Acupuncture (AC) has been demonstrated to improve outcomes in ischemic stroke. However, its precise mechanism of action remains incompletely understood. This study investigated the effects of AC in a rat model of cerebral ischemia-reperfusion injury (CIRI). A middle cerebral artery occlusion/reperfusion (MCAO/R) model was established to create CIRI rats. Rats were randomly assigned to the Sham, MCAO/R, AC + MCAO/R, IRP2(+) + MCAO/R, IRP2(-) + MCAO/R and IRP2(-) + AC + MCAO/R groups. Neurological function was assessed using the Garcia scores. TTC staining was used to observe cerebral infarction volume ratios. Transmission electron microscopy (TEM) examined mitochondrial structure in neurons within the ischemic penumbra. Colorimetric assays measured Fe2+ content in neurons, along with ferroptosis-related biomarkers Reactive oxygen species (ROS), Glutathione peroxidase 4 (GPX4), Malondialdehyde (MDA) and Glutathione (GSH). Immunofluorescence staining detected Transferrin receptor 1 (TFR1), Ferritin (FER), Ferroportin (FPN) and Iron regulatory protein 2 (IRP2) expressions in the ischemic penumbra. RT-qPCR measured IRP2 mRNA expression, whilst RNA immunoprecipitation (RIP) analysis assessed the impact of each group on IRP2 binding to IRE. Our results showed AC improved neurological deficits, reduced infarct volume, decreased Fe2+ overload, enhanced antioxidant markers GPX4 and GSH, and lowered lipid peroxidation levels of ROS and MDA. It also attenuated mitochondrial structural damage related to ferroptosis. Immunofluorescence staining and RT-qPCR analyses revealed that AC downregulated TFR1 and FER expression whilst upregulating FPN and IRP2 expression. Furthermore, RNA immunoprecipitation assays revealed enhanced binding affinity between IRP2 and IRE after AC. These findings indicate that AC inhibits neuronal ferroptosis after CIRI and protects brain tissue, possibly by promoting IRP2-IRE binding, modulating neuronal iron metabolism, and reducing lipid peroxidation.

Cerebral ischemia-reperfusion injury (CIRI) is the second leading cause of death worldwide, posing a huge risk to human life and health. Therefore, investigating the pathogenesis underlying CIRI and developing effective treatments are essential. Ferroptosis is an iron-dependent mode of cell death, which is caused by disorders in iron metabolism and lipid peroxidation. Previous studies demonstrated that ferroptosis is also a form of autophagic cell death, and nuclear receptor coactivator 4(NCOA4) mediated ferritinophagy was found to regulate ferroptosis by interfering with iron metabolism. Ferritinophagy and ferroptosis are important pathogenic mechanisms in CIRI. This review mainly summarizes the link and regulation between ferritinophagy and ferroptosis and further discusses their mechanisms in CIRI. In addition, the potential treatment methods targeting ferritinophagy and ferroptosis for CIRI are presented, providing new ideas for the prevention and treatment of clinical CIRI in the future.

We have previously reported that the expression of miR-34c-5p was up-regulated during acupuncture treatment in the setting of a cerebral ischemia/reperfusion injury (CIRI), indicating that miR-34c-5p plays an important role in healing from a CIRI-induced brain injury. This study sought to evaluate the effects of acupuncture on miR-34c-5p expression and autophagy in the forward and reverse directions using a rat focal cerebral ischemia/reperfusion model. After 120 minutes of middle cerebral artery occlusion and reperfusion, rats were treated with acupuncture at the ''Dazhui'' (DU20), ''Baihui'' (DU26) and ''Renzhong'' (DU14) points. Neurologic function deficit score, cerebral infarct area ratio, neuronal apoptosis and miR-34c-5p expression were evaluated 72 hr after treatment. The autophagy agonist RAPA and the antagonist 3MA were used to evaluate the neuro protective effects of autophagy-mediated acupuncture. We found that acupuncture treatment improved autophagy in the brain tissue of CIRI rats. Acupuncture reversed the negative effects of 3MA on CIRI, and acupuncture combined with RAPA further enhanced autophagy. We also found that acupuncture could increase miR-34c-5p expression in hippocampal neurons after ischemia/reperfusion. Acupuncture and a miR-34c agomir were able to enhance autophagy, improve neurologic deficits, and reduce the cerebral infarct area ratio and apoptosis rate by promoting the expression of miR-34c-5p. Silencing miR-34c resulted in a significantly reduced activating effect of acupuncture on autophagy and increased apoptosis, neurologic deficit symptoms, and cerebral infarct area ratio. This confirms that acupuncture can upregulate miR-34c-5p expression, which is beneficial in the treatment of CIRI.

