基于CD4+CD25+Foxp3+Tregs免疫调控网络探讨“从心论治”易损斑块作用机制

The rupture and hemorrhage of vulnerable plaque is the leading cause of acute cardio-cerebro-vascular events. Thereare still some difficulties in clinical treatment by Western Medicine.It has been proved that the web of CD4+CD25+Foxp3+Tregs has participated in the development of atherosclerosis and vulnerable plaque, however its specific mechanism has not been elucidated. Our previous studies found that Dingxin Recipe could treat coronary heart disease and prevent sudden death.The experiment also found that DXR can reduce AS damage and stabilizevulnerable plaque.In Traditional Chinese Medicine, it is said that “the heart could maintains blood and vessel”. It is a good interpretation of the relationship between the heart and blood vessels.We suggest that vulnerable plaque should be treated from heart. So, we have optimized DingxinRecipe, presumably the principal effect of drugs are into the Heart Sutra. Our pre-experiment found that the combination with berberine, salvia, Semen and Ganodermacould reduce AS damage and stabilize vulnerable plaque in mice, and it also could regulate Tregs network.So, we suppose that treatment from heart could stabilize atherosclerotic plaque through modulating the web regulated by CD4+CD25+Foxp3+ Tregs. The aim of this study is to explore the molecular mechanism of treatment from heart and CD4+CD25+Foxp3+Tregsweb in vulnerable plaque in rabbit model and reveal its target point in cell level.Furthermore, we aim to deepen the understanding of "the heart could maintain blood and vessel" in Traditional Chinese Medicine.

易损斑块破裂出血是急性心脑血管事件的主因，临床治疗仍比较棘手。目前认为CD4+CD25+Foxp3+Tregs免疫网络参与了易损斑块发生全过程，但其具体机制不清。依据中医“心主血脉”理论，我们对易损斑块疗效确切的定心方进行优化，认为该方入心经药物（黄连、丹参、酸枣仁和灵芝）可能发挥了主要功效。初步研究发现上述药物配伍能显著减轻小鼠动脉粥样硬化损伤，稳定斑块，并能调节Tregs网络。我们推测从心论治可通过调控Tregs免疫网络稳定斑块。本项目拟采用体内结合体外的方法，以从心论治为干预手段，利用易损斑块家兔模型，深入研究Tregs网络调控易损斑块机制及从心论治的干预效应；在细胞水平采用家兔主动脉平滑肌细胞与Tregs共培养模式，从信号传导方面探讨从心论治对Tregs的调节机制，以阐明Tregs在易损斑块中的发生机理，揭示从心论治干预易损斑块作用机制及分子靶点，深化对“心主血脉”中医理论认识。

易损斑块破裂出血是急性心脑血管事件的主因，临床治疗手段虽在不断发展中，但治疗现状与效果仍然不容乐观，仍是心血管临床较为棘手的难题。本项目依据中医“心主血脉”基本理论，提出“从心论治动脉粥样硬化性疾病”的科学假说，从CD4+CD25+Foxp3+Tregs免疫网络着手，初步探讨了中药从心论治方（CXLZF）防治易损斑块的药效和分子机制研究。采用高脂喂养ApoE基因敲除（ApoE-/-）小鼠建立易损斑块模型，分别采用Tregs和CXLZF干预小鼠。结果发现，Tregs和CXLZF均能有效调节小鼠血脂水平，减轻血清及斑块处炎症反应，调控斑块处基质金属蛋白酶（MMPs）表达，缩小AS斑块面积，从而减轻AS进展。立体水平发现，Tregs能显著调节MASMCs与HUVEC细胞的抗炎能力，CXLZF可能主要通过Tregs免疫网络抑制ox-LDL诱导HUVEC细胞的炎症损伤，并且MAPK信号家族可能参与这一过程。为进一步挖掘CXLZF的主要药效成分，课题组探讨了其主要成分小檗碱抗AS损伤的药理作用，发现小檗碱在调节血脂水平、降低炎症因子表达、减轻AS斑块损伤等方面与原方类似，并且能显著调节AS小鼠及巨噬细胞中PBEF的表达，该蛋白可能是小檗碱抗AS易损斑块损伤的重要分子靶点。