在分子动力学模拟指导下自组装构建可同时递释氢气的中药复方组分双重响应内耳给药系统及防治噪声性聋的研究

In the treatment of Noise-induced hearing loss (NIHL), how to achieve selective distribution at the pathogenic site and targeted release of traditional Chinese medicine (TCM) in order to improve the efficacy and minimize the side effects, has become an urgent problem to be solved. There exists an increase of reactive oxygen species (ROS) accompanying low pH in the inner ear while NIHL attacks. Hydrogen can alleviate NIHL as a therapeutic agent with unique advantage. Therefore, on the basis of the pathological characteristics as well as our previous findings, we will perform an innovative study on self-assembled dual-responsive inner ear drug delivery system loaded with compound components of TCM formulae simultaneously delivering hydrogen through the intermolecular weak interaction under the guidance of molecular dynamics simulation. Calcium carbonate and chitosan coordinated with iron ions will be applied to fabricate pH-responsive core and ROS-responsive shell of the nanoparticles (NPs), respectively. Acting as TCM partner, salviae miltiorrhizae and panax notoginseng are going to be employed as model drugs and their hydrophobic and hydrophilic components including tanshinone IIA and panax notoginseng saponins will be loaded in the NPs. Nano-calcium hydride will be encapsulated into mPEG-PLGA so as to achieve slow and steady hydrogen. The composite nano-drug delivery system could not only exert TCM-hydrogen synergistic therapeutic effects, but also automatically release compound components at the pathogenic site of the inner ear. We will variously examine intelligent response mechanism, hydrogen sustained-release behavior, in vivo/in vitro transporting and distributing law, biocompatibility and pharmacodynamics by means of in vitro and in vivo experiments with the cell, organism and animal level. The experimental results would demonstrate the potential of the formulated nanoparticulate drug delivery system served as TCM multicomponent delivery carrier via inner ear administration. In summary, this study will help to supply scientific information for developing design theory and methods of TCM compound prescription.

在噪声性聋的防治中，如何实现中药在病灶部位有针对性地释药，尽量提高疗效、减少毒副作用，已成为一个亟待解决的关键问题。本项目在前期研究基础上，基于噪声性聋病发时活性氧水平增加和伴随的低pH病理特点以及氢气的独特优势和作用，以分子动力学模拟为理论指导，利用分子间弱相互作用自组装构建可同时递释氢气的中药复方组分双重响应内耳给药系统。该系统分别以碳酸钙、配位铁离子的壳聚糖为pH响应内核和氧化响应外壳并同时包载药对丹参-三七脂溶性和水溶性组分，将纳米氢化钙包裹于mPEG-PLGA缓慢持续性地释放氢气，不仅能发挥中药与氢气的协同治疗作用，而且药物可在病灶部位实现自动调节释放。项目通过体外与体内试验，在细胞、组织和整体动物水平上多层次考察该给药系统的智能响应释药机理、缓释氢气特点、体内外转运分布机制、生物相容性及药效学规律，探讨其作为中药复方组分内耳给药载体的潜力，为中药制剂的设计理论和方法提供参考。

耳聋（听力损失）在国内外都是常见病，全球五分之一人口即15亿人存在不同程度的听力损失，其中超过4亿人患有残疾性听力损失。感音神经性聋包括噪声性聋、药源性聋、老年性聋等，其中噪声性聋是由外界噪声诱发，以耳蜗毛细胞损伤为主要机制的进行性感音神经性聋。血迷路屏障（血内耳屏障）严重阻碍了全身用药时中药药效成分进入内耳，极大限制了中药对耳聋的治疗效果。尽管临床应用可避开血迷路屏障的内耳局部给药越来越受到重视，但如何实现中药有效组分在耳蜗病灶部位的靶向输送及智能化释药，减少在体内其他部位的分布与作用，从而在修复听力损失的同时又能降低毒副作用，是亟待解决的一个重要难题。基于耳聋病发时活性氧水平增加和伴随的低pH病理特点，本项目引入现代医学理论和制剂新技术，以分子动力学模拟为理论指导，构建环境响应性的新型内耳给药系统，以实现体内定位递送与智能释药一体化。“质量源于设计”，本项目从剂型角度深入研究，构建了可负载氢气、具pH/氧化响应功能等一系列递药系统，并进行了结构表征与体外性质考察，通过耳蜗毛细胞模型、斑马鱼模型、在体圆窗膜模型和豚鼠体内实验全面研究递药系统的体内外转运行为。制得的递药系统具有双重响应性能，局部给药后能以完整形态穿过圆窗膜递送至耳蜗并被毛细胞摄取；透膜机制主要是以网格蛋白、小窝蛋白介导等多种内吞方式进入圆窗膜，然后胞吐排出到内耳外淋巴；而且其毒性低，对毛细胞、斑马鱼、内耳组织的生物相容性良好。在此基础上，进一步设计和制备了可递释氢气与丹参、三七有效组分的双重响应等纳米递药系统，考察了递药系统体外性质及经内耳局部给药的转运分布特征和疗效，结果显示纳米载体可有效携载中药多组分至耳蜗，提高了局部生物利用度，延长了体内滞留时间，并能有效防治听力损失。综上，本项目的研究结果不仅为内耳局部给药系统的设计提供理论指导，也为中药复方内耳局部制剂的研发提供技术支撑，具有较高的学术价值和应用前景。