贯叶金丝桃素通过星形胶质细胞介导的免疫抑制微环境促进缺血性脑卒中神经血管再生和功能恢复的研究

Our recent study and preliminary data found that hyperforin could activate astrocytes which could promote neurogenesis and long-term functional recovery through the secretion of interleukin (IL-6) and IL-17A in the delayed phases of stroke recovery, but the underlying mechanisms are unclear. Immune suppression after brain injury is critical for neurovascular regeneration; and T helper-2 type (Th2) cells and regulatory T cells (Tregs) play important roles in the immune suppression after brain injury. In neurodegenerative diseases, astrocytes can be induced to become non-professional antigen presenting cells (APCs), which mainly stimulate CD4+ T cells to differentiate into Th2 cells. In addition, IL-6 can also promote Th2 differentiation from CD4+ T cells. Our preliminary results showed for the first time that astrocytes could also be induced to express the major histocompatibility complex class Ⅱ (MHC-Ⅱ) and intercellular cell adhesion molecule-1 (ICAM-1) on its membrane to become non-professional APC during stroke recovery. We also found significant dysfunction of Tregs (IL-17A-secreting Tregs) in the ischemic brain during stroke recovery. According to these findings, we hypothesize that hyperforin can activate astrocyte which could act as non-professional APC and secret IL-6 to promote Th2 differentiation from CD4+ T cells. In addition, hyperforin stimulates astrocytes to secret IL-17A during stroke recovery to inhibit Tregs dysfunction in a negative feedback manner, and ultimately exert immune suppression and promote neurovascular regeneration and functional recovery. In summary, the findings will provide a new strategy for developing drugs to promote neurorepair in the later phases of stroke recovery.

我们前期发现，贯叶金丝桃素可促进缺血性脑卒中（CIS）慢性恢复期星形胶质细胞分泌IL-6和IL-17A，进而显著促进神经再生和功能恢复，但具体机制不明。脑损伤后免疫抑制微环境对神经血管再生至关重要，其中Th2和Tregs对该微环境起关键作用。星形胶质细胞在神经退行性疾病可被诱导为非专职抗原提呈细胞（APCs），刺激CD4+ T分化为Th2；IL-6也可促进CD4+ T的Th2分化。我们首次发现，CIS慢性恢复期星形胶质细胞也可被诱导表达MHC-Ⅱ和ICAM-1成为非专职APCs，同时存在Tregs功能障碍（分泌IL-17A）。提出研究假设：贯叶金丝桃素激活星形胶质细胞，通过分泌IL-6和其抗原提呈功能促进CD4+ T细胞的Th2分化，以及通过分泌IL-17A负反馈调节Tregs功能，共同营造免疫抑制微环境，促进神经血管再生和功能恢复。本课题可为CIS的功能恢复治疗提供新策略和新药物。

缺血性脑卒中急性期的治疗窗口短，恢复期治疗时间窗很宽，通过促进恢复期神经血管再生可促进长期神经功能的恢复。我们建立C57BL/6、Tlr4Lps-d和IL-17a-/-小鼠大脑中动脉栓塞（MCAO）模型模拟缺血性脑卒中，并通过体外原代培养脑室下区神经祖细胞增殖和分化实验，以及体外原代培养缺血侧大脑星形胶质细胞。我们通过体内外研究发现:. 1.缺血性脑卒中恢复期持续给予贯叶金丝桃素（hyperforin）可通过增加星形胶质细胞合成和分泌IL-17A，IL-17A促进缺血侧脑组织VEGF表达，进而促进血管再生和长期神经功能恢复；. 2.hyperforin通过抑制脑卒中后抑郁和焦虑，促进缺血侧脑组织TGF-β表达，进而促进海马神经再生和改善认知功能障碍；. 3.hyperforin通过TLR-4促进恢复期星形胶质细胞合成和分泌IL-6，进而增加Tregs向缺血侧脑组织浸润，增加Tregs来源细胞因子IL-10和TGF-β表达，以及促进缺血侧脑组织Th2细胞分化，最终促进缺血侧脑组织免疫抑制微环境形成，促进神经血管再生和神经功能恢复。. 4.缺血侧脑组织免疫抑制微环境形成还可通过社会支持介导的心脏合成和分泌ANP，ANP作用于室管膜和脉络丛上皮细胞NPR-A，促进其分泌TGF-β，增加缺血侧脑组织Tregs。. 我们的研究表明hyperforin通过星形胶质细胞诱导的缺血侧脑组织免疫抑制微环境形成对神经血管再生和功能恢复至关重要，可为缺血性脑卒中恢复期治疗提供新策略。