中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2013年
2期
147-150
,共4页
角膜新生血管%重组canstatin蛋白%碱烧伤%血管内皮生长因子%碱性成纤维细胞生长因子
角膜新生血管%重組canstatin蛋白%堿燒傷%血管內皮生長因子%堿性成纖維細胞生長因子
각막신생혈관%중조canstatin단백%감소상%혈관내피생장인자%감성성섬유세포생장인자
Corneal neovascularization%Recombinant canstatin protein%Alkali burn%Vascular endothelial growth factor%Basic fibroblast growth factor
背景 碱烧伤后角膜新生血管(CNV)形成可引起致盲性眼病,最近研究表明血管内皮生长因子(VEGF)和碱性成纤维细胞生长因子(bFGF)是诱导血管生成的主要调节因子,有关canstatin蛋白在CNV生成中作用的报道较少. 目的 探讨重组canstatin蛋白对小鼠碱烧伤后CNV的抑制作用及其机制. 方法 采用1 mol/L NaOH溶液烧伤小鼠右眼角膜的方法建立40只雌性BALB/c小鼠碱烧伤CNV动物模型,然后用随机数字表法将模型动物随机分为对照组和实验组,每组20只小鼠20只眼,均取右眼为实验眼,分别给予生理盐水和5 mg/L重组canstatin蛋白溶液点眼,每日4次.分别于碱烧伤后1、3、7、14 d在裂隙灯显微镜下检查CNV生长情况,并测量和计算CNV面积.分别于上述时间点获取小鼠角膜组织,采用Westernblot法检测各组小鼠角膜中VEGF和bFGF蛋白的表达,用增强化学发光法(ECL)对检测结果进行分析. 结果 裂隙灯显微镜检查显示,实验组碱烧伤后3、7、14 d CNV面积分别为(1.98±0.31)、(6.21±0.44)和(9.83±0.72) mm2,明显低于相应时间点对照组的(2.92±0.41)、(8.04±0.56)和(11.78±0.84) mm2,差异均有统计学意义(t3 d=4.332,P=0.005；t7 d=11.729,P=0.000；t14d=14.562,P=0.000).Western blot检测结果显示,角膜碱烧伤后实验组1、3、7、14 d VEGF蛋白在角膜组织中表达的灰度值明显高于对照组,差异均有统计学意义(t1d =-3.980,P＜0.001；t3 d=-10.020,P＜0.001；t7d=--4.355,P＜0.001；t14d =-8.156,P＜0.001)；实验组bFGF蛋白在角膜组织中表达的灰度值明显高于对照组,差异均有统计学意义(t1d=-3.488,P＜0.001；t3d=-2.124,P=0.013；t7d =-1.977,P=0.028;t14 d =-4.542,P＜0.001). 结论 重组canstatin蛋白通过下调VEGF蛋白和bFGF蛋白在角膜组织中的表达抑制小鼠碱烧伤后CNV的形成.
揹景 堿燒傷後角膜新生血管(CNV)形成可引起緻盲性眼病,最近研究錶明血管內皮生長因子(VEGF)和堿性成纖維細胞生長因子(bFGF)是誘導血管生成的主要調節因子,有關canstatin蛋白在CNV生成中作用的報道較少. 目的 探討重組canstatin蛋白對小鼠堿燒傷後CNV的抑製作用及其機製. 方法 採用1 mol/L NaOH溶液燒傷小鼠右眼角膜的方法建立40隻雌性BALB/c小鼠堿燒傷CNV動物模型,然後用隨機數字錶法將模型動物隨機分為對照組和實驗組,每組20隻小鼠20隻眼,均取右眼為實驗眼,分彆給予生理鹽水和5 mg/L重組canstatin蛋白溶液點眼,每日4次.分彆于堿燒傷後1、3、7、14 d在裂隙燈顯微鏡下檢查CNV生長情況,併測量和計算CNV麵積.分彆于上述時間點穫取小鼠角膜組織,採用Westernblot法檢測各組小鼠角膜中VEGF和bFGF蛋白的錶達,用增彊化學髮光法(ECL)對檢測結果進行分析. 結果 裂隙燈顯微鏡檢查顯示,實驗組堿燒傷後3、7、14 d CNV麵積分彆為(1.98±0.31)、(6.21±0.44)和(9.83±0.72) mm2,明顯低于相應時間點對照組的(2.92±0.41)、(8.04±0.56)和(11.78±0.84) mm2,差異均有統計學意義(t3 d=4.332,P=0.005；t7 d=11.729,P=0.000；t14d=14.562,P=0.000).Western blot檢測結果顯示,角膜堿燒傷後實驗組1、3、7、14 d VEGF蛋白在角膜組織中錶達的灰度值明顯高于對照組,差異均有統計學意義(t1d =-3.980,P＜0.001；t3 d=-10.020,P＜0.001；t7d=--4.355,P＜0.001；t14d =-8.156,P＜0.001)；實驗組bFGF蛋白在角膜組織中錶達的灰度值明顯高于對照組,差異均有統計學意義(t1d=-3.488,P＜0.001；t3d=-2.124,P=0.013；t7d =-1.977,P=0.028;t14 d =-4.542,P＜0.001). 結論 重組canstatin蛋白通過下調VEGF蛋白和bFGF蛋白在角膜組織中的錶達抑製小鼠堿燒傷後CNV的形成.
