CAJ | 학술논문

目的了解SD大鼠视觉发育关键期内视网膜厚度的变化,探讨细胞凋亡的相关作用.方法实验研究.选择新生SD大鼠50只,其中10只(20只眼)于生后14、18、21、24及42 d行双眼眼底相干光断层扫描(OCT),测量视网膜厚度.另10只大鼠(20只眼)分别于生后14、18、21、24及42 d各取2只大鼠(4只眼),提取视网膜组织总RNA,实时荧光定量PCR法检测Bcl-2、Bax和Caspase-3 mRNA表达水平.另20只大鼠(40只眼)于各对应时间点取4只(8只眼),制作眼球石蜡切片行HE染色并行视网膜厚度测量.同时取10只大鼠(20只眼)于各对应时间点取2只(4只眼)制作冰冻切片行TUNEL染色.对不同时间点的视网膜厚度,Bcl-2、Bax及Caspase-3的mRNA相对表达量进行方差分析,对两种方法所测视网膜厚度进行线性回归分析.结果大鼠生后14、18、21、24及42 d视网膜厚度OCT测量值分别为(243.42±13.83)、(218.78±8.21)、(195.42±8.02)、(195.74±14.85)及(190.79±11.70)μm,差异有统计学意义(F=15.425,P=0.000).在大鼠生后3周内,视网膜厚度逐渐变薄,此后变化不显著.其结果与HE切片视网膜厚度测量呈线性相关(R=0.794,P=0.000).比较不同时间点大鼠视网膜组织Bcl-2、Bax和Caspase-3 mRNA相对表达量,差异均有统计学意义(F=18.684,F=47.307,F:49.611;P=0.000).Bax和Caspase-3 mRNA表达呈增加趋势,并在生后24 d呈现峰值,生后42 d表达量明显下降.TUNEL染色结果显示生后18 d至42 d视网膜神经节细胞层、内核层和外核层均可见较多凋亡细胞,生后42 d视网膜细胞凋亡明显减少.结论视觉发育关键期内大鼠视网膜厚度逐渐变薄.Cirrus HD-OCT可较准确测量大鼠视网膜厚度,是观察视网膜形态学变化的有效检查方法之一.视觉发育关键期内,细胞凋亡参与大鼠视网膜发育,可能影响视网膜厚度的变化.
목적 료해SD대서시각발육관건기내시망막후도적변화,탐토세포조망적상관작용.방법 실험연구.선택신생SD대서50지,기중10지(20지안)우생후14、18、21、24급42 d행쌍안안저상간광단층소묘(OCT),측량시망막후도.령10지대서(20지안)분별우생후14、18、21、24급42 d각취2지대서(4지안),제취시망막조직총RNA,실시형광정량PCR법검측Bcl-2、Bax화Caspase-3 mRNA표체수평.령20지대서(40지안)우각대응시간점취4지(8지안),제작안구석사절편행HE염색병행시망막후도측량.동시취10지대서(20지안)우각대응시간점취2지(4지안)제작빙동절편행TUNEL염색.대불동시간점적시망막후도,Bcl-2、Bax급Caspase-3적mRNA상대표체량진행방차분석,대량충방법소측시망막후도진행선성회귀분석.결과 대서생후14、18、21、24급42 d시망막후도OCT측량치분별위(243.42±13.83)、(218.78±8.21)、(195.42±8.02)、(195.74±14.85)급(190.79±11.70)μm,차이유통계학의의(F=15.425,P=0.000).재대서생후3주내,시망막후도축점변박,차후변화불현저.기결과여HE절편시망막후도측량정선성상관(R=0.794,P=0.000).비교불동시간점대서시망막조직Bcl-2、Bax화Caspase-3 mRNA상대표체량,차이균유통계학의의(F=18.684,F=47.307,F:49.611;P=0.000).Bax화Caspase-3 mRNA표체정증가추세,병재생후24 d정현봉치,생후42 d표체량명현하강.TUNEL염색결과현시생후18 d지42 d시망막신경절세포층、내핵층화외핵층균가견교다조망세포,생후42 d시망막세포조망명현감소.결론 시각발육관건기내대서시망막후도축점변박.Cirrus HD-OCT가교준학측량대서시망막후도,시관찰시망막형태학변화적유효검사방법지일.시각발육관건기내,세포조망삼여대서시망막발육,가능영향시망막후도적변화.
Objective To observe the changes of retinal thickness during critical period plasticity in rat, and to investigate whether apoptosis participates in the structural forming of retina. Methods Experimental research. 50 normal newborn pups of SD rat were randomly selected in the experiment In vivo consecutive scanning of retinal image was taken and the retinal thickness from RPE to ILM was recorded in 10 pups (20 eyes) with spectral domain optical coherence tomography (OCT) at postnatal day 14 (P14), P18, P21, P24 and P42. Bcl-2, Bax, Caspase-3 Mrna was assessed with fluorescent quantitative real-time polymerase chain reaction after total RNA extracted from 4 retinas of 2 pups at each time point from P14 to P42. Histological measurement of retinal thickness of sections with HE staining from 4 pups (8 eyes) at each time point was compared with the results of OCT scanning. TUNEL staining was used to detect the apoptotic cells in retinal cyrosections of 2 pups (4 eyes) at the same time point The data were analyzed by One-way ANOVA and Linear Regression through SPSS 11.5 software. Results There was significant difference between the retinal thickness measured through OCT in pups from P14 to P42 (F = 15.425 ,P = 0. 001). And the values were (243. 42 ± 13. 83) μm at P14, ( 218. 78 ± 8. 21) (μm at P18, (195.42 ± 8. 02) μm at P21, (195. 74 ± 14. 85) μm at P24, (190. 79 ± 11. 70) um at P42. The retinal thickness measured through OCT decreased significantly during the first 3 weeks after birth. The results of OCT measurement had linear correlation with histology measurement ( R = 0. 794, P = 0. 000 ) . There was significant difference between Mrna expression of Bcl-2, Bax and Caspase-3 in pups from P14 to P42 ( F = 18.684, F=47. 307,F =49. 611 ;P =0.000). The relative expression of Bax and Caspase-3 peaked at P24 while Bcl-2 was much more stable. There were a lot of apoptotic cells in the ganglion cells layer, the inner nuclear layer and the outer nuclear layer during P18 to P24 by TUNEL staining. And the apoptosis alleviated at P42. Conclusions The retinal thickness decreases when the retina continues to develop during critical period plasticity. Cirrus HD-OCT can be used as an effective instrument to show the layers of retina in rat in vivo. Apoptosis participates in the course of retinal development which possibly leads to the thinning of retina.