高胆固醇饮食加铜水喂养WHBE兔引起的脑组织病理学改变

Brain histopathological changes in WHBE rabbits fed with high cholesterol diet plus copper drinking water

目的观测散发性WHBE兔阿尔茨海默症(AD)模型脑组织病理学变化。方法取雄性WHBE兔30只,随机分成3组:正常对照(NC)组,高胆固醇饮食(HCD)组,高胆固醇饮食+铜饮水(HCD+Cu^(2+))组,每组10只;另取10只老年WHBE兔作为老年(Senile)组。NC组和Senile组给予普通饲料,HCD组给予2%胆固醇饲料,HCD+Cu^(2+)组给予2%胆固醇饲料和添加0.12 ppm铜饮水,连续造模12周。造模12周时,取血测定总胆固醇(TC)和β淀粉样蛋白(Aβ)1-42水平;取部分脑组织测定脑皮质和海马组织中超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量,另取冠状切片脑组织行免疫组化染色观察Aβ、β-分泌酶1(BACE1)、磷酸化tau(p-tau)蛋白的阳性表达情况,同时切片行刚果红和皮尔苏斯基氏(Bielschowsky)染色分别观察老年斑和神经纤维缠结情况。结果老年组WHBE兔的体重明显高于NC组(P<0.01),各组血浆TC、Aβ1-42明显高于NC组(P<0.05,P<0.01);且各组脑组织中SOD活性明显低于NC组(P<0.05),MDA含量显著高于NC组(P<0.05,P<0.01)。免疫组化染色显示,各组脑组织中Aβ、BACE1、p-tau蛋白阳性表达均显著高于NC组(P<0.05,P<0.01),且HCD+Cu^(2+)组脑组织中BACE1和p-tau蛋白阳性表达亦显著高于HCD组(P<0.05,P<0.01)。刚果红和Bielschowsky染色显示HCD组、HCD+Cu^(2+)组和老年组WHBE兔脑组织中观察到大量的老年斑和神经纤维缠结。结论高胆固醇饮食或复合添加微量铜饮水能诱导散发性AD模型WHBE兔脑部明显的AD病理学变化,包括氧化损伤、脑内Aβ沉积增多、老年斑和tau病理学等改变,WHBE兔可用于神经退行性疾病动物模型的研究。

Objective To observe the pathological changes of brain tissues in the WHBE rabbit model of sporadic Alzheimer disease（ AD）. Methods Thirty 3-4-month old male WHBE rabbits were randomly divided into 3 groups：normal control（ NC） group,high cholesterol diet（ HCD） group,high cholesterol diet ＋ copper drinking water（ HCD ＋Cu^2＋） group,10 in each group. Another 10 senile（ 36-48-month old） male WHBE rabbits were taken as senile group.The NC group and the senile group were fed a normal basic diet,the HCD group fed a 2% cholesterol diet,and the HCD＋ Cu^2＋ group fed a 2% cholesterol diet plus 0. 12 PPM copper drinking water for 12 weeks. The levels of total cholesterol（ TC） and β-amyloid protein（ Aβ） 1-42 were measured at 12 weeks. The activity of superoxide dismutase（ SOD） and the content of malondialdehyde（ MDA） in the cortex and the hippocampus were detected. In addition,the covered area of Aβ,β-site APP cleaving enzyme 1（ BACE1） and phosphorylated tau（ p-tau） protein in coronal sections of brain tissues were also observed by immunohistochemical staining. The senile plaques and the neurofibrillary tangles were observed by Congo red and Bielschowsky staining,respectively. Results The body weight of WHBE rabbits in the senile group was significantly higher than that of the NC group（ P〈0. 01）,and the plasma TC and Aβ1-42 in each group were significantly higher than that in the NC group（ P〈0. 05,P〈0. 01）. The activity of SOD in brain tissues was significantly lower than that of NC group（ P〈0. 05）,and the MDA content was significantly higher than that of NC group（ P〈0. 05,P〈0. 01）. Immunohistochemical staining showed that the covered area of Aβ,BACE1 and p-tau in brain tissues of all groups were significantly higher than that of NC group（ P〈0. 05,P〈0. 01）,and the covered area of BACE1 and p-tau protein in the brain tissues of HCD ＋ Cu^2＋ group was also significantly higher than that of the HCD group（ P〈0. 05,P〈0. 01）. Congo red and Bielschowsky staining showed that the number of senile plaques and neurofibrillary tangles were observed in the brain tissues of the HCD,HCD ＋ Cu^2＋ and senile groups. Conclusions High cholesterol diet or supplemented with trace copper drinking water can induce obvious AD pathological changes in WHBE rabbit models of sporadic AD with obvious oxidative damage,increased Aβ deposition and senile plaque in the brain,and pathological changes of tau. WHBE rabbit can be used in the study of animal models of neurodegenerative diseases.