Objective: This study aimed to observe the effects of acupuncture on iron metabolism in the hippocampus of rats with cerebral ischemia-reperfusion injury (CIRI) and to explore the mechanism of acupuncture on neural repair in CIRI rats. Methods: After general feeding for 3 days, 16 of 48 healthy Sprague-Dawley rats were selected according to a random number table to form a sham-occlusion group, and the models were prepared in the remaining rats. After successful modeling, the rats were randomized into model and acupuncture groups, with 16 rats in each group. The CIRI models were prepared using the modified Zea Longa suture-occlusion method. In the acupuncture group, the rats were fixed and acupuncture was delivered at "Shuigou" (GV26), "Dazhui" (GV14), and "Baihui" (GV20), and the needles were manipulated once every 15 min. In the sham-occlusion and model groups, rats were fixed without acupuncture. The intervention was performed for 30 min in each group, once every 12 h, seven times. The degree of neurological impairment was assessed by using the modified Garcia score. Cerebral blood flow (CBF) was monitored using a laser speckle blood flow imaging system. The cerebral infarction area ratio was measured using triphenyltetrazolium chloride staining. Ferric ion aggregation in brain tissue was noted using Prussian blue staining. The ferric ion and glutathione (GSH) contents of the hippocampal tissue on the ischemic side were detected using ferric ion and GSH kits. The protein expression of transferrin receptor 1 (TFR1), iron regulatory protein 2 (IRP2), and ferroportin (FPN) was detected via western blotting. Results: (1) Before intervention, compared with the sham-occlusion group, the Garcia neurological score was significantly reduced in the model and acupuncture groups (both P < 0.01). After the intervention, compared with the sham-occlusion group, the Garcia neurological score in the model group decreased significantly ( P < 0.01), and the score in the acupuncture group was elevated compared with the model group ( P < 0.05). Compared to before the intervention, the Garcia neurological score increased after the intervention in the acupuncture group ( P < 0.05). (2) Compared with the sham-occlusion group, in the model group, CBF was reduced, the infarct area was significantly enlarged, ferric ion aggregation increased in the hippocampus, the content of ferric ion increased, and that of GSH decreased significantly (both P < 0.01), while the protein expression of TFR1 and IRP2 was upregulated and that of FPN was downregulated (both P < 0.05). When compared with the model group, in the acupuncture group, CBF was increased, the infarct area was reduced ( P < 0.01), the ferric ion aggregation was reduced in the hippocampus, the content of ferric ion decreased, and that of GSH increased significantly ( P < 0.05, P < 0.01); the protein expression of TFR1 and IRP2 was downregulated and that of FPN was upregulated (both P < 0.05). Conclusion: Acupuncture can alleviate CIRI, which may be related to the regulation of iron homeostasis- associated transferrin in the hippocampus, attenuation of ferric ion aggregation in hippocampal tissue, reduction of lipid peroxidation, and inhibition of ferroptosis. (c) 2024 World Journal of Acupuncture - Moxibustion House. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co. Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)

目的 研究针刺对脑缺血再灌注损伤(cerebral ischemia reperfusion injury,CIRI)大鼠海马区环状RNA_009775 表达的影响,探索针刺对CIRI可能的作用机制.方法 将 39只SD大鼠随机分为假手术组、模型组、针刺组,每组 13只.除假手术组外,其余两组均采用线栓法制备CIRI模型.造模后,针刺组大鼠捆绑固定行针刺干预,间隔 12h进行 1次,共 7次;假手术组与模型组大鼠仅捆绑不针刺.Garcia评分法、TTC染色法评估针刺疗效;基因芯片技术筛选针刺组/模型组、模型组/假手术组核心共同差异表达circRNA;qRT-PCR法对芯片筛选结果进行验证;TargetScan软件预测核心circRNA的下游miRNA及mRNA,构建核心circRNA的调控网络,并对mRNA进行基因本体功能富集分析;Western blot法验证神经元核抗原抗体(neuron specific nuclear protein,NEUN)表达量.结果 干预后,与假手术组相比,模型组大鼠Garcia评分、NEUN和circRNA_009775 表达量显著降低(P＜0.01),脑梗死面积比显著升高(P＜0.01);与模型组相比,针刺组脑梗死面积比显著降低(P＜0.01),Garcia评分、NEUN和circRNA_009775 表达量显著升高(P＜0.05 或P＜0.01).芯片筛选核心共同差异表达circRNA,经qRT-PCR验证circRNA_009775 表达趋势与芯片结果一致,结合靶关系预测,构建了含有 1个circRNA、12个miRNA、31个mRNA的调控网络.基因本体功能富集分析显示mRNA主要涉及神经系统发育、神经元生成等生物过程.结论 针刺具有改善CIRI大鼠神经功能缺损、降低脑梗死面积、部分减轻海马区神经元损伤的作用,其机制可能与调控circRNA_009775 介导的潜在调控网络、促进神经系统发育、神经元生成等生物过程有关.

