摘要
背景:目前视网膜缺血再灌注损伤仍没有一个理想的治疗方案,组织细胞工程尤其是干细胞工程的兴起为视网膜损伤的治疗提供了新思路。目的:以视网膜电图b波、视网膜形态学、神经节细胞密度及内层视网膜厚度为指标,观察骨髓间充质干细胞视网膜下移植对大鼠视网膜缺血再灌注损伤的影响。设计、时间及地点:完全随机对照实验,于2007-01/12在潍坊医学院眼科研究所完成。材料:选择清洁级Wistar成年大鼠48只。方法:采用密度梯度离心法培养骨髓间充质干细胞。将48只大鼠按随机数字表法分为4组,每组12只。缺血再灌注损伤组采用Daniel法制备视网膜缺血再灌注损伤模型;缺血再灌注损伤后24h,骨髓间充质干细胞移植组视网膜下移植Hoechst33324标记的骨髓间充质干细胞2×108L-1;磷酸盐缓冲液组输注等量的磷酸盐缓冲液;正常对照组不做任何处理。主要观察指标:缺血再灌注损伤后6h,24h,3d,7d,14d,28d动态检测各组视网膜电图b波;缺血再灌注损伤后28d荧光显微镜下观察骨髓间充质干细胞移植组视网膜Hoechst33324阳性细胞表达情况;缺血再灌注损伤后28d处死各组大鼠,分别对视网膜进行苏木精-伊红、尼氏染色观察,检测并比较各组神经节细胞密度及视网膜内层厚度的变化。结果:缺血再灌注损伤后各时间点缺血再灌注损伤组大鼠视网膜电图b波逐渐降低,均显著低于正常对照组(P<0.01);缺血再灌注损伤后28d骨髓间充质干细胞移植组视网膜电图b波显著高于缺血再灌注损伤组(P<0.01)。缺血再灌注损伤后28d骨髓间充质干细胞移植组视网膜各层均可见Hoechst阳性细胞,核染成蓝色。缺血再灌注损伤组视网膜神经节细胞密度与内层视网膜厚度显著小于正常对照组与骨髓间充质干细胞移植组(P<0.05～0.01),缺血再灌注损伤组与磷酸盐缓冲液组差异无显著性意义(P>0.05)。结论:视网膜缺血再灌注损伤后24h骨髓间充质干细胞视网膜下移植,可以改善视网膜电图b波、增加神经节细胞密度及内层视网膜厚度,从而达到减轻视网膜缺血再灌注损伤的目的。
BACKGROUND： At present, there is not an ideal therapeutic regimen for retinal ischemia-reperfusion injury. Histiocyte engineering, especially the emerging of stem cell engineering, provides a new idea for retinal injury. OBJECTIVE： To observe the effect of bone marrow mesenchymal stem cells （MSCs） on retinal ischemia-reperfusion injury after subretinal transplantation indexed by the b-wave of electroretinogram （ERG）, retinal morphology, density of retinal ganglial cells （RGC） and thickness of inner retina. DESIGN, TIME AND SETTING： The randomized controlled study was performed at Eye Institute of Weifang Medical University from January to December in 2007. MATERIALS： Forty-eight Wistar rats of clean grade were used in the study. METHODS： Forty-eight rats were randomly divided into the normal control group （n= 12）, retinal ischemia-reperfusion injury group （n=12）, MSCs transplantation group （n=12） and phosphate buffer saline （PBS） group （n=12）. MSCs were obtained from healthy rats by the density gradient centrifugation method. The retinal ischemia-reperfusion injury model was made by Daniel＇s method. Twenty-four hours after injury, MSCs labeled by Hochest33324 （2×10^5/mL） and PBS were interfused into the model by subretinal transplantation. There was no treatment in the normal control group. MAIN OUTCOME MEASURES： b-wave amplitude of ERG was dynamically examined in each group at 6 hours, 24 hours, 72 hours, 7 days, 14 days and 28 days after ischemia-reperfusion. Twenty-four days after ischemia-reperfusion, the positive expression of retina Hoechst33324 was observed under a fluorescence microscope. Haematoxylin-eosin staining and Nissl staining were used to observe the changes density of RGCs and thickness of the inner retina. RESULTS： b-wave amplitude of ERG decreased gradually in the retinal ischemi-reperfusion injury group at different time points after ischemia-reperfusion, which was significantly lower compared with the normal control group （P 〈 0.01）. And b-wave in the transplantation group was significantly greater compared with the retinal ischemia-reperfusion injury group at 28 days after ischemia-reperfusion （P 〈 0.01）.The Hochest positive cells observed in the retina and the nuclear were stained blue 28 days after ischemia-reperfusion. The RGC and inner retina in the retinal ischemia-reperfusion injury group was significantly lower than that in the normal control group and in the MSCs transplantation group （P 〈 0.05-0.01）. There was no significant difference between the retinal ischemia-reperfusion injury group and PBS group （P 〉 0.05）. CONCLUSION： The subretinal transplantation of bone MSCs 24 hours after ischemia-reperfusion can improve b-wave of ERG, increase density of RGCs and thickness of inner retina, to alleviate the retinal ischemia-reperfusion injury.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2008年第47期9261-9264,共4页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
"骨髓间充质干细胞移植对视网膜缺血再灌注损伤功能修复作用的实验研究"山东省自然科学基金项目(Y2007C084)~~
