原位可注射壳聚糖基温敏水凝...管狭窄中分阶段释药应用研究_李宁.pdf

现代生物医学进展Progress in Modern Biomedicine Vol.23NO.1JAN.2023 基础研究 doi:10.13241/ki.pmb.2023.01.001原位可注射壳聚糖基温敏水凝胶的制备及其在预防 ESD 术后食管狭窄中分阶段释药应用研究*李宁1宁敏2宛新建2刘坚华3何伟娜3（1 上海交通大学基础医学院基础医学实验教学中心 上海 200025；2 上海交通大学附属上海市第六人民医院消化内科 上海 200233；3 上海交通大学基础医学院药物化学与生物信息学中心 上海 200025）摘要 目的：构建一种可以分阶段释放药物的原位可注射水凝胶，通过直接注射在 ESD（Endoscopic Submucosal Dissection，内镜黏膜下剥离术）术后伤口处，形成水凝胶敷料，起到保护伤口的作用。同时凝胶中的两种药物通过分阶段释放的方式，更好地促进伤口的无瘢痕愈合，为 ESD 术后食管狭窄的预防提供一种新的参考方案。方法：在壳聚糖/-甘油磷酸钠（CS/-GP）温敏水凝胶的体系中加入聚多巴胺（PDA），制备壳聚糖/-甘油磷酸钠/聚多巴胺（CS/-GP/PDA）水凝胶。通过在载药水凝胶中加入聚乙二醇-聚乳酸-羟基乙酸（PEG-PLGA）纳米载药微粒制备 CS/-GP/PDA/NPs 双载药水凝胶，通过两种载药体系的复合，实现药物的分阶段释放。通过流变学实验测定 CS/-GP、CS/-GP/PDA、CS/-GP/PDA/NPs 凝胶体系的相转变温度以及凝胶强度。通过高效液相色谱法检测 CS/-GP/PDA/NPs 水凝胶中两种药物的释放动力学特征。通过 CCK-8 细胞增殖实验评价 CS/-GP/PDA、CS/-GP/PDA/NPs 温敏水凝胶的生物相容性。在体外猪食管中，模拟 ESD 术后创口，通过内镜辅助将水凝胶母液注射在伤口处，并通过内镜观察水凝胶的凝胶状态。结果：得到了粘附性显著增强的壳聚糖/-甘油磷酸钠/聚多巴胺（CS/-GP/PDA）凝胶体系。流变学实验证明聚多巴胺（PDA）的加入可以显著降低水凝胶的凝胶温度，缩短原位成胶时间。CCK-8 实验显示 CS/-GP/PDA、CS/-GP/PDA/NPs 凝胶体系无潜在的细胞毒性。在体外猪食管模拟实验中，将其凝胶母液注射在伤口处后，可原位形成凝胶，且凝胶贴合伤口，具有较强的粘附性。通过体外释药速率测定，验证了 CS/-GP/PDA/NPs 水凝胶中所载两种药物释放速率存在明显差异，可实现药物的分阶段释放。结论：设计的 CS/-GP/PDA/NPs 凝胶体系适用于 ESD 术后的伤口修复，并能够实现分阶段释药，对于预防 ESD 术后食管狭窄具有潜在的应用价值。关键词：内镜黏膜下剥离术；食管狭窄；壳聚糖水凝胶；原位注射；温敏凝胶；分段释药中图分类号：R735.1；R-33文献标识码：A文章编号：1673-6273（2023）01-01-08In Situ Forming Injectable Hydrogel for Phased Dosing in PreventingPost-ESD Esophageal Stricture*LI Ning1,NING Min2,WAN Xin-jian2,LIU Jian-hua3,HE Wei-na3(1BasicMedicalExperimentTeachingCenter,ShanghaiJiaoTongUniversityCollegeofBasicMedicalSciences,Shanghai,200025,China;2 Department of Gastroenterology,Shanghai Jiao Tong University Affiliated Sixth Peoples Hospital,Shanghai,200233,China;3MedicalChemistryandBioinformaticsCenter,ShanghaiJiaoTongUniversityCollegeofBasicMedicalSciences,Shanghai,200025,China)ABSTRACT Objective:To construct an insitu injectable Chitosan-based hydrogel for phased dosing,of which the sol solution is in-jected directly into the wound after ESD(Endoscopic Submucosal Dissection)surgery to form a hydrogel dressing in situ.After introduc-ing drug-loaded nanoparticles into the hydrogel dressing,the dual drug-loaded hydrogel can promote the scar-free healing of woundsthrough different release kinetics of the drugs contained in the gel and nanoparticles,which will be a good solution for preventingpost-ESD esophageal stricture.Methods:Polydopamine(PDA)is added to chitosan/sodium glycerophosphate(CS/-GP)thermosensitivehydrogeltopreparechitosan/sodiumglycerophosphate/polydopamine(CS/-GP/PDA)hydrogel.TheCS/-GP/PDA/NPsdualdrug-loadedhydrogel