目的:探讨针刺对脑缺血再灌注损伤(CIRI)大鼠海马区细胞凋亡的影响及机制.方法:SD大鼠共75只,随机分为假手术组(sham)、脑缺血再灌注损伤模型组(CIRI)和针刺治疗组(CIRI+AC),利用大脑中动脉栓塞法制备CIRI大鼠模型,CIRI+AC组大鼠接受针刺"大椎"、"水沟"、"百会"穴治疗.Garcia评分法检测大鼠神经功能,TTC染色法检测大鼠脑梗死面积,免疫组织化学染色检测海马区caspase-3阳性细胞数量,Western Blot检测海马区caspase-9表达,TUNEL检测细胞凋亡.结果:针刺前,CIRI组和CIRI+AC组神经功能评分较sham组显著降低(P＜0.01);针刺24、72 h后,CIRI+AC组神经功能评分较CIRI组显著升高(P＜0.01),脑梗死面积比缩小(P＜0.05),患侧脑组织细胞凋亡率降低(P＜0.01);与针刺24 h后比较,72 h后CIRI组患侧脑组织细胞凋亡率及海马区caspase-3阳性细胞数量增加(P＜0.05),CIRI+AC组患侧脑组织细胞凋亡率及海马区caspase-3阳性细胞数量明显降低(P＜0.05).与CIRI组比较,针刺24 h后CIRI+AC组患侧海马区caspase-9表达显著升高(P＜0.01),针刺72 h后caspase-9表达显著降低(P＜0.01).结论:持续针刺可改善大鼠神经功能和细胞凋亡情况,且针刺对CIRI大鼠海马区细胞凋亡的抑制具有时效性.

ObjectiveTo explore the mechanisms of acupuncture against cerebral ischemia/reperfusion injury （CIRI） through observing the expression of circular RNA HDAC2 （circHDAC2） in the hippocampus of rats.MethodsSD rats were randomly divided into sham-operation， model and acupuncture groups， with 13 rats in each group. The rat model of CIRI was established by middle cerebral artery occlusion. In the acupuncture group， acupuncture was delivvered at “Dazhui” （GV14）， “Shuigou” （GV26） and “Baihui” （GV20）， and the needles were retained for 30 min each time and acupuncture was conducted once every 12 h for a total of 7 sessions. Before and after intervention， using modified Garcia scale， the neurological function of the rats were evaluated， and TTC staining was employed to determine the cerebral infarct area. Gene chip technology was used to screen the circRNAs with differential expressions in the ischemic hippocampus， and the circRNAs with co-differential expression （co-DE circRNAs） in the model group/sham-operation group， and the acupuncture group/model group separately. Among those circRNAs， the core circRNAs were screened according to P value， fold change （FC） and gene ontology （GO） analysis； and their expressions in the ischemic hippocampus were determined using quantitative real-time PCR （qPCR）. Based on the verification results， a competing endogenous RNA （ceRNA） prediction network was constructed. The expression levels of microRNA （miRNA） and mRNA with high node centrality in the prediction network were validated by qPCR.ResultsBefore intervention， compared with the sham-operation group， the modified Garcia score of each modeling group decreased （P<0.01）. After intervention， the modified Garcia score was reduced and the cerebral infarct area ratio increased （P<0.01） in the model group when compared with the sham-operation group. In the acupuncture group， the modified Garcia score was higher and the cerebral infarct area ratio lower （P<0.01） than those of the model group. The microarray results of gene chip showed that 16 co-DE circRNAs were down-regulated in the model group and up-regulated in the acupuncture group， and 7 co-DE circRNAs up-regulated in the model group and down-regulated in the acupuncture group. The core circHDAC2 and circNTRK2 were screened according to P value， FC and the enrichment number of GO entries. QPCR results showed that， compared with the sham-operation group， the expression of circHDAC2 and circNTRK2 of the ischemic hippocampal tissue was down-regulated in the model group （P<0.01）； and that of circHDAC2 and circNTRK2 up-regulated in the acupuncture group when compared with the model group （P<0.01）. The relevant ceRNA regulatory network was constructed for circHDAC2 and the prediction results showed that the regulatory networks contained 12 miRNAs and 31 mRNAs. Results of verifying miRNA with high network node centrality and mRNA relevant with nerve regulation showed that， when compared with the sham-operation group， the expression levels of miR-29a， miR-29b and the solute carrier family 30 member 3 （SLC30A3） mRNA in the ischemic hippocampus were down-regulated （P<0.01）； and those of miR-3065 and mercaptopyruvate sulfurtransferase （MPST） up-regulated （P<0.01） in the model group. Compared with the model group， the expressions of miR-29a， miR-29b and SLC30A3 mRNA in the ischemic hippocampus were up-regulated （P<0.01，P<0.05）， while that of miR-3065 down-regulated （P<0.05） in the acupuncture group.ConclusionAcupuncture significantly improves the neurological function and reduces the cerebral infarct area in CIRI rats， which may be related to the regulation of hippocampal circHDAC2/miR-3065/SLC30A3 axis.

