中华实验眼科杂志
中華實驗眼科雜誌
중화실험안과잡지
CHINESE JOURNAL OF EXPERIMENTAL OPHTHALMOLOGY
2013年
1期
65-69
,共5页
于靖%崔尘%赵红梅%王克生
于靖%崔塵%趙紅梅%王剋生
우정%최진%조홍매%왕극생
胰岛素样生长因子结合蛋白%增生性玻璃体视网膜病变%大鼠/模型%视网膜
胰島素樣生長因子結閤蛋白%增生性玻璃體視網膜病變%大鼠/模型%視網膜
이도소양생장인자결합단백%증생성파리체시망막병변%대서/모형%시망막
Insulin-like growth factor binding protein%Proliferative vitreoretinopathy%Rat/ model%Retina
背景 增生性玻璃体视网膜病变(PVR)是导致视网膜脱离复位手术失败的主要原因之一.PVR玻璃体蛋白质组学研究发现,胰岛素样生长因子结合蛋白-6(IGFBP-6)是PVR患者玻璃体和血清中的特异蛋白质,且其在严重PVR患者中的含量明显高于轻度PVR. 目的 探讨IGFBP-6在PVR大鼠模型中的表达变化. 方法 选取7周龄SPF级雄性Wistar大鼠70只,采用抛硬币法随机将动物分为PVR模型组34只和对照组36只.PVR模型组大鼠左眼玻璃体腔内注入透明质酸酶1U(商品单位),注射后10 min待玻璃体液化后,玻璃体腔内再注入1×106视网膜色素上皮(RPE)细胞悬液5μl和1×10 7富含血小板的血浆5μl;对照组大鼠以同样的方法注射无菌生理盐水10μl.各组大鼠玻璃体腔注射后1、2、3、4周行眼底检查并按Francine标准对PVR进行分级.分别于造模后1、2、4、8周处死各组大鼠,抽取动物心脏血,切取肝脏和视网膜,并制备切片,采用实时荧光定量PCR法检测大鼠肝脏组织和视网膜组织中IGFBP-6 mRNA的表达,采用ELISA法检测大鼠玻璃体和血清中IGFBP-6的质量浓度,对各组大鼠的测量指标进行比较. 结果 利用实时荧光定量PCR法分别从肝脏和视网膜中提取高纯度的IGFBP-6 mRNA.PVR模型组大鼠造模后第4周视网膜中IGFBP-6 mRNA含量为(3.79±1.33) ×10-4,明显低于对照组的(8.32±2.96) ×10-4,差异有统计学意义(t=3.420,P＜0.01)；PVR模型组大鼠造模后第4周视网膜中IGFBP-6 mRNA含量明显低于造模后第1、2、8周,差异均有统计学意义(P＜0.05),而PVR模型组大鼠造模后各时间点肝脏中IGFBP-6 mRNA含量与对照组比较以及PVR模型组大鼠组内各时间点间的比较差异均无统计学意义(P＞0.05).PVR 1、2、3级组大鼠视网膜中IGFBP-6 mRNA含量明显低于对照组,差异有统计学意义(P＞0.05),但不同等级的PVR大鼠间差异无统计学意义(P＞0.05).ELISA检测结果显示PVR 1、2、3级组大鼠玻璃体中IGFBP-6的质量浓度分别为(221.00±19.32)、(229.63±18.89)和(225.70±26.71) μg/L,明显高于对照组的(173.25±21.11) μg/L,差异有统计学意义(P＜0.05),但PVR组各级之间比较差异无统计学意义(t=1.240、1.46,P＞0.05).PVR模型组大鼠和血清中IGFBP-6质量浓度高于对照组大鼠,4周时两组间差异有统计学意义(P＜0.05). 结论 PVR大鼠玻璃体和血清中IGFBP-6质量浓度均增加,玻璃体中增加的IGFBP-6可能来源于局部的自分泌.
