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摘要：
目的 探讨电针（electroacupuncture, EA）对脑缺血再灌注损伤后血管再生的影响及肿瘤转移相关蛋白1（metastasis‑associated genes 1, MTA1）在其中的作用。 方法 80只8~10周龄C57小鼠采用随机数字表法分为5组（每组16只）：假手术组（Sham组）、脑缺血再灌注损伤组［大脑中动脉栓塞（middle cerebral artery occlusion, MCAO）组］、EA治疗组（EA组）、对照干扰小RNA（small interfering RNA, siRNA）组（Control siRNA组）、MTA1 siRNA组。Sham组不做任何处理；MCAO组行MCAO，在建模后第3天仅接受麻醉及针刺，不给予EA刺激，连续5 d；EA组MCAO建模后第3天，在“百会”穴进行EA治疗，连续5 d；Control siRNA组MCAO建模后在缺血半暗带区注射对照siRNA，后续处理同EA组；MTA1 siRNA组MCAO建模后在半暗带区注射MTA1 siRNA，后续处理同EA组。采用神经功能学评分、2，3，5‑氯化三苯基四氮唑（2, 3, 5‑triphenyltetrazolium chloride, TTC）和TUNEL染色法分别测定神经功能、脑梗死面积和神经元凋亡数量，同时采用Western blot法检测MTA1、裂解的胱天蛋白酶‑3（cleaved caspase‑3）和血管性血友病因子（von willebrand factor, vWF）蛋白水平；RT‑PCR法测定MTA1 mRNA水平；免疫荧光法标记血管内皮并计算血管密度；比较各组小鼠MCAO术前、术中及术后PaO2、PaCO2、pH、脑血流量及肛温变化情况。 结果 与MCAO组比较，EA组和Control siRNA组神经功能学评分、vWF蛋白水平及血管密度增加（P<0.05），而脑梗死面积、cleaved caspase‑3蛋白水平及神经元凋亡数量减少（P<0.05）；EA组MTA1蛋白水平及MTA1 mRNA水平增加（P<0.05）。与Control siRNA组比较，MTA1 siRNA组神经功能学评分、vWF蛋白水平及血管密度降低（P<0.05），脑梗死面积、cleaved caspase‑3蛋白水平及神经元凋亡数量增加（P<0.05）。与Sham组比较，其余4组小鼠术中脑血流量明显减少（P<0.05），MCAO组和EA组MTA1蛋白水平及MTA1 mRNA水平增加（P<0.05）。其余各组指标差异均无统计学意义（P>0.05）。 结论 EA通过上调MTA1促进血管生成改善脑缺血再灌注损伤后神经功能。
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Abstract: Objective To investigate the effect of electroacupuncture (EA) on angiogenesis after cerebral ischemia and the role of metastasis associated protein 1 (MTA1) during this process. Methods According to the random number table methods, 60 C57 mice aging 8‒10 weeks were randomly divided into five group (n=12): a Sham group, a middle cerebral artery occlusion (MCAO) group, an EA treatment (EA) group, a control small interfering RNA (siRNA) group (Control siRNA group) and a MTA1 siRNA group. The Sham group did not receive any processing. The MCAO group underwent MCAO, and were subjected to acupuncture and anesthesia on Day 3 after modeling, without EA for consecutive five days. The EA group were subjected to EA at the acupoint Baihui on Day 3 after MCAO modeling for consecutive five days. The Control siRNA group was injected with control siRNA in the penumbra immediately after MCAO modeling, followed by the same treatment with those in the EA group. The MTA1siRNA group was injected with MTA1 siRNA in the penumbra immediately after MCAO modeling, followed by the same treatment with those in the EA group. Then, their neurological function, cerebral infarct volume and the number of apoptotic neurons were measured by neurological function score, 2, 3, 5‑triphenyltetrazolium chloride (TTC) staining and terminal deoxynucleotidyl transferase‑mediated dUTP‑biotin nick end labeling (TUNEL) staining, respectively. The levels of MTA1, cleaved caspase‑3 and von willebrand factor (vWF) protein were detected by Western blot. The levels of MTA1 mRNA were determined by real‑time polymerase chain reaction (RT‑PCR). The density of microvessels was detected by immunofluorescence. The four groups were compared for the changes in arterial partial pressure of oxygen (PaO2), arterial partial pressure of carbon dioxide (PaCO2), pH value, cerebral blood flow and anal temperature before, during, and after MCAO surgery. Results Compared with the MCAO group, the EA group and Control siRNA group showed increases in neurological function score, the levels of vWF protein and the density of microvessels (P<0.05), as well as decreases in infarct volume, the levels of cleaved caspase‑3 and the number of apoptotic neurons (P<0.05); the levels of MTA1 protein and MTA1 mRNA in the EA group increased (P<0.05). Compared with the Control siRNA group, the MTA1 siRNA group presented decreases in neurological function score, the levels of vWF protein and the density of microvessels (P<0.05), as well as increases in infarct volume, the levels of cleaved caspase‑3 and the number of apoptotic neurons (P<0.05). Compared with the Sham group, the cerebral blood flow in the other four groups significantly decreased during operation (P<0.05), the levels of MTA1 protein and MTA1 mRNA increased in the MCAO and EA groups (P<0.05). There was no statistical difference in other indicators among other groups (P>0.05). Conclusion EA enhances angiogenesis and improves neurological function after cerebral ischemia through up‑regulating MTA1.
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