探讨"心胀咽痛,刺太冲必除之"理论依据,扩展太冲穴临床应用.从心与肝生理病理、气血调节联系、经络循行理论以及标本根结理论四方面分析,得出肝疏泄失常致心主血脉、心主神志功能异常,故治心脏疾患需先调肝清其源;肝与心,气与血的调节相互影响,调气为主,调血为辅,不可忽视肝调节气机对维持其他脏腑生理功能的重要性;太冲穴为肝经的原穴,针刺其可治疗胸腹部疾病;针刺太冲可调理气机不畅,血行不利所致心神疾患,故"心胀咽痛,刺太冲必除之"理论可指导临床应用太冲穴治疗胸闷心烦、短气、咽痛、卧不安等.

Objective To investigate the effects of abdominal Tuina (Chinese therapeutic massage) on behavioral function, 5-hydroxytryptamine 1A receptor (5-HT 1A R), and synapsin-1 (Syn1) in neonatal rats with hypoxic-ischemic brain injuries (HIBI). Methods Forty healthy neonatal rats, born of 5 specific pathogen-free healthy pregnant rats, were randomly divided into a group for modeling ( n =28) and a sham operation group ( n =12) on the 7th day of birth. In the group for modeling, 24 neonatal rats with HIBI successfully established by the Rice method were randomly divided into a model group ( n =12) and an abdominal Tuina group ( n =12). The abdominal Tuina group was given abdominal Tuina for 28 d from 24 h after modeling, and the other groups were put under the same conditions but without any treatments. Rats in each group were subjected to suspension tests on the 7th, 14th, 21st, and 28th days of intervention. After the intervention, the rat hippocampal tissue was collected and stained with hematoxylin-eosin to observe the pathological changes in the rat hippocampal CA1 region. The 5-HT 1A R expression in rat hippocampal CA1 region was detected by immunehistochemistry. The Syn1 expression in rat hippocampus was measured by Western blotting method. Results The cells were disordered, and edema and necrosis appeared in the hippocampal CA1 region of the model group. Cell arrangement was clear, and edema was improved obviously in the hippocampal CA1 region of the abdominal Tuina group. Compared with the sham operation group, the suspension test scores, the number of 5-HT 1A R positive cells, and Syn1 protein expression in the hippocampus decreased significantly in the model group after 21 d and 28 d of interventions ( P <0.05). Compared with the model group, the suspension test scores, the number of 5-HT 1A R positive cells, and Syn1 protein expression increased significantly in the abdominal Tuina group after 21 d and 28 d of interventions ( P <0.05). Conclusion Abdominal Tuina improves the behavioral function of upper limbs and up-regulates the expression levels of 5-HT 1A R and Syn1 in the hippocampus of neonatal HIBI rats.

"或针结,针着大肠泄水穴"指以病邪凝结经络,气血运行受阻为主要病机之结病,可针刺大肠俞和二间两个穴位.经络腧穴理论中,大肠俞属足太阳膀胱经,是大肠之经气转输、聚结体表所在,有疏调肠胃、宣导气血之功,二间为大肠经的水穴,可针泻用以治疗大肠经的各种实证.标本气街理论中,大肠俞属于背俞穴,位于标部,二间穴属于五输穴,位于本部,本部与标部穴位相配可流转十二经气,利于气血运行.气街与标本关系密切,大肠俞位于背部,可治疗相应脏腑疾病.气血津液与脏腑学说中,针刺二间穴可以使大肠传化糟粕,调节津液代谢的功能恢复正常.针刺大肠俞可宣泄水液,调节膀胱的津液代谢.基于以上理论探讨"或针结,针着大肠泄水穴"中选穴及配穴方法切实可行,可为临床治疗结病提供参考.

