微囊化内皮抑素球周注射的实验研究  

Experimental research of Endostatin microcapsulized by periocular injection

作　　者：詹文芳[1] 蔡善君[1] 刘锐[1] 谢兵[1] 李红[1] 宿罡[1] 

机构地区：[1]遵义医学院附属医院眼科,563003

出　　处：《中华眼科杂志》2009年第11期1027-1032,共6页Chinese Journal of Ophthalmology

基　　金：基金项目：贵州省科技厅自然基金资助项目（黔科合J字[2005]2061号）

摘　　要：目的检测乳化-内部凝胶化制得的海藻酸钙-壳聚糖（CAC）内皮抑素微囊的体外释放情况，大鼠眼球周注射后眼内组织分布情况及生物相容性。方法实验研究。采用乳化-内部凝胶化方式制备微囊，紫外分光光度法、考马斯亮蓝法检测内皮抑素囊外释放情况；32只SD大鼠随机分成4组，每组8只。A组：微囊化ES组，球周注射微囊化内皮抑素20μl（2．5g/L）；B组：单纯内皮抑素组，球周注射内皮抑素蛋白20μl（2．5g／L）；C组：空微囊组，球周注射空微囊20μl；D组：生理盐水组，球周注射生理盐水20μl。免疫组织化学法、ELISA法、Western—blot法检测内皮抑素分布、浓度和蛋白表达；观察SD大鼠的日常活动，进食情况和体重，以断颈法将大鼠处死取心、肝、脾、肺、。肾及球周组织（球周注射部位周围组织）行病理学检查。采用独立样本设计定量资料t检验对数据进行分析。结果十二烷基硫酸钠-聚丙烯酰胺凝胶电泳检测可见内皮抑素蛋白条带，3、7、14、21d各时点上清液中内皮抑素蛋白质浓度逐渐增加。A组：球周注射后7、14d，内外核层和RPE层见内皮抑素蛋白阳性表达；B组：7d时，以上组织可见内皮抑素蛋白阳性表达，14d后无蛋白阳性表达；C组和D组7、14d以上组织均未见内皮抑素蛋白阳性表达，表明内皮抑素蛋白经CAC微囊化后通过球周注射可以穿过巩膜壁进入球内。A组7、14d，眼球组织匀浆内皮抑素蛋白浓度分别为每只眼（63．16±7．64）μg/L和（33．2±5．77）μg/L，而B组分别为（33．2±2．89）μg/L，（15．73±2．08）μg/L，A组明显大于B组且差异具有统计学意义（t=6．364、4．920，P=0．003、0．008）；大鼠日常活动和进食情况均正常，14d后病理学检查心肝、脾、肺、肾及球周组织未见异常。结论微囊化内皮抑素具有缓控释功能，球周注射微囊化内皮抑素，Objective To detect the release of endostatin in microcapsules of calcium alginate gel by emulsification-internal gelatification technology, to observe the distribution of Endostatin microcapsulized in the eye by periocular injection and biocompatibility. Methods The calcium alginate gel microcapsules encapsulating endostatin were prepared by emulsification-internal gelatification technology, the release of endostatin in microcapsules in vitro was examined by uv-Spectrophotometry. 32 SD mice were divided into randomly four groups： group A, periocular injection of endostatin microcapsulized 20μl（2.4 g/L） ; group B, periocular injection of endostatin protein 20 μl （2. 5 g/L）; group C, periocular injection of null- Microcapsules 20 μl ;group D, periocular injection of saline 20μl. Western blot, immunohistochemical and enzyme-linked immunosorbent assay were used. Protein expression and pathology of the heart, liver, spleen, lung, kidney and periocular tissue were examed. Daily activities and eating condition were observed, t-test was wsed to analyze the data. Results SDS polyacrylamidedel electrophoresis showed strap of ES protein. Concentration of ES protein in superior schedule fluid gradually increased at 3,7,14,21 days. A group： ES protein expression were observed in inner nuclear layer,outer nuclear layer and RPE lamina after periocular injection 7 or 14 days. B group： ES protein expression were observed in above tissues at 7 days, but not at 14 days. C and D group： ES protein expression were not observed in above tissues at 7 or 14 days. The results showed that Endostatin microcapsulized was able to pass through the sclera and spread out in various retinal tissues. Concertration of endostatin in the homogenates of eyeball was （63.16 ± 7.64）μg · L-^-1 · eye^-1 and （ 33.2 ± 5.77 ） μg · L^- 1 ·eye^ -1 respectively after one and two weeks in A group, but （33.2 ± 2. 89 ） μg · L^- 1 ·eye^ -1 and （ 15.73 ±2.08 ） μg · L^- 1 ·eye^ -1, The concertration of endostatin w

关 键 词：胶囊 壳聚糖 内皮抑素类 投药 局部 

分 类 号：R686[医药卫生—骨科学]