杜华康,陆苑,金阳,陈玉颖,王永林,李勇军,刘文
DU Huakang,LU Yuan,JIN Yang,CHEN Yuying,WANG Yonglin,LI Yongjun,LIU Wen

• 1:贵州医科大学贵州省药物制剂重点实验室&省部共建药用植物功效与利用国家重点实验室
• 2:贵州医科大学药学院
• 3:贵州医科大学民族药与中药开发应用教育部工程研究中心

摘要(Abstract)：

目的 探讨载阿霉素(DOX)纳米粒温敏凝胶(DOX-NPs-Gel)复合体系的体内缓释性能、抗肿瘤作用及瘤内滞留性能。方法 18只SD雄性大鼠随机均分为DOX组、DOX-碘油组(4 g/L DOX溶液1 mL与碘油1 mL混匀现配，5 mL/kg)及DOX-NPs-Gel组(5 mg/kg DOX+2 g/L DOX),腹腔注射，分别于给药后不同时间点经眼底静脉丛穿刺取血，采用超高效液相色谱-串联质谱(UPLC-MS/MS)技术检测血浆中DOX的浓度，利用WinNonLin 8.1软件拟合主要药代参数[半衰期(t_(1/2))、峰浓度(C_(max))、曲线下面积(AUC)、清除率(CL)、平均驻留时间(MRT)];培养、收集小鼠肝癌细胞株H22细胞制成悬液，单次接种于90只雄性ICR小鼠右腋皮下构建荷瘤模型，分2部分进行实验，第一部分取荷瘤小鼠36只均分为生理盐水组(注射用生理盐水，5 mL/kg)、空白NPs-Gel组(100 mg空白NPs冻干粉分散于1 mL空白Gel, 5 mL/kg)、碘油组(碘油，5 mL/kg)、DOX组(2 g/L DOX,5 mL/kg)、DOX-碘油组(4 g/L DOX溶液1 mL与碘油1 mL混匀现配，5 mL/kg)及DOX-NPs-Gel组(2 g/L DOX,5 mL/kg),瘤内注射、给药1次，观测10 d,给药后每2天称体质量1次并计算各组小鼠的体质量变化百分比，称重后同时测量各组小鼠瘤体宽度、长度并计算肿瘤体积(V),观测结束时脱颈处死、剥离肿瘤组织，称取各组小鼠瘤体质量并计算抑瘤率，然后制作切片采用苏木精-伊红(HE)染色观察组织学特征；第二部分取荷瘤小鼠54只均分为DOX组、DOX-碘油组及DOX-NPs-Gel组，给药剂量和途径同前，给药后12、24、48、72、96及120 h时取各组3只小鼠脱颈处死，剥离肿瘤组织，采用UPLC-MS/MS检测瘤体组织中各时间点的DOX瘤内滞留率。结果 体内缓释性能研究结果显示，相比于其余组，DOX-NPs-Gel组大鼠t_(1/2)延长、CL降低(P<0.05);抗肿瘤作用考察结果显示，与其它治疗组相比，DOX-NPs-Gel组小鼠肿瘤组织生长最慢，瘤重降低(P<0.05),抑瘤率最高(76.55%),HE染色显示肿瘤组织出现大范围凋亡和坏死情况；瘤内滞留性能考察结果显示，DOX-NPs-Gel组小鼠第5天时滞留率为(44.14±3.84)%,其余组第4天时滞留率已降至0%。结论 与DOX和DOX-碘油相比，DOX-NPs-Gel在动物体内具有更强的缓释性能、抗肿瘤作用及瘤内滞留性能。
Objective To investigate the extended release performance in vivo, anti-tumor effect and intratumor retention performance of doxorubicin-loaded nanoparticles thermosensitive gel(DOX-NPs-Gel) composite system. Methods Eighteen SD male rats were randomly divided into doxorubicin(DOX) group, DOX-iodine oil group(1 mL 4 g/L DOX solution mixed with 1 mL iodine oil on site, 5 mL/kg) and DOX-NPs-Gel group(5 mg/kg DOX+2 g/L DOX); then subjects received intraperitoneal injection; blood was taken through the fundus venous plexus at different time points after administration of drugs; the concentration of DOX in plasma was detected by ultra-performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS), and the main pharmacokinetic parameters were fitted by WinNonLin 8.1 software [half life period(t_(1/2)), C_(max), area under the curve(AUC), clearance(CL), mean residence time(MRT)]. Rats hepatocarcinoma strain H22 was cultivated and collected for suspension; single vaccination was performed in right axilla subcutaneous part of 90 male ICR rats for load tumor. Such experiment was divided into two parts: part one, 36 rats with load tumor were evenly divided into normal saline group(injected with normal saline, 5 mL/kg), blank NPs-Gel group(100 mg blank NPs lyophilized powder distributed in 1 mL blank Gel, 5 mL/kg), iodine oil group(iodine oil, 5 mL/kg), DOX group(2 g/L DOX,5 mL/kg), DOX-iodine oil group(1 mL 4 g/L DOX mixed with 1 mL iodine oil on site, 5 mL/kg) and DOX-NPs-Gel group(2 g/L DOX,5 mL/kg); intratumor injected for one time and observed for 10 d, body mass was measured every two days after drug administration and calculated changes of body mass ration of all groups; the size and length of tumor was measured after weighing and tumor volume(V) was calculated; all the mice were sacrificed by neck removal after observation finished, tumor tissue was stripped, tumor mass was weighed and tumor inhibition rate was calculated; histological features of tumor tissue was observed by slicing and HE staining. Part two, 54 load tumor mice were evenly divided into DOX group, DOX-iodine oil group, and DOX-NPs-Gel group; all subjects followed the same pattern of dosage and experiment method. At 12, 24, 48, 72, 96, and 120 h after drug administration, three mice from each group were sacrificed by neck removal, tumor tissue was stripped, UPLC-MS/MS was adopted to detect DOX intratumor retention rate of tumor tissue at different time points. Results Compared with other groups, in vivo extended-release performance study showed that the half-life(t_(1/2)) of DOX-NPs-GEL group was prolonged, and CL rate decreased(P<0.05). Findings of anti-tumor effect study indicated that, the growth of tumor tissues in DOX-NPs-Gel group was the slowest, the tumor weight decreased(P<0.05), the tumor inhibitory rate was the highest(76.55%) compared with other treatment group. After HE staining, the tumor tissues showed large scale apoptosis and necrosis. The results of intratumor retention showed that the retention rate of DOX-NPs-Gel was(44.14±3.84) % on the 5~(th )day, and the retention rate of other groups dropped to 0% on the 4~(th )day. Conclusion Compared with DOX and DOX-iodide, DOX-NPs-Gel showed stronger extended-release properties, antitumor effects and intratumor retention in animals.

关键词(KeyWords)： 纳米粒;温敏凝胶;药代动力学;抗肿瘤作用;缓释作用;肝动脉化疗栓塞术
nanoparticles;thermosensitive gel;pharmacokinetics;antitumor effect;extended-release effect;transcatheter arterial chemoembolization

Abstract:

Keywords:

基金项目(Foundation): 中央引导地方科技专项项目(黔科中引地[2018]4006);; 贵州省科技计划项目(黔科合平台人才[2016]5613,5677)

作者(Author): 杜华康,陆苑,金阳,陈玉颖,王永林,李勇军,刘文
DU Huakang,LU Yuan,JIN Yang,CHEN Yuying,WANG Yonglin,LI Yongjun,LIU Wen

DOI: 10.19367/j.cnki.2096-8388.2024.03.006

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