揹景 增生性玻璃體視網膜病變(PVR)是導緻視網膜脫離複位手術失敗的主要原因之一.PVR玻璃體蛋白質組學研究髮現,胰島素樣生長因子結閤蛋白-6(IGFBP-6)是PVR患者玻璃體和血清中的特異蛋白質,且其在嚴重PVR患者中的含量明顯高于輕度PVR. 目的 探討IGFBP-6在PVR大鼠模型中的錶達變化. 方法 選取7週齡SPF級雄性Wistar大鼠70隻,採用拋硬幣法隨機將動物分為PVR模型組34隻和對照組36隻.PVR模型組大鼠左眼玻璃體腔內註入透明質痠酶1U(商品單位),註射後10 min待玻璃體液化後,玻璃體腔內再註入1×106視網膜色素上皮(RPE)細胞懸液5μl和1×10 7富含血小闆的血漿5μl;對照組大鼠以同樣的方法註射無菌生理鹽水10μl.各組大鼠玻璃體腔註射後1、2、3、4週行眼底檢查併按Francine標準對PVR進行分級.分彆于造模後1、2、4、8週處死各組大鼠,抽取動物心髒血,切取肝髒和視網膜,併製備切片,採用實時熒光定量PCR法檢測大鼠肝髒組織和視網膜組織中IGFBP-6 mRNA的錶達,採用ELISA法檢測大鼠玻璃體和血清中IGFBP-6的質量濃度,對各組大鼠的測量指標進行比較. 結果 利用實時熒光定量PCR法分彆從肝髒和視網膜中提取高純度的IGFBP-6 mRNA.PVR模型組大鼠造模後第4週視網膜中IGFBP-6 mRNA含量為(3.79±1.33) ×10-4,明顯低于對照組的(8.32±2.96) ×10-4,差異有統計學意義(t=3.420,P＜0.01)；PVR模型組大鼠造模後第4週視網膜中IGFBP-6 mRNA含量明顯低于造模後第1、2、8週,差異均有統計學意義(P＜0.05),而PVR模型組大鼠造模後各時間點肝髒中IGFBP-6 mRNA含量與對照組比較以及PVR模型組大鼠組內各時間點間的比較差異均無統計學意義(P＞0.05).PVR 1、2、3級組大鼠視網膜中IGFBP-6 mRNA含量明顯低于對照組,差異有統計學意義(P＞0.05),但不同等級的PVR大鼠間差異無統計學意義(P＞0.05).ELISA檢測結果顯示PVR 1、2、3級組大鼠玻璃體中IGFBP-6的質量濃度分彆為(221.00±19.32)、(229.63±18.89)和(225.70±26.71) μg/L,明顯高于對照組的(173.25±21.11) μg/L,差異有統計學意義(P＜0.05),但PVR組各級之間比較差異無統計學意義(t=1.240、1.46,P＞0.05).PVR模型組大鼠和血清中IGFBP-6質量濃度高于對照組大鼠,4週時兩組間差異有統計學意義(P＜0.05). 結論 PVR大鼠玻璃體和血清中IGFBP-6質量濃度均增加,玻璃體中增加的IGFBP-6可能來源于跼部的自分泌.
배경 증생성파리체시망막병변(PVR)시도치시망막탈리복위수술실패적주요원인지일.PVR파리체단백질조학연구발현,이도소양생장인자결합단백-6(IGFBP-6)시PVR환자파리체화혈청중적특이단백질,차기재엄중PVR환자중적함량명현고우경도PVR. 목적 탐토IGFBP-6재PVR대서모형중적표체변화. 방법 선취7주령SPF급웅성Wistar대서70지,채용포경폐법수궤장동물분위PVR모형조34지화대조조36지.PVR모형조대서좌안파리체강내주입투명질산매1U(상품단위),주사후10 min대파리체액화후,파리체강내재주입1×106시망막색소상피(RPE)세포현액5μl화1×10 7부함혈소판적혈장5μl;대조조대서이동양적방법주사무균생리염수10μl.각조대서파리체강주사후1、2、3、4주행안저검사병안Francine표준대PVR진행분급.분별우조모후1、2、4、8주처사각조대서,추취동물심장혈,절취간장화시망막,병제비절편,채용실시형광정량PCR법검측대서간장조직화시망막조직중IGFBP-6 mRNA적표체,채용ELISA법검측대서파리체화혈청중IGFBP-6적질량농도,대각조대서적측량지표진행비교. 결과 이용실시형광정량PCR법분별종간장화시망막중제취고순도적IGFBP-6 mRNA.PVR모형조대서조모후제4주시망막중IGFBP-6 mRNA함량위(3.79±1.33) ×10-4,명현저우대조조적(8.32±2.96) ×10-4,차이유통계학의의(t=3.420,P＜0.01)；PVR모형조대서조모후제4주시망막중IGFBP-6 mRNA함량명현저우조모후제1、2、8주,차이균유통계학의의(P＜0.05),이PVR모형조대서조모후각시간점간장중IGFBP-6 mRNA함량여대조조비교이급PVR모형조대서조내각시간점간적비교차이균무통계학의의(P＞0.05).PVR 1、2、3급조대서시망막중IGFBP-6 mRNA함량명현저우대조조,차이유통계학의의(P＞0.05),단불동등급적PVR대서간차이무통계학의의(P＞0.05).ELISA검측결과현시PVR 1、2、3급조대서파리체중IGFBP-6적질량농도분별위(221.00±19.32)、(229.63±18.89)화(225.70±26.71) μg/L,명현고우대조조적(173.25±21.11) μg/L,차이유통계학의의(P＜0.05),단PVR조각급지간비교차이무통계학의의(t=1.240、1.46,P＞0.05).PVR모형조대서화혈청중IGFBP-6질량농도고우대조조대서,4주시량조간차이유통계학의의(P＜0.05). 결론 PVR대서파리체화혈청중IGFBP-6질량농도균증가,파리체중증가적IGFBP-6가능래원우국부적자분비.
