目的:检测可卡因-苯丙胺调节转录肽(CART)蛋白疫苗在吗啡镇痛及其耐受中的作用。方法:利用基因克隆技术构建pGEX-4T3-CART55-102表达质粒,通过谷胱甘肽S转移酶(GST)亲和层析法纯化出GST-CART蛋白。实验分为6组:空白对照组,生理盐水组,GST+弗氏佐剂组,CART蛋白疫苗5μg组、10μg组和20μg组,免疫两次后热板法对各组进行痛反应检测。皮下注射6mg/kg吗啡进行镇痛效应检测,计算最大镇痛效应百分率(MPE%)。随后建立吗啡耐受模型,并于末次给药12h后皮下注射6mg/kg吗啡进行耐受效应检测。结果:CART蛋白疫苗本身对基础痛阈没有影响(P〉0.05)。10μg CART蛋白疫苗组显著降低了吗啡镇痛效应(P〈0.05)。对比生理盐水组,疫苗组显示出抗耐受的潜力,其中10μg组MPE%增高具有显著性差异(P〈0.05)。结论:CART蛋白疫苗本身对痛反应没有影响,但削弱了吗啡的镇痛效应,同时对吗啡镇痛耐受具有拮抗作用。
Objective: To study the effect of cocaine-and amphetamine-regulated transcript (CART) protein vaccine on morphine analgesia and tolerance. Methods: The expression plasmid pGEX-4T3-CART was constructed by gene cloning. The CART protein was purified by glutathione s-transferase (GST)-affinity chromatography. The experiment included 6 groups: blank control,normal saline (NS),GST+Freund's adjuvant,and CART protein vaccine (5 μg, 10 μg and 20 μg) groups. After immunization for twice,all groups were tested in hot plate. Morphine analgesia effect was evaluated through s.c. injection with 6 mg/kg morphine solution,calculated by MPE%. Then morphine tolerance model was established,and the tolerance to morphine was tested by s. c. injection with 6 mg/kg morphine 12 h after the last injection. Results: CART vaccine itself had no pronounced effect on the pain threshold (P〉0.05). CART vaccine at 10 μg significantly depressed the analgesic effect of morphine analgesia (P〉0.05). Compared with NS group, vaccine groups showed a potential antagonizing tolerance effect, especially in the 10 μg group,with the MPE% significantly increased (P〈0.05). Conclusion: CART vaccine itself has no influence on the pain response; however,it can impair the analgesia effect of morphine and can antagonize the analgesia tolerance to morphine.