目的 探讨针刺对脑缺血再灌注损伤(CIRI)大鼠脑保护作用与海马组织差异表达核心基因、转录网中信使RNAs(mRNAs)功能之间的联系,为针刺调控环状RNAs(circRNAs)抗CIRI后神经损伤研究提供参考.方法 将54只SD大鼠随机分为假手术组、模型组和针刺组,每组18只,后两组采用改良Longa线栓法制备大脑中动脉阻塞再灌注模型,假手术组仅剥离血管,不插线栓.干预期内针刺组捆绑于鼠板并针刺"大椎""水沟""百会"穴,均匀捻转刺激1 min,频率为90次/min,15 min后行针1次,共留针30 min,每12 h干预1次,共7次.假手术组、模型组捆绑于鼠板不针刺.改良加西亚(Garcia)评分法检测大鼠神经功能状况,2,3,5-氯化三苯基四氮唑(TTC)染色法观察脑梗死面积;通过基因芯片技术筛选缺血侧海马组织差异表达基因,整理出模型组/假手术组、针刺组/模型组共同差异表达的环状RNAs(co-DE circRNAs),根据P值、差异倍数(FC)、基因本体(GO)数据库条目富集数量筛选出高表达的核心co-DE circRNAs,分析核心co-DE circRNAs来源基因的GO功能;统计缺血侧海马CA1区尼氏染色阳性细胞数;通过实时荧光定量逆转录聚合酶链式反应(RT-qPCR)法验证核心co-DE circRNAs表达量,选取表达趋势与芯片结果一致的circRNAs进行竞争性内源RNA(ceRNA)网络预测,对预测网络中的mRNAs进行GO功能富集分析.结果 干预前,与假手术组比较,各造模组改良Garcia 神经功能评分显著降低(P<0.05);干预后,与假手术组比较,模型组脑梗死面积比显著升高,改良 Garcia 神经功能评分和CA1区尼氏染色阳性细胞数显著降低(P<0.05);与模型组比较,针刺组脑梗死面积比显著降低,改良Garcia 神经功能评分和CA1区尼氏染色阳性细胞数显著升高(P<0.05);组内干预前后比较,针刺组改良Garcia 神经功能评分显著升高(P<0.05).芯片结果显示,模型组下调/针刺组上调、模型组上调/针刺组下调的co-DE circRNAs为23个,根据FC>1.25和GO数据库条目富集数量筛选出核心基因为环状RNA组蛋白去乙酰化酶2基因(circHDAC2)、环状RNA神经营养受体酪氨酸激酶2基因(circNTRK2);GO数据库分析显示,二者富集于神经元的产生、神经系统发育、神经元分化等条目.验证其表达量发现,与假手术组比较,模型组大鼠缺血侧海马组织circHDAC2、circNTRK2 表达量均显著下调(P<0.05);与模型组比较,针刺组大鼠缺血侧海马组织circHDAC2、circNTRK2表达量均显著上调(P<0.05).circHDAC2表达趋势与芯片结果一致,由此对circHDAC2 构建相关 ceRNA调控网络,调控网络中mRNAs的GO分析主要富集在神经元的产生、神经元分化的调控、轴突引导、神经系统发育、神经元死亡的调节等条目.结论 针刺可改善CIRI大鼠改良 Garcia神经功能评分与脑梗死面积比,降低缺血侧海马组织的神经元损伤,其作用可能与调控缺血侧海马组织circHDAC2表达以及激发调控网络中mRNA的神经元产生、神经元分化、神经元细胞体功能等有关.

国医大师熊继柏教授不仅是临床大家,同时,从教40余年,其治学及教育教学水平也得到了学界公认.熊教授在教育教学中体现出了以学识铸信念、以医理示逻辑、以经典为根本、以临床为指规等教育思想,强调在教学中输出学识、启发思维、融贯医理、突出临床.这是宝贵的学术资源,能为高等中医院校教育提供有指导价值的思路与方法,值得深入的发掘与传承.

目的 神庭穴是针灸临床常用腧穴之一,通过分析、整理涉及神庭穴应用的古代文献资料,总结归纳神庭穴在古代临床中的应用规律.方法 以"神庭""天庭"为检索词,通过检索《中华医典》中所载全部古医籍,对神庭穴主治病症、配伍主治病症、刺灸法、刺激量等相关条文归纳整理并建立数据库.结果 共检索出与神庭相关的文献条文287条,囊括67本古籍,与神庭穴主治相关条文159条,古人运用神庭穴治疗20种病症,涉及内科、五官科、儿科等多门学科.其中单穴及配伍主治病症频次最高的均为心系病症,配伍腧穴共119个,最常与百会、上星、前顶等督脉穴位配伍应用.涉及神庭穴刺灸法的条文共有126条,主要的治疗方法是灸法,刺激量以灸3壮、7壮为多.结论 古代文献中神庭穴的应用内容记载丰富,治疗疾病的病种广泛,可为神庭穴的现代临床应用提供参考.

