太子参抗心肌缺血的药效物质基础及作用机制研究

Ischemic heart disease (IHD) is a type of serious cardiovascular disorders and remains the leading cause of death and heart failure worldwide. Due to complex pathological reasons, there are no effective therapies for its prevention and treatment. Although surgical treatment has been widely used in clinical therapy, its high recurrence rate and need of additional surgical treatments precluded its wide acceptance. Chemotherapy is still the main strategy for the prevention and treatment of IHD. The effectiveness and use of Pseudostellaria heterophylla in the treatment of IHD can be found in folkloric medicines, but its effective compounds (except polygaccharides) and mechanism of action are still unclear. Previous investigations showed that fractions enriched in polysaccarides (PHP) and glycosides (PHS) were the two active fractions against cardiomyocyte hypoxia, with potency comparable to the positive control NAC. Target cell extraction and UPLC-MS/MS analysis has led to the hit of 14 potential active compounds, while isolation and pharmacological studies showed that the two compounds were anti-hypoxic compounds. In the current project, various pharmacological and chemical isolation techniques will be employed to evaluate and isolate the chemical constituents present in the active fractions for effective anti-myocardial ischemia agents and to know the effective anti-myocardial ischemic compounds of P. heterophylla and its mechanism of action.

缺血性心脏病是一类严重的心血管系统疾病，是导致死亡和心功能衰竭的主要原因。由于发病机理复杂，这类疾病仍无非常有效的防治手段。尽管手术治疗常在临床上被采用，但由于术后复发严重，药物治疗仍是这类疾病的主要防治方式。太子参在民间曾被用来治疗心悸，研究也证实其具有抗心肌缺血作用，但除多糖外的其它作用物质及作用机制尚不清楚。前期研究发现，太子参富含苷类组分（PHS）和富含多糖组分是其抗心肌缺氧的有效组分，其作用与阳性药N-乙酰-L-半胱氨酸相当。靶细胞萃取结合液质联用分析对PHS的筛选得到了14个潜在活性单体。针对其中两个单体的分离和药效评价发现它们均为抗心肌缺氧有效成分。本课题拟在已有基础上，通过有效组分的体内药效、靶细胞萃取和液质联用分析、药效物质的分离纯化和作用机制研究，明确太子参抗心肌缺血的物质基础和作用机制，为太子参的开发利用和抗心肌缺血活性单体的开发奠定基础。

太子参在民间常被用来治疗心悸，研究也证实其具有抗心肌缺血作用，但抗心肌缺血的药效物质基础尚不清楚，本课题通过体外CoCl2诱导H9c2心肌细胞缺氧模型对太子参进行有效组分筛选，结果发现太子参富含多糖组分和富含苷类组分具有降低胞内SOD、LDH和ROS，上调MDA的作用。表明太子参多糖组分和苷类物质为其有效组分。体外Na2S2O4诱导的H/R损伤H9c2心肌细胞的保护作用实验也证实富含苷类组分作用最强。机制研究表明，太子参多糖组分对大鼠心肌I/R损伤的改善作用可能与其具有降低氧化应激反应，改善心肌细胞超微结构，减少心肌细胞凋亡，下调Bax、Bcl-2和Caspase-3凋亡基因的表达量有关。苷类组分对心肌细胞的保护作用可能与抑制心肌细胞脂质过氧化相关。.采用靶细胞萃取技术、UPLC-DAD-MS分析以及现代分离纯化技术从富含苷类有效成分中分离出苷类物质9个，其它类型化合物12个。包括7个新化合物，分别为：Pseudoheterin A–E, 3-呋喃甲醇-D-吡喃半乳糖苷和9-(furan-3- yl methyl) -9H -purin-6-amine。其中水杨酸、咖啡因、胸苷、二氢阿魏酸甲酯、苯甲酸、hemerocallisamine VI、异麦芽酚葡萄糖苷、N-(4-Ethoxyphenyl) acetamide和白藜芦醇苷为太子参中首次分离的化合物。单体化合物经药效评估筛选发现Pseudoheterins A，B, E和白藜芦醇苷低、中、高剂量组的细胞存活率明显升高（P<0.05， <0.01，或＜0.001）表明其具有心肌保护作用，是太子参的有效成分，基本明确了太子参的药效物质基础。.为进一步明确卡波林类单体化合物（Pseudoheterin类）对心肌保护作用的机制，课题组对Pseudoheterin E合成得到前体化合物KBL-X。实验发现KBL-X能显著抑制LDH（P＜0.01）释放量与MDA（P＜0.05）含量，并且能显著提高GSH、SOD活性（P＜0.05）。机制研究显示KBL-X可能通过PKC依赖性途径对H9c2心肌细胞发挥抗细胞凋亡作用。.本课题研究基本明确太子参抗心肌缺血的物质基础，基本阐明了太子参有效组分的抗心肌缺血作用机制，为太子参的开发应用和抗心肌缺血活性单体药物的研发奠定了实验基础。.