Background Proliferative vitreoretinopathy (PVR) is one of the major causes of retinal detachment surgery failure.Based on proteomic studies of PVR vitreous,the insulin-like growth factor binding protein-6 (IGFBP-6) protein was specifically expressed in the vitreous and serum of PVR patients.Furthermore,its expression level is higher in the vitreous and serum in severe PVR patients than that in mild PVR patients.Objective This experiment was to detect the expression of IGFBP-6 in a PVR rat model.Methods Seventy 7-week old male SPF Wistar rats were included and were randomized into the PVR model group and control group.A mixture of RPE-J cell suspension(5 μl) and platelet-rich plasma (5 μl) was intravitreally injected in the left eyes of adult Wistar rats to establish the PVR model,and normal saline solution was administered in the same way in the control group.The rat eyes were clinically examined 1 week,2,3 and 4 weeks after injection,and PVR was graded based on the criteria of Francine.The animals were sacrificed after 1 week,2,4 or 8 weeks for the preparation of retinal sections and liver extraction.Expression levels of IGFBP-6 mRNA in the rat retina and liver were assayed by real-time Q-PCR.The expression of IGFBP-6 protein in the rat serum and vitreous was detected by ELISA.The use of animals complied with the Regulations for the Administration of Affairs Concerning Experimental Animals by State Science and Technology Commission.Results Purified IGFBP-6 RNA was extracted from the liver and retina of Wistar rat and quantified by real-time Q-PCR.The expression level of IGFBP-6 mRNA in retina was (3.79± 1.33) × 10-4 in the PVR model rats,showing a significant decline in comparison with the control rats with a level of(8.32±2.96) × 10 4,4 weeks after injection (t =3.42,P＜0.01).The expression of IGFBP-6 mRNA in the 4th week was significantly lower than that of 1 week,2 or 8 weeks after the establishment of the PVR model(P＜0.05).No significant difference was found in the IGFBP-6 mRNA level in the liver between the PVR group and control group(27.60± 14.01 × 10 4 vs.25.01 ± 12.04 ×10-4,respectively),as well as among the different time points(P＞0.05).IGFBP-6 mRNA content in the retina was significantly reduced in grades 1,2 or 3 of the PVR groups compared with the control group(P＞0.05),but there was no significant difference among the different grades of PVR groups (P＞0.05).Concentrations of IGFBP-6 protein in grades 1,2 and 3 of the PVR model group were (221.00 ± 19.32),(229.63 ± 18.89) and (225.70 ± 26.71) μg/L,with a significant elevation in comparison with (173.25 ±21.11) μg/L of the control group (t =2.14,P＜0.05).However,there was no significant change among the different grades of PVR groups(t=1.24,1.46,P＞0.05).The concentrations of IGFBP-6 protein in the vitreous and serum were higher in PVR rat samples (vitreous:225.44±19.36 μg/L;serum:108.48 ± 15.78 μg/L) than in control rats (vitreous:173.25 ± 21.11 μg/L,serum:95.96 ±17.40 μg/L)(P＜0.05).Conclusions The concentrations of IGFBP-6 protein in the vitreous and serum increase in PVR rats.The results indicate that the increased IGFBP-6 in the vitreous might be a localized autocrine secretion of the eye.