目的 探讨霉帕霉素(RAPA)作用下针刺激活Ⅲ型PI3K-Beclin1通路抗脑缺血再灌注后细胞自噬的研究,为临床针刺治疗脑血管疾病提供一定的实验依据.方法 SD健康55只大鼠,随机分为假手术组11只、造模组44只.造模组参考线栓法制备大鼠CIRI模型.造模成功后再随机分为模型组、针刺组、模型+RAPA组和针刺+RAPA组.大鼠生命体征稳定后,假手术组、模型组、模型+RAPA组只捆绑固定不针刺;针刺组和针刺+RAPA组捆绑固定并针刺"大椎"、"人中"、"百会"穴,针后行捻转手法1 min(频率90次/min),每15 min行捻转手法1 min,留针30 min,每12h针刺1次,共7次.干预前及72h后进行改良Garcia评分观察大鼠神经功能损伤程度,TTC染色法观察大鼠脑梗死面积比,Western Blot法检测Ⅲ型PI3K、Beclin-1、LC3Ⅱ/Ⅰ、Lamp2、P62蛋白的表达水平,透射电镜观察自噬小体.结果 ①改良Garcia神经功能评分:干预前,与假手术组比较,各造模组改良Garcia评分显著降低(P＜0.01).干预后,与假手术组比较,各造模组改良Garcia评分明显降低(P＜0.01);与模型组比较,针刺组、模型+RAPA组和针刺+RAPA组改良Garcia评分明显升高(P＜0.01).组内干预前后比较,针刺组、模型+RAPA组和针刺+RAPA组较干预前改良Garcia评分上升(P＜0.01).②脑梗死面积比:与假手术组比较,各造模组脑梗死面积比明显升高(P＜0.01);与模型组比较,针刺组、模型+RAPA组和针刺+RAPA组大鼠脑梗死面积比明显降低(P＜0.01);与针刺组、模型+RAPA组比较,针刺+RAPA组大鼠脑梗死面积比降低(P＜0.01).③Ⅲ型PI3K、Beclin-1、LC3Ⅱ/Ⅰ、Lamp2、P62表达水平:与假手术组比较,模型组Ⅲ型PI3K、Bec-lin-1、LC3Ⅱ/Ⅰ、Lamp2表达水平显著降低,P62表达水平显著增高(P＜0.01);与模型组比较,针刺组、模型+RAPA组、针刺+RAPA组Ⅲ型PI3K、Beclin-1、LC3Ⅱ/Ⅰ、Lamp2表达显著升高,P62表达水平降低(P＜0.01);与模型+RAPA组比较,针刺+RAPA组LC3Ⅱ/Ⅰ表达水平升高,P62表达水平降低(P＜0.01).④大鼠海马神经元自噬超微结构:假手术组神经元细胞呈轻微水肿,胞膜、核膜清晰、完整,Ly、SL、ASS、AP个别存在.模型组神经元细胞水肿较严重,大多数线粒体局部基质溶解,嵴减少;SL少量存在,未见典型自噬结构.模型+RAPA组神经元细胞呈中度水肿,细胞膜完整,观察到Ly、ASS数量较多.针刺组、针刺+RAPA组神经元细胞整体结构尚可,未见明显损伤,胞膜、核膜清晰、完整,Ly、SL存在,可见ASS、AP.结论 针刺激活Ⅲ型PI3K-Beclin1通路,增加自噬标志物LC3Ⅱ/Ⅰ、Lamp2的表达,降低溶酶体标志物P62的表达,以激活适度自噬减轻CIRI.

目的:探讨针刺对脑缺血再灌注损伤(CIRI)大鼠脑组织的保护作用,观察针刺对CIRI大鼠缺血侧海马组织环状RNAs(circRNAs)差异表达的影响,并对其进行基因本体(GO)分析.方法:6～8周龄SD大鼠54只,运用随机数字法随机分为造模组和假手术组(sham组),造模成功后再随机分为模型组(model组)和针刺组(AC组),每组18只.采用改良Longa线栓法制备大脑中动脉闭塞再灌注(MCAO/R)模型,激光散斑成像仪监测造模前、MCAO手术后及再灌注后脑血流量,假手术组只剥离血管,不插入线栓;干预期间模型组和假手术组只捆绑不针刺,针刺组捆绑+针刺.采用改良加西亚(Garcia)评分法对神经功能进行评定,TTC染色法检测脑梗死面积,Western blot法检测神经元核抗原(NeuN)的表达,尼氏染色法观察缺血侧海马组织神经元损伤程度,基因芯片微阵列分析筛选出缺血侧海马组织差异表达的circRNAs,并对模型组/假手术组、针刺组/模型组共同差异表达circRNAs的来源基因进行GO分析.结果:与假手术组比较,模型组大鼠脑梗死面积比显著升高(P<0.01),Garcia神经功能评分、NeuN表达量和海马CA1区尼氏染色阳性细胞数显著降低(P<0.05或P<0.01);与模型组比较,针刺组脑梗死面积比显著降低(P<0.01),神经功能评分、NeuN表达量和尼氏染色阳性细胞数显著升高(P<0.05或P<0.01).芯片筛选结果显示,与假手术组比较,模型组上调的circRNAs个数为288,下调个数为315;与模型组比较,针刺组上调的差异表达circRNAs个数为33,下调个数为18(FC>1.25,P<0.05);其中模型组/假手术组、针刺组/模型组共同差异表达的circRNAs个数为23个;GO分析显示共同差异表达circRNAs的来源基因功能涉及神经系统发育,神经元的产生、发育、分化及投射,头部、大脑及海马的发育,突触的形成、发育、延伸及运输等.结论:CIRI大鼠缺血侧海马组织circRNAs在造模后及针刺干预后均存在差异表达.针刺能显著改善CIRI大鼠的神经功能和脑梗死面积,减轻海马组织神经元损伤,其机制可能与针刺调控缺血侧海马组织多种circRNAs的差异表达及激发其促进神经元发育分化、抗神经损伤等功能有关.

