重组腺病毒介导疱疹病毒胸苷激酶基因在昆明小鼠体内分布行为研究  

In vivo biodistribution study of recombinant adenovirus-mediated herpes simplex virus thymidine kinase gene in Kunming mice

作　　者：吕鹏[1] 王越[2] 周剑峰[3] 靳洪涛[1] 王灵芝[1] 包捷[1] 李静莉[2] 李海艳[1] 卢运萍[1] 黄明芳[1] 李成合[1] 刘岩[1] 董坚 薛鹏 王爱平[1] 魏金锋[1] 

机构地区：[1]中国医学科学院北京协和医学院新药安全评价研究中心,北京100050 [2]中国食品药品检定研究院,北京100050 [3]华中科技大学同济医学院附属同济医院,武汉43003 [4]美国哈姆纳健康医学研究所,北卡罗来纳27709

出　　处：《药物分析杂志》2014年第4期595-601,共7页Chinese Journal of Pharmaceutical Analysis

基　　金：supported by Shenzhen Tiandakang gene Engineering Co. Ltd and in part by Beijing Union - Genius Pharmaceutical Technology Development Co. Ltd.

摘　　要：目的:开展重组腺病毒-胸苷激酶基因制剂(adenovirus-mediated delivery of herpes simplex virus thymidine kinase gene,ADV-tk)局部或静脉用药后于动物体内的分布行为的研究。方法:分别采用绿色荧光蛋白(green fluorescent protein,GFP)和原位杂交法检测分析重组腺病毒及胸苷激酶于肝脏、脾脏、淋巴结、肠系膜、肾上腺、胸腺、肾脏、甲状腺、前列腺、卵巢等组织分布的情况。结果判断采用免疫组织化学评分法(immuno-histochemical scores,IHS),并结合阳性细胞百分比及阳性细胞染色强弱2个方面进行评价。结果:(1)3种不同给药途径的组织分布特征具有相似性,ADV或tk阳性表达的组织脏器从强到弱分布依次为:肝脏、脾脏、淋巴结、肠系膜、肾上腺、胸腺、肾脏、甲状腺、前列腺、卵巢。ADV或tk mRNA的丰度具有一致性。但不同给药途径在组织分布丰度上有明显区别,由强至弱分别为:局部注射>腹腔注射>静脉注射,在组织分布特征上具有一致性。(2)小鼠腹腔注射后,ADV及tk的各组织分布动态变化上具有一致性,腹腔注射后1 d ADV及tk即具有高表达,7 d表达最强,14 d下降60%左右,21 d残留20%左右,32 d基本消失。(3)随着ADV-GFP给药剂量的增强,ADV的组织分布表现为剂量依赖性逐步增强。剂量为病毒颗粒数每1 kg为6.7×108时,ADV主要分布于肝脏、脾脏、淋巴结和肠系膜,分布较低。当剂量达到病毒颗粒数每1 kg为6×109时,ADV在10个器官的分布以肝脏、脾脏、淋巴结、肠系膜和肾上腺分布增加最为明显。当剂量增加到病毒颗粒数每1 kg为1×1012时,ADV在10个器官的分布继续增加;在病毒颗粒数每1 kg为1×1013的大剂量病毒剂量情况下,肝脏的转染效率增加不太显著,但在其他阳性器官的分布增加。结论:本实验提示,采用每周用药1次,病毒颗粒数每1 kg为1×1012,局部或局部血管用药的给药方案可以得到肝脏有效转染强度,有望�Objective： To explore the histological distribution of adenovirus - mediated herpes simplex virus thymi- dine kinase （ ADV - tk） gene after topical or intravenous administration in Kunming mice. Methods： Histological distribution characteristics of ADV - GFP and ADV - tk in Kunming mice were studied. The biodistribution of ADV and tk in liver, spleen, lymph nodes, mesenterie, adrenal, thymus, kidney, thyroid, prostate and ovary etc. was detec-ted by GFP and thymidine kinase mRNA （fiber coding region） expression. Quantitation of immunoreactivity was ac- complished using an immune - histochemical scoring system （IHS） which was an image analysis - based system. Results： （1） The tissue distribution characteristics of three different administrations were similar. The results showed that the order of positive activity of ADV and tk was liver 〉 spleen 〉 lymphnodes 〉 mesentery 〉 adrenal 〉 thymus 〉 kidney 〉 thyroid 〉 prostate 〉 ovary. While, the immunohistochemical scoring analysis showed there was a significant difference of the abundance of ADV and tk in tissues among different injection ways , and the order was ： local injection 〉 intraperitoneal injection 〉 intravenous injection, while the histological distribution characteristics of ADV and tk were consistent. （2） The biodistribution characteristics of ADV were consistent with tk after intraperito- neal injection at different times. ADV and tk were highly expressed after 1 day, while the strongest expression ap- peared after 7 days,and then dropped to 60% after 14 days;there was only about 20% left after 21 days,which fi- nally disappeared after 32 days. （3） Tissue distribution of ADV was dose - dependent. At a dose of 6.7 ×10^12 virus particles per kg, ADV was mainly found in the liver, spleen, and mesenteric lymph nodes, while the abundance was low. At the dose of 6 ×10^9 virus particles per kg, there was significant accumulation in liver, spleen, lymph nodes, mesentery, and adrenal gland. When the dose was in

关 键 词：重组腺病毒 胸苷激酶 生物分布 绿色荧光蛋白 自杀基因 免疫组织化学评分法 

分 类 号：R917[医药卫生—药物分析学]