摘要
目的建立SD大鼠后囊膜混浊(posterior capsular opacification,PCO)动物模型,检测Rho/Rock通路中的关键因子Rho A、Rho激酶Ⅰ(Rho associated coiledcoil forming protein kinase-Ⅰ,RockⅠ)和肌球蛋白轻链(phosphorylated myosin light chain,p-MLC)在PCO中的表达。探讨Rho/Rock通路在PCO形成过程中的作用。方法选用健康SD大鼠30只,随机分为对照组(0d)和实验组(3 d、7 d、14 d、28 d),每个时间点各6只。所有大鼠(右眼)均行晶状体囊外摘出术,显微镜下观察不同时间点术眼前节的炎症反应及PCO发生情况。ELISA法检测不同时间点房水中TGF-β的表达。处死大鼠后摘除眼球行组织病理学检查,免疫组织化学和Western Blot法检测Rho A、RockⅠ、p-MLC在晶状体后囊上的表达。结果随着术后观察时间的推移,PCO混浊程度逐渐增加,HE染色显示晶状体上皮细胞不同程度的增生、移行。对照组术后0 d、3 d、7 d、14 d、28 d房水中TGF-β的表达量分别为(37.28±1.31)μg·L-1、(40.29±3.17)μg·L-1、(42.12±3.45)μg·L-1、(60.42±4.75)μg·L-1、(68.69±8.06)μg·L-1,实验组各时间点与对照组相比,差异有统计学意义(F=137.158,P<0.05);不同时间点两两相比,差异均有统计学意义(均为P<0.05)。术后0 d、3 d、7 d、14 d和28 d晶状体后囊膜上Rho A蛋白相对表达量(Rho A/GAPDH)分别为1.00±0.00、1.49±0.01、1.57±0.03、1.77±0.02和1.97±0.02,实验组各时间点与对照组相比,差异有统计学意义(F=94.546,P<0.05);不同时间点两两相比,差异均有统计学意义(均为P<0.05)。RockⅠ蛋白的表达量(RockⅠ/GAPDH)分别为1.00±0.00、0.35±0.12、1.42±0.14、1.78±0.09和1.95±0.14,实验组各时间点与对照组相比,差异有统计学意义(F=30.942,P<0.05);不同时间点两两相比,差异均有统计学意义(均为P<0.05)。p-MLC蛋白的相对表达量(p-MLC/GAPDH)分别为1.00±0.00、1.44±0.08、1.47±0.13、1.63±0.10和2.12±0.20,实验组各时间点与对照组相比,差异有统计学意义(F=34.313,P<0.05);不同时间点两两相比,差异均有统计学意义(均为P<0.05)。结论 Rho A、RockⅠ、p-MLC在PCO形成过程中表达逐渐增多,Rho/Rock信号转导通路可能与PCO的发生、发展有关。
Objective To establish animal model of posterior capsular opacifica- tion （PCO） in SD rats and detect the expressions of key factors of Rho/Rock pathway in PCO tissue, including RhoA, Rho associated coiledcoil forming protein kinase- I （Rock I ） and phosphorylated myosin light chain （p-MLC）. Methods Thirty healthy SD rats were randomly divided into control group （0 day） and experimental group （3 days,7 days, 14 days,28 days）. Extracapsular extraction was performed in all the rats （ right eyes）. Anterior segment inflammation and development of PCO were observed by slit lamp microscope after surgery. The expression of transfer growth factor-β （TGF-β） in aqueous humor was detected by ELISA. Then the rats were killed and the right eyes were removed for histopathological examination. Immunohistochemistry and Western Blot were performed to detect the expression of RhoA, Rock I ,p-MLC in posterior cap- stiles. Results The posterior capsular opacification developed gradually after surgery. HE staining showed different degrees of proliferation and migration of lens epithelial cells. After operation, the content of TGF-β in the aqueous humor at 0 day, 3 days, 7 days,14 days,28 days were （37.28 ± 1.31 ） μg · L^-1 , （40.29 ± 3.17） μg · L^-1 , （42. 12 ± 3.45） μg · L^-1 ,（60.42 ±4.75） μg · L^-1 ,（68.69 ±8.06） μg · L^-1 ,respectively,the differ- ence of TGF-[3 content was statistically significant between the experimental group and control group （F= 137. 158 ,P =0. 000） ,and the TGF-β content was statistically signifi- cant among different experiment groups （all P 〈 0.05 ）. The expression of RhoA（ RhoA/GAPDH） at 0 day,3 days,7 days, 14 days,28 days were 1.00 ± 0.00,1.49 ±0.01,1.57 ± 0.03,1.77± 0.02,1.97 ± 0.02, re- spectively, the difference of RhoA content was statistically significant between the experimental group and control group （ F =94.545 ,P =0. 000） ,and the RhoA content was statistically significant among different experiment groups（ all P 〈0.05 ）. The expression of Rock Ⅰ （ Rock I/GAPDH） at 0 day,3 days,7 days, 14 days,28 days were 1.00 ±0. 00,0.35 ± 0.12,1.42 ± 0.14,1.78± 0.09,1.95 ± 0.14, respectively, the difference of Rock Ⅰ content was statistically significant between the experi- mental group and control group （F = 30.942 ,P = 0.000）, and the Rock I content was statistically significant among different experiment groups （ all P 〈0.05）. The expression of p-MLC（p-MLC/GAPDH） at 0 day,3 days,7 days,14 days,28 days were 1.00 ±0.00,1.44 ±0.08,1.47 ±0.13,1.53 ±0.10,2.12 ±0.20, respectively, the difference of p-MLC content was statistically significant between the experimental group and control group （F = 34.313 ,P = 0. 000 ）, and the p-MLC content was statisti- cally significant among different experiment groups （all P 〈 0.05）. Conclusion The expressions of RhoA, Rock I and p- MLC gradually increase during the development of PCO, indicating that the Rho/Rock signal pathway may be involved in PCO formation.
出处
《眼科新进展》
CAS
北大核心
2016年第2期105-109,共5页
Recent Advances in Ophthalmology
基金
河南省科学技术重点攻关项目(编号:142102310053)~~
作者简介
作者简介：杜文文，女，1988年7月出生，河南郑州人，在读硕士研究生。联系电话：18538030520；E-mail：441325974@qq．com
通讯作者：张凤妍，E-mail：zhang-fengyanx@aliyun．com.