目的:利用大脑中动脉闭塞/再灌注(MCAO/R)法制备大鼠脑缺血再灌注损伤(CIRI)模型,研究针刺对CIRI模型大鼠脑组织中Ⅲ型磷脂酰肌醇-3-羟激酶(PI3K-Ⅲ)/Beclin-1通路的影响.方法:利用MCAO/R方法制备SD大鼠CIRI模型,分别给予针刺和(或)自噬抑制剂3-甲基腺嘌呤(3-MA)进行治疗.利用采用Gar-cia评分法检测大鼠神经功能,TTC染色法检测大鼠脑梗死面积,Western Blot法检测海马组织PI3K-Ⅲ、Beclin-1和B淋巴细胞瘤-2基因(Bcl-2)蛋白的表达水平,TUNEL法检测细胞凋亡.结果:MCAO/R手术后大鼠神经功能评分降低(P＜0.01),TTC染色观察到脑梗死,同时海马区PI3K-Ⅲ、Beclin-1和Bcl-2表达显著下调(P＜0.01),脑皮质细胞凋亡增多(P＜0.01).针刺治疗不但提高了大鼠神经功能评分(P＜0.01),并且减少了脑梗死面积(P＜0.01);同时显著上调了海马组织中PI3K-Ⅲ、Beclin-1和Bcl-2表达(P＜0.01),并减少了脑细胞凋亡(P＜0.01).3-MA阻断了针刺的治疗作用,而且使脑损伤更加严重.结论:针刺可改善CIRI大鼠神经功能并减轻神经损伤,其机制可能与激活PI3K-Ⅲ/Beclin-1通路增加细胞自噬并减少细胞凋亡有关.

目的 从海马体基因表达方面探讨针刺大椎、百会、水沟穴治疗脑缺血再灌注损伤的可能作用机制.方法 39只SD大鼠随机分为假手术组、模型组、针刺组各13只.模型组和针刺组大鼠使用线栓法制备大鼠脑缺血再灌注损伤模型.造模后,针刺组大鼠捆绑后针刺"百会""水沟""大椎"穴,留针30min,每15 min捻转1次,平补平泻,共针刺7次,每次间隔12 h;假手术组、模型组只捆绑不针刺.于针刺前、针刺后进行神经功能缺损评分;针刺后TTC染色观察脑梗死情况,采用circRNAs微阵列芯片分析大鼠缺血侧海马区circRNAs差异表达,并用qRT-PCR法验证差异基因的表达.结果 针刺前,与假手术比较,针刺组、模型组神经功能评分显著降低(P＜0.01);针刺后,针刺组神经功能评分较本组针刺前提高,亦高于模型组(P＜0.05).与模型组比较,针刺组脑梗死面积比下降(P＜0.05).CircRNAs微阵列芯片检测结果显示,模型组/假手术组差异表达的circRNAs共215个,其中上调59个,下调156个;针刺组/假手术组差异表达的circRNAs共5个,其中上调1个,下调4个;针刺组/模型组差异表达circRNAs 2个,均上调.模型组/假手术组与针刺组/模型组共表达的差异基因有2个,分别为circ-011989和circ-009775.选取共表达差异基因Ilog2 FCI≥1.5最高的差异基因circ-011989进行验证,结果显示,与假手术组比较,模型组大鼠海马区circ-011989表达量减少(P＜0.05);与模型组比较,针刺组大鼠海马区circ-011989表达量增加(P＜0.05).结论 针刺"大椎""百会""水沟"穴可改善脑缺血再灌注损伤模型大鼠神经功能,减小脑梗死面积,其机制可能与调控缺血侧海马区circ-011989的表达有关.

