ApoE基因缺失小鼠视网膜及Bruch膜组织形态观察

Morphous observation on retina and Bruch’s membrane in ApoE-/-mice

目的探讨ApoE基因缺失(apolipoprotein E-deficient,ApoE-/-)小鼠视网膜及Bruch膜的组织形态改变。方法随机将24只2月龄C57BL小鼠分为高脂组和普食组,每组12只,另用2月龄ApoE-/-小鼠12只作为普食组,喂养4个月后检测各组动物血浆总胆固醇(total cholesterol,TC)、低密度脂蛋白(low density lipoprotein,LDL)、甘油三脂(triglyceride,TG)含量,光镜和电镜下观察视网膜外核层(outer nuclear layer,ONL)细胞、视网膜色素上皮(retinal pigment epithelium,RPE)细胞和Bruch膜形态结构改变,半定量检测视网膜ONL、RPE细胞数和Bruch膜厚度。结果 6月龄ApoE-/-普食组小鼠、6月龄C57BL高脂组小鼠、6月龄C57BL普食组小鼠血浆TC的含量分别为(9.72±0.41)mmol·L-1、(6.97±0.54)mmol·L-1、(5.15±0.31)mmol·L-1,LDL的含量分别为(1.05±0.16)mmol·L-1、(1.25±0.10)mmol·L-1、(0.43±0.08)mmol·L-1,TG的含量分别为(0.78±0.16)mmol·L-1、(0.56±0.19)mmol·L-1、(0.53±0.17)mmol·L-1,ONL细胞核面积分别为(23208.00±716.91)μm2、(24269.00±1263.13)μm2、(26952.00±2449.10)μm2,RPE细胞数目分别为10.20±0.83、11.70±0.62、13.80±1.01,Bruch膜厚度分别为(0.87±0.04)μm、(0.77±0.07)μm、(0.59±0.03)μm。TC:6月龄ApoE-/-普食组小鼠比6月龄C57BL高脂组和普食组明显增高(P<0.001);LDL:6月龄ApoE-/-普食组小鼠比6月龄C57BL普食组明显增高(P<0.001);TG:6月龄ApoE-/-普食组小鼠比6月龄C57BL普食组明显增高(P<0.05),比6月龄C57BL高脂组增高(P>0.05)。光镜下观察发现:6月龄ApoE-/-普食组小鼠视网膜ONL、RPE细胞排列紊乱、萎缩;半定量结果显示,6月龄ApoE-/-普食组小鼠比6月龄C57BL普食组视网膜RPE细胞明显减少(P<0.001),ONL细胞数显著减少(P<0.01),Bruch膜显著增厚(P<0.001)。6月龄ApoE-/-普食组小鼠比6月龄C57BL高脂组视网膜RPE细胞明显减少(P<0.01),ONL细胞数减少(P>0.05),Bruch膜显著增厚(P<0.05)。电镜观察发现:6月龄ApoE-/-普食组小鼠RPE基底膜皱褶减少,胞质内线粒体轻度肿胀,可见核边聚现象,Bruch膜纤维结构不均匀,弹力层断裂,有空泡出现。结论 ApoE-/-普食组小鼠出现显著高脂血症,引起视网膜ONL、RPE及Bruch膜呈现AMD的病理改变,表明ApoE-/-与AMD具有明显相关性。

Objective To explore the morphological characteristics of retina and Bruch＇s membrane in ApoE-/- mice. Methods Twenty-four C57BL mice aged two months were randomly divided into high fat diet group and normal diet group, 12 cases in each group, and 12 ApoE-/- mice aged two months were chosen as normal diet group. After being fed for 4 months, levels of total plasma cholesterol （ TC）, low density lipoprotein （LDL） and trigiyceride （TG） in each group were determined, respectively, morphological characteristics of retinal outer nuclear layer cells （ ONLC）, retinal pig- ment epithelium cells （RPEC） and Bruch＇ s membrane were observed under light and electron microscope, and amounts of ONLC and RPEC, Bruch＇ s membrane thickness were examined by semi-quantitative histopathology. Results Levels of TC in ApoE-/- mice of normal diet group and C57BL mice of high fat diet and normal diet groups were （9.72±0.41）mmol ·L^-1 ,（6.97 ＋0.54） mmol·L^-1,（5.15±0.31） mmo·L^-1 ,respec- tively,levels of LDL were （ 1.05 ±0.16） mmol·L^-1, （ 1.25 ±0.10） mmol·L^-1 , （0.43± 0.08） mmol·L^-1, respectively, levels of TG were （ 0.78±0. 16 ） mmol·L^-1, （ 0.56±0.19）mmol·L^-1, （0. 53 ± 0. 17） mmol·L^-1, respectively, area of ONLC were （23 208.00± 716.91 ）μm, （24 269.00±1253.13 ） μm, （ 26 952.00± 2449.10 ） μn, respec- tively, counts of RPEC were 10.20±0.83,11.70±0.62,13.80±1.01, respectively, thick- ness of Bruch＇ s membranes were （0. 87 ± 0.09） μm, （0.77 ±0. 07 ） Win, （0. 59 ± 0. 03 ）μm, respectively. Levels of TC in ApoE-/- mice of normal diet group were significantly higher than that in C57BL mice of both high fat diet and normal diet groups（P 〈0.001 ） ; Levels of LDL in ApoE-/- mice of normal diet control group were significantly higher than that in C57BL mice of normal diet group（P 〈 0.001 ） ; Levels of TG in ApoE-/- mice of normal diet group were higher than that in C57BL mice of normal diet group （ P 〈 0. 05 ）, higher than that in C57BL mice of high fat diet group（P 〉 0.05 ）. Under light mi- croscope, ONLC and RPEC were disorganized and atrophic in ApoE-/- mice of normal diet control tzrouo. Bv semi-ouantitative histooatholo~.＇v, counts of ONLC and RPEC inApoE-/- mice of normal diet group were significantly less than those in C57BL mice of normal diet groups （P 〈 0.01 and 0. 001 ） ,and thickness of Bruch＇ s membranes was higher than that in C57BL mice of normal diet group（P 〈0.001 ）. While, counts of RPEC in ApoE-/- mice of normal diet group were less than that in C57BL mice of high fat diet group（P 〈0.01 ）, counts of ONLC decreased （P 〉0.05 ）, and Bruch＇ s membranes thickness was higher than in C57BL mice of high fat diet group （P 〈 0.05 ）. Under electron microscope in ApoE-/- mice of normal diet group, reductus in RPE basement membrane reduced,mitochondria swollen slightly, the nuclei aggregation was seen, fiber in Bruch＇ s membrane was irregulared with broken elastic layer and vacuoles. ion Hyperlipidemia can be seen in ApoE-/- mice, which can lead to age-relat- ed macular degeneration in ONLC, RPEC and Bruch＇ s membrane. It is indicated that ApoE-/- is related with age-related macular degeneration.