细胞层和齿状回颗粒细胞层,二者在CA1、CA3区表达较强;标记部位为胞膜、胞浆,部分神经元在胞浆进核周处存在不连续的点、线状共标记部位。Co IP实验显示CD147与成熟型NCT相互作用,不与非成熟型NCT相互作用。结论:CD147和NCT在SAMP8小鼠海马区锥体细胞层表达较高,推测二者可能与海马的学习记忆功能密切相关;CD147与NCT存在相互作用的关系,推测CD147可能参与调节γ-分泌酶活性。

Objective： To investigate the expression of extracellular matrix metalloproteinase inducer （CD147） and nicastrin （NCT） in the hippocampus of senescence-accelerated-prone mouse 8 （SAMP8） mouse by immunofluorescence histochemistry and eo-immunoprecipitation （CoIP） techniques, it provides theoretical basis to the treatment for Alzheimer＇s dis- ease relevant policies and direction. Methods： Male SAMP8 mice were obtained from 3-month-old （n = 6）. Three mice were peffused with 4% paraformaldehyde followed by removed the brains and embedded in paraffin, and then coronal sections were used for immunofluorescence histochemical tests to observe the existence of co-localization of CD147 with NCT. The other three mice were used for CoIP test, the hippocampus were lysed and the protein was incubated with anti-CD147 antibody followed by incubated with Protein A/G sepharose overnight. The supernatant were used for Western Blot test to detect the interrelation between CD147 and NCT. Results： The positive labeling of CD147 and NCT appeared mainly in the CA1, CA3, CA4 pyramidal cells and granulazcell of the dentate gyrus in the CA1, CA3, CA4 of SAMP8 mice, where the most intensive areas appeared in the CA1 and CA3. Labeling sites appeared both in cell membrane and cytoplasm, and the co-localization were observed in near around the nucleus in some of neurons, which presented discrete points or lines. The CoIP test showed that CD147 interacted with mature NCT nor immature NCT. Conclusion： CD147 and NCT express more intensive in the pyramidal ceils of hippocampus of SAMP8 mice, speculated that they may participate in the function of learning and memory associated with hippocampus. CD147 interacts with NCT in hippocampus of SAMP8 mice, speculated that CD147 may adjust the activity of γ-secretase complex.