OBJECTIVE To observe the effects of acupuncture on the expression of type Ⅲ phosphatidylinositol 3-hydroxykinase (PI3K) and Beclin-1 in hippocampus of rats with cerebral ischemia/reperfusion injury (CI/RI), so as to explore the mechanism of acupuncture in regulating type Ⅲ PI3K pathway to activate autophagy in the hippocampal neurons of CI/RI rats. METHODS SD rats were randomly divided into sham operation group (n=11) and operation group. Then after successful modeling, rats in the operation group were randomly divided into model, acupuncture, model+3-MA and acupuncture+3-MA groups, with 11 rats in each group. The model of CI/RI was established by occlusion of the middle cerebral artery. Rats in the model+3-MA and acupuncture+3-MA groups were injected with 3-MA (400 nmol/ 5 μL) 5 μL into the lateral ventricle 30 min before reperfusion. Rats in the acupuncture and acupuncture+3-MA groups were punctured with filiform needles at "Dazhui" (GV14), "Shuigou" (GV26) and "Baihui" (GV20) and stimulated manually once every 15 min. The acupuncture intervention was conducted for 30 min each time, once every 12 h for a total of 7 times. The degree of neurological impairment was evaluated 2 h after reperfusion and after intervention by Garcia score. After intervention, the percentage of cerebral ischemic area was observed by TTC staining, the protein expression levels of type Ⅲ PI3K, Beclin-1, microtubule-associated protein 1 light chain 3B (LC3B), lysosome associated membrane protein 2 (Lamp2) and P62 in ischemic hippocampal tissue were detected by Western blot, the ultrastructure of neurons in ischemic hippocampus was observed by transmission electron microscopy (TEM). RESULTS Compared with the sham operation group, the Garcia score was decreased (P<0.01), the percentage of cerebral ischemic area was increased (P<0.01), the expression levels of type Ⅲ PI3K, Beclin-1, LC3B-Ⅱ/Ⅰ, Lamp2 proteins were decreased (P<0.01), and the expression level of P62 protein was increased (P<0.01) in ischemic hippocampal tissue in the model group. Compared with the model group, the Garcia score was increased (P<0.01), the percentage of cerebral ischemic area was decreased (P<0.01), the expression levels of type Ⅲ PI3K, Beclin-1, LC3B-Ⅱ/Ⅰ, Lamp2 proteins were increased (P<0.01, P<0.05) and the expression level of P62 was decreased (P<0.01) in ischemic hippocampal tissue in the acupuncture group； the percentage of cerebral ischemic area was increased (P<0.05), the expressions of type Ⅲ PI3K and Beclin-1 were decreased (P<0.01) and the expression level of P62 protein was increased (P<0.05) in ischemic hippocampal tissue in the mo-del+3-MA group. Compared with the model +3-MA group, the Garcia score was increased (P<0.05), the percentage of cerebral ischemic area was decreased (P<0.01), the expression levels of type Ⅲ PI3K, Beclin-1, LC3B-Ⅱ/Ⅰ in ischemic hippo-campal tissue were increased (P<0.01, P<0.05) in the acupuncture+3-MA group. Compared with the acupuncture group, the Garcia score was decreased, the percentage of cerebral ischemic area was increased (P<0.05, P<0.01), the expression levels of type Ⅲ PI3K, Beclin-1, Lamp2 proteins were decreased (P<0.01, P<0.05) and P62 protein was increased (P<0.05) in ischemic hippocampal tissue in the acupuncture+3-MA group. The results of TEM showed that the edema of neurons was heavier, and few hypolysosomes existed in the model group; there was no obvious damage to neuronal structure, intracellular matrix was abundant, and a few lysosomes existed in the acupuncture group; the neuronal cells had mild edema and primary lysosomes were present in the acupuncture +3-MA group. CONCLUSION Acupuncture can improve the symptoms of neurological impairment and reduce the percentage of cerebral ischemic area in rats with CI/RI. The mechanism may be related to regulating type Ⅲ PI3K/Beclin-1 pathway, up-regulating the expressions of autophagy related factors LC3B-Ⅱ and Lamp2, and down-regulating the expression of P62.

国医大师熊继柏是驰名海内外的中医临床大家,同时从教40余年,精湛的临床技艺与教学水平得益于他深厚的治学功底.本文围绕熊继柏教授所倡导的"中医的生命力在于临床"这一思想展开研究,总结出这一思想是熊教授治学的核心理念,其在治学上将读书作为中医治学基石,主张通过有目的、有方法、有层次地读书而融贯医理、博采众长,进而在临床中能执全纠偏、活学活用.这一理念的提出与贯彻,对今天中医学术研究、教育教学乃至临床实践有重要的指导意义.

目的 探讨针刺对脑缺血再灌注损伤(cerebral ischemia reperfusion injury,CIRI)大鼠的脑保护作用及缺血侧海马组织铁死亡的影响.方法 65只健康SD大鼠随机分成假手术组、模型组、针刺组、药物组,药物组5只,其他每组20只.线栓法制备大鼠脑缺血模型,2 h后拔出线栓约10 mm,建立CIRI模型.大鼠生命体征稳定后,药物组采用依达拉奉水溶液(5 mg/kg)腹腔注射后捆绑30 min;假手术组、模型组注射同等药物剂量生理盐水,并捆绑30 min;针刺组注射同等药物剂量生理盐水并选取"人中""大椎""百会"穴进行捆绑针刺,留针30 min.均1次/12 h,共7次.采用改良Garcia评分观察大鼠神经功能损伤程度;TTC染色检测脑梗死面积;铁离子、活性氧(reactive oxygen species,ROS)及丙二醛(malondialdehyde,MDA)试剂盒检测海马组织铁离子、ROS及MDA含量.结果(1)干预前,与假手术组相比,其余各组改良Garcia评分显著降低(P<0.01).干预后,与假手术组相比,模型组改良Garcia神经功能评分明显降低,脑梗死面积显著扩大(P<0.01);与模型组相比,针刺组、药物组改良Garcia神经功能评分升高,脑梗死面积缩小(P<0.05,P<0.01).与干预前相比,针刺组、药物组改良Garcia神经功能评分升高(P<0.05).(2)与假手术组相比,模型组海马组织铁离子、ROS及MDA含量均明显增加(P<0.01);与模型组相比,针刺组海马组织铁离子、ROS及MDA含量均明显降低(P<0.01).结论 针刺可以抑制CIRI后大鼠神经细胞铁死亡,发挥脑保护作用,可能与调节神经细胞铁离子代谢、减轻海马组织铁离子集聚、降低脂质过氧化水平有关.