배경 감소상후각막신생혈관(CNV)형성가인기치맹성안병,최근연구표명혈관내피생장인자(VEGF)화감성성섬유세포생장인자(bFGF)시유도혈관생성적주요조절인자,유관canstatin단백재CNV생성중작용적보도교소. 목적 탐토중조canstatin단백대소서감소상후CNV적억제작용급기궤제. 방법 채용1 mol/L NaOH용액소상소서우안각막적방법건립40지자성BALB/c소서감소상CNV동물모형,연후용수궤수자표법장모형동물수궤분위대조조화실험조,매조20지소서20지안,균취우안위실험안,분별급여생리염수화5 mg/L중조canstatin단백용액점안,매일4차.분별우감소상후1、3、7、14 d재렬극등현미경하검사CNV생장정황,병측량화계산CNV면적.분별우상술시간점획취소서각막조직,채용Westernblot법검측각조소서각막중VEGF화bFGF단백적표체,용증강화학발광법(ECL)대검측결과진행분석. 결과 렬극등현미경검사현시,실험조감소상후3、7、14 d CNV면적분별위(1.98±0.31)、(6.21±0.44)화(9.83±0.72) mm2,명현저우상응시간점대조조적(2.92±0.41)、(8.04±0.56)화(11.78±0.84) mm2,차이균유통계학의의(t3 d=4.332,P=0.005；t7 d=11.729,P=0.000；t14d=14.562,P=0.000).Western blot검측결과현시,각막감소상후실험조1、3、7、14 d VEGF단백재각막조직중표체적회도치명현고우대조조,차이균유통계학의의(t1d =-3.980,P＜0.001；t3 d=-10.020,P＜0.001；t7d=--4.355,P＜0.001；t14d =-8.156,P＜0.001)；실험조bFGF단백재각막조직중표체적회도치명현고우대조조,차이균유통계학의의(t1d=-3.488,P＜0.001；t3d=-2.124,P=0.013；t7d =-1.977,P=0.028;t14 d =-4.542,P＜0.001). 결론 중조canstatin단백통과하조VEGF단백화bFGF단백재각막조직중적표체억제소서감소상후CNV적형성.
Background Alkali burn-induced corneal neovascularization(CNV) usually leads to blindness.Recently,study determined that vascular endothelial growth factor(VEGF) and basic fibroblast growth factor(bFGF) were the main regulating factors inducing angiogenesis,and canstatin proteins can inhibit the growth of VEGF and bFGF and thereby inhibit CNV growth.Objective The present study was to investigate the effect of recombinant canstatin protein on mouse CNV induced by alkali burn and its mechanism.Methods CNV models were induced in the right eyes of 40 female BALB/c mice by sticking the 2.0 mm filtering paper with 1 mol/L NaOH at the central cornea for 10 seconds.The animals then were randomized into two groups.Recombinant canstatin protein drops(5 mg/L) was topically administered 4 times daily in the mice of the experimental group,and normal saline solution was used at the same way in the control group.Corneas of the mice were examined under the slit lamp to calculate the CNV area 1 day,3,7,14 days after modeling.The mice were sacrificed at above time points and corneas were obtained.The expressions of VEGF protein and bFGF protein in cornea were detected by Western blot,and the results were analyzed by enhanced chemiluminescence(ECL).The use of the experimental animals complied with the Instructive Notions with Respect to Caring for Laboratory Animals by State Ministry of Science and Technology.Results In 3,7 and 14 days after establishment of models,the area of CNV was (1.98-0.31) mm2,(6.21 ±0.44) mm2 and (9.83±0.72) mm2 in the experimental group,and that in the control group was (2.92 ± 0.41) mm2,(8.04 ± 0.56) mm2 and (11.78 ±0.84) mm2,showing significant reduce in the mice treated with recombinant canstatin protein(t3d =4.332,P=0.005 ;t7 d =11.729,P =0.000 ;t14 d =14.562,P =0.000).Western blot analysis showed that there was significant increase in the gray scale of VEGF protein 1 day,3,7,14 days following alkali burn in the experimental group compared with the control group(t1 d =-3.980,P＜0.001 ;t3d =-10.020,P＜0.001 ;t7d =-4.355,P＜0.001 ;t14 d =-8.156,P＜0.001),and the gray scale of bFGF was significantly ascent at various time points in the the experimental in comparison with the control groups (t1 d =-3.488,P＜0.001 ; t3 d =-2.124,P =0.013; t7 d =-1.977,P =0.028; t14 d =-4.542,P＜0.001).Conclusions Topical application of recombinant canstatin protein drops inhibits CNV growth induced by alkali burn by down-regulating the expressions of VEGF and bFGF proteins.
