中华眼科杂志
中華眼科雜誌
중화안과잡지
Chinese Journal of Ophthalmology
2015年
11期
831-838
,共8页
刘超启%孙哲%曹立宁%王方
劉超啟%孫哲%曹立寧%王方
류초계%손철%조립저%왕방
肽碎片%淀粉样β肽类%注射,眼内%视网膜%视网膜色素上皮%细胞衰老%小鼠
肽碎片%澱粉樣β肽類%註射,眼內%視網膜%視網膜色素上皮%細胞衰老%小鼠
태쇄편%정분양β태류%주사,안내%시망막%시망막색소상피%세포쇠로%소서
Peptide fragments%Amyloid beta-peptides%Injections,intraocular%Retina%Retinal pigment epithelium%Cell aging%Mice
目的 探讨小鼠视网膜下注射淀粉样蛋白β(1-42)寡聚体(OAβ1-42)后视网膜组织的变化.方法 实验研究.120只8周龄C57BL/6N小鼠单纯随机抽样法分为双蒸水组和OAβ(1-42)组,分别在视网膜下腔注射双蒸水和浓度为0.312 5mmol/L的OAβ(1-42).注射后于不同时间点对实验眼进行观察,包括眼底照相及自发荧光成像、视网膜电图检查、视网膜组织切片HE染色、神经视网膜铺片β-半乳糖苷酶染色、视网膜色素上皮/脉络膜铺片老化标志物免疫荧光染色、视网膜组织中老化相关因子的免疫印迹法及荧光定量PCR检测.实验数据采用独立样本t检验进行统计学分析.结果 与对照组比较,注射后1周时实验组小鼠眼底照相可见视网膜组织萎缩伴自发荧光增强;暗适应视网膜电图显示a波振幅均值下降为(39.94±7.75)μV[对照组(225.27±28.94)μV,t=12.45,P＜0.001],b波振幅均值下降为(185.55±4.62)μV[对照组(873.78±43.80)μV,t=27.06, P＜0.001];HE染色可见视网膜色素上皮细胞色素分布紊乱,视网膜厚度显著变薄(t=75.13,P＜0.001);β半乳糖苷酶染色及免疫荧光染色老化标志物表达明显增高;以上表现持续到注射后8周.同时分子生物学检测提示,注射后1周时实验组小鼠视网膜组织老化相关因子较对照组表达显著增高.结论 OAβ(1-42)在小鼠视网膜下聚集后可导致视网膜组织老化,使其视觉功能下降,该表现与AMD临床表现类似.
目的 探討小鼠視網膜下註射澱粉樣蛋白β(1-42)寡聚體(OAβ1-42)後視網膜組織的變化.方法 實驗研究.120隻8週齡C57BL/6N小鼠單純隨機抽樣法分為雙蒸水組和OAβ(1-42)組,分彆在視網膜下腔註射雙蒸水和濃度為0.312 5mmol/L的OAβ(1-42).註射後于不同時間點對實驗眼進行觀察,包括眼底照相及自髮熒光成像、視網膜電圖檢查、視網膜組織切片HE染色、神經視網膜鋪片β-半乳糖苷酶染色、視網膜色素上皮/脈絡膜鋪片老化標誌物免疫熒光染色、視網膜組織中老化相關因子的免疫印跡法及熒光定量PCR檢測.實驗數據採用獨立樣本t檢驗進行統計學分析.結果 與對照組比較,註射後1週時實驗組小鼠眼底照相可見視網膜組織萎縮伴自髮熒光增彊;暗適應視網膜電圖顯示a波振幅均值下降為(39.94±7.75)μV[對照組(225.27±28.94)μV,t=12.45,P＜0.001],b波振幅均值下降為(185.55±4.62)μV[對照組(873.78±43.80)μV,t=27.06, P＜0.001];HE染色可見視網膜色素上皮細胞色素分佈紊亂,視網膜厚度顯著變薄(t=75.13,P＜0.001);β半乳糖苷酶染色及免疫熒光染色老化標誌物錶達明顯增高;以上錶現持續到註射後8週.同時分子生物學檢測提示,註射後1週時實驗組小鼠視網膜組織老化相關因子較對照組錶達顯著增高.結論 OAβ(1-42)在小鼠視網膜下聚集後可導緻視網膜組織老化,使其視覺功能下降,該錶現與AMD臨床錶現類似.