was prepared by adding drug-loaded polyethylene glycol-poly lactic acid-co-glycolic acid(PEG-PLGA)nanoparticles to thedrug-loaded hydrogel,and the staged release of the drugs was achieved through the combination of the two drug carrier systems.Thephase transition temperature and rheological properties of CS/-GP,CS/-GP/PDA,CS/-GP/PDA/NPs gels were determined by rheo-*基金项目：国家自然科学基金项目（21504104）；上海市卫健委 优青计划 （2017YQ080）；上海市教委 晨光项目 （16GC14）作者简介：李宁（1990-），男，硕士研究生，实验员，主要研究方向：设计构建水凝胶载药系统，E-mail:通讯作者：宛新建，男，博士生导师，主任医师，E-mail:；刘坚华，男，硕士生导师，副研究员，E-mail:；何伟娜，女，硕士生导师，副研究员，E-mail:（收稿日期：2022-05-23 接受日期：2022-06-18）1现代生物医学进展Progress in Modern Biomedicine Vol.23NO.1JAN.2023logical tests.The release kinetics of the two drugs in CS/-GP/PDA/NPs hydrogels was detected by high performance liquid chromatog-raphy(HPLC).The biocompatibility of CS/-GP/PDA,CS/-GP/PDA/NPs gels was evaluated by CCK-8 cell proliferation experiments.In the porcine esophagus in vitro,the wound that would generate after ESD was simulated.The sol solution was injected into the woundwith the aid of endoscopy,and the state of the hydrogel was observed through an endoscope.Results:A chitosan/-sodium glycerophos-phate/polydopamine(CS/-GP/PDA)gel with significantly enhanced adhesion was prepared.Rheological experiments proved that theaddition of PDA could significantly reduce the gel temperature of the hydrogel and shorten the in situ gelling time.CCK-8 experimentsshowed that CS/-GP/PDA and CS/-GP/PDA/NPs gel systems both exhibit minimal cytotoxicity.In the porcine esophagus simulationexperiment in vitro,after the sol was injected into the wound,the gel could be formed in situ,and the gel adhered to the wound strongly.Through measuring the drug release rate,it was verified that the release rate of the two drugs contained in the CS/-GP/PDA/NPs hydro-gel was significantly different,suggesting that the phased dosing could be achieved.Conclusion:The designed CS/-GP/PDA/NPs gelsystem is suitable for post-ESD wound repair,and could achieve phased dosing,revealing its potential for preventing post-ESDesophageal stricture.Key words:ESD;Esophageal stricture;Chitosan hydrogel;in situ injection;Thermosensitive gel;Phased dosingChinese Library Classification(CLC):R735.1;R-33Document code:AArticle ID:1673-6273(2023)01-01-08前言食管癌是一类发生在消化道的癌症，具有较高的发病率及死亡率，据世界卫生组织 2020 年的统计数据显示，全球食管癌一年的新发病例为 60.41 万，其中我国就有 32.44 万新发病例，占全球发病总数的 53.7%，而全球 54.4 万例的死亡病例也使得对食管癌的治疗至关重要1。在食管癌早期，癌细胞仅存在于食管黏膜层和黏膜下层，尚未发生淋巴结转移，可直接通过手术进行切除治疗2。内镜黏膜下剥离术（End