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Prevention of reperfusion liver injury by SOD-(DIVEMA) conjugate

SOD-(DIVEMA) 结合物预防再灌注肝损伤

基本信息

批准号：
06455003
负责人：
TODOROKI Takeshi
金额：
$ 4.42万
依托单位：
University of Tsukuba
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1994
资助国家：
日本
起止时间：
1994 至 1995
项目状态：
已结题

项目摘要

Procedures for prolongation of superoxide dismutase (SOD) activity both in vitro and in vivo were established by synthesizing the conjugates with the copolymer of the divinyl ether and maleic anhydride (DIVEMA), which have average molecular weigh of 30,000 and 5,000 and narrow molecular weigh distribution (Wm/Mn=1.4). The enzymatic activity of SOD decreased to 50% within 139 min.in human plasma, on the contrary SOD-DIVEMA showed no decrease in 24 hr. SOD injected into mice lost 50% of its activity within 6 min., whereas a half life of SOD in conjugate was 69 min. The conjugating of DIVEMA markedly diminished antigenicity of SOD and increased the stability for temperature. While, an intravital fluorescence epi-illuminated television microscopic observation and analyzing system was newly developed by this Grant in order to estimate the efficacy of conjugating with DIVEMA on the prevention of reperfusion injury of the liver microcirculation. Sodium fluorescein and Rodamin-6G were used for staining serum and leukocyte. The liver microcirculation was recorded under our newly developed epi-illumination video microscope for 30min.after decramping the portal vein. Cramping of the portal vein was continued for 30 min.and a tested drug was administered one min.prior to decramping. The perfusion rate (No.of perfusing sinusoid per acine/Total number of sinusoid), the number of sticking and rolling leukocytes in the lobulus, and the rate of sticking and rolling leukocytes (No.of sticking and rolling leukocytes/total No.of passsing leukocyte) in the post sinusoidal central venule were used as indicators for the reperfusion injury of the liver. Administration of SOD-DIVEMA conjugate prevented reperfusion injury significantly (p<0.05) in compare to SOD administration. On the other hand SOD showed insignificantly better microcirculation estimated in every applied indicators than non treated control.

通过合成平均分子量分别为30,000和5,000、分子量分布较窄(Wm/Mn=1.4)的二乙烯基醚与马来酸酐的共聚物(DIVEMA)缀合物，建立了延长体内外超氧化物歧化酶(SOD)活性的方法。人血浆中SOD的酶活性在139分钟内下降至50%，相反SOD-DIVEMA在24小时内没有显示下降。注射到小鼠体内的 SOD 在 6 分钟内失去了 50% 的活性，而结合物中 SOD 的半衰期为 69 分钟。 DIVEMA 的结合显着降低了 SOD 的抗原性并增加了温度稳定性。同时，本次Grant还新开发了活体荧光落射电视显微观察分析系统，以评估与DIVEMA结合预防肝脏微循环再灌注损伤的功效。使用荧光素钠和Rodamin-6G对血清和白细胞进行染色。解除门静脉痉挛后，在我们新开发的落射照明视频显微镜下记录肝脏微循环30分钟。门静脉痉挛持续30分钟，并在解除痉挛前一分钟施用测试药物。以灌注率（每根腺泡灌注血窦数/血窦总数）、小叶内粘滚白细胞数、肝窦后中央微静脉粘滚白细胞率（粘滚白细胞数/通过白细胞总数）作为肝脏再灌注损伤的指标。与SOD施用相比，SOD-DIVEMA缀合物的施用显着预防再灌注损伤（p<0.05）。另一方面，SOD 在每个应用指标中显示出比未处理对照更好的微循环。