목적 탐토소서시망막하주사정분양단백β(1-42)과취체(OAβ1-42)후시망막조직적변화.방법 실험연구.120지8주령C57BL/6N소서단순수궤추양법분위쌍증수조화OAβ(1-42)조,분별재시망막하강주사쌍증수화농도위0.312 5mmol/L적OAβ(1-42).주사후우불동시간점대실험안진행관찰,포괄안저조상급자발형광성상、시망막전도검사、시망막조직절편HE염색、신경시망막포편β-반유당감매염색、시망막색소상피/맥락막포편노화표지물면역형광염색、시망막조직중노화상관인자적면역인적법급형광정량PCR검측.실험수거채용독립양본t검험진행통계학분석.결과 여대조조비교,주사후1주시실험조소서안저조상가견시망막조직위축반자발형광증강;암괄응시망막전도현시a파진폭균치하강위(39.94±7.75)μV[대조조(225.27±28.94)μV,t=12.45,P＜0.001],b파진폭균치하강위(185.55±4.62)μV[대조조(873.78±43.80)μV,t=27.06, P＜0.001];HE염색가견시망막색소상피세포색소분포문란,시망막후도현저변박(t=75.13,P＜0.001);β반유당감매염색급면역형광염색노화표지물표체명현증고;이상표현지속도주사후8주.동시분자생물학검측제시,주사후1주시실험조소서시망막조직노화상관인자교대조조표체현저증고.결론 OAβ(1-42)재소서시망막하취집후가도치시망막조직노화,사기시각공능하강,해표현여AMD림상표현유사.
Objective To investigate the alterations of retinal tissue induced by OAβ(1-42) subretinal injection in normal C57BL/6N mice.Methods Experimental study.One hundred and twenty C57BL/6N mice aged 8 weeks participated in this study.Among them, 60 eyes received subretinal injection of OAβ(1-42);another 60 received double distilled water as control.The concentration of OAβ(1-42) was 0.312 5 mmol/L.We took fundus photograph and autofluorescence, electroretinogram(ERG), haematoxylineosin staining of retinal paraffin sections, β-galactosidase staining of neural retinal flatmounts, immunofluorescence of senescence marker P16INK4a of RPE/choroidal flatmounts, Westeru blot and RT-PCR of senescence relative cytokines IL-6, TGF-β1 before and after the injection.The data were analyzed by independent sample t-test.Results Comparing with controls, 1 week post injection, the experimental group eyes had atrophied area with high autofluorescence site, and showed evident decrease in amplitudes of a wave (39.94±7.75)μV comparing with the controls (225.27±28.94)μV(t=12.45, P＜0.001) in scotopia ERG, so did the amplitude of b wave (185.55±4.62)μV comparing with the controls (873.78±43.80)μV(t=27.06, P＜0.001), retinal structures appeared significant loss of retinal thickness (t=75.13, P＜0.001) along with hypo-or hyperpigmentation;the experimental eyes had high frequency of blue-green deposits in retinal β-gal staining;the green fluorescence sites of RPE/choroidal flatmounts were clear and bright, indicating higher expression of P16INK4a in the nucleus;these results were the same till 8 weeks post operation.Western blot and RT-PCR vertified conspicuous increases in expression of TGF-β1, IL-6 in retinas of experimental eyes compared with the control ones 1 week post injection.Conclusions OAβ(1-42) subretinal injection could induce visual function deficiency and retinal tissue senescence.All the manifestations were AMD-like retinopathy, which implied that Aβ(1-42) might be an essential clue in human AMD pathology.
