电针刺激对脊髓挫伤模型大鼠神经细胞凋亡的影响

BACKGROUND: Electroacupuncture has promoting effects on the functional recovery of the injured spinal cord, can decrease pain, and elevate postoperative effect after acute spinal cord contusion.  OBJECTIVE: To observe the effect of electroacupuncture on apoptosis in the injured site after spinal cord contusion, and analyze its neuroprotective effects on neurological function in rats with spinal cord contusion. METHODS: A total of 66 adult female Sprague-Dawley rats were randomly divided into: sham surgery group (n=20), spinal cord contusion group (n=20), electroacupuncture stimulation group (n=20) because six rats were excluded due to modeling failure and death. Before model establishment, at 1, 3 days, 1, 2, 3 and 4 weeks after model establishment, motor functions were evaluated by BBB score and the inclined plate test. At 3 days after model establishment, apoptosis of nerve cells could be detected in the site of injury in each experimental group using TUNEL assay. mRNA and protein expression of bax, bcl-2 and caspase-3 was detected surrounding the injury site using RT-PCR and western blot assay. Morphological changes in the site of injury could be observed using hematoxylin-eosin staining. The regeneration of nerve fibers was observed using HRP tracing.  RESULTS AND CONCLUSION: (1) Motor function score was significantly increased at various time points in the 2nd week of treatment in the electroacupuncture stimulation group than in the spinal cord contusion group (P < 0.05). (2) Apoptotic index was significantly lower in the electroacupuncture stimulation group than in the spinal cord contusion group at 3 days after model establishment (P < 0.05). (3) mRNA and protein expression of bax and caspase-3 was significantly lower in the electroacupuncture stimulation group than in the spinal cord contusion group at 72 hours (P < 0.05); bcl-2 gene and protein expression was significantly higher (P < 0.05). (4) The number of HRP-positive nerve fibers was highest in the sham surgery group, followed by electroacupuncture stimulation group, and lowest in the spinal cord contusion group at 4 weeks (P < 0.05). Results indicated that electroacupuncture plays a protective role on the spinal cord contusion by reducing apoptosis of nerve cells at the site of injury.      

激光损伤视网膜后神经元凋亡及GFAP表达的研究

目的 观察兔视网膜激光损伤后神经元细胞有无凋亡改变及视网膜Ｍｕｌｌｅｒ细胞胶质纤维酸性蛋白（ＧＦＡＰ）的表达变化。方法 应用末脱氧核苷酸转移酶介导的ｄＵＴＰ－Ｘ切口末标记法（ＴＵＮＥＬ法）标记凋亡细胞。应用免疫组化染色显示视网膜Ｍｕｌｌｅｒ细胞ＧＦＡＰ表示。结果伤后６ｈ、１、３、７ｄ视网膜各层可见散在分布的ＴＵＮＥＬ阳性凋亡细胞,尤以外核层多见,伤后３ｄ,视网膜可见Ｍｕｌｌｅｒ细胞ＧＦＡＰ表达;伤     

米诺环素抑制谷氨酸诱导的大鼠视网膜神经细胞凋亡

目的：观察米诺环素对谷氨酸诱导的大鼠视网膜神经细胞凋亡的抑制效应及其作用机制。方法:取原代培养的SD乳鼠视网膜神经细胞，随机分为正常对照组、米诺环素对照组（米诺环素20 μmol/L）、谷氨酸组（谷氨酸1 mmol/L）和米诺环素治疗组（米诺环素20 μmol/L＋谷氨酸1 mmol/L）。干预1 h后采用Annexin V/PI流式细胞仪计数凋亡细胞数量，同时检测Rh123以评估线粒体膜电位改变。干预12 h后行MTT检测细胞活性；另取细胞培养上清液做一氧化氮合酶（NOS）的活力单位检测。结果:干预1 h后，流式细胞仪Annexin V/PI检测凋亡细胞比例：正常对照组、米诺环素对照组、谷氨酸组、米诺环素治疗组分别为5.1％、4.3％、15.2％、8.3％。Rh123各组阳性率分别为67.1%、54.2%、27.5%、32.4%。干预12 h后，MTT检测各组细胞吸光度均值分别为： 0.093±0.008、0.099±0.012、0.038±0.008、0.088±0.016。治疗组与正常组之间无显著差异（P>0.05），谷氨酸组与其它各组之间均存在显著差异（P<0.05）。NOS活力单位检测，以正常对照组均数为1，另3组与其比值的均数分别为： 0.987±0.219、1.513±0.472、1.176±0.259。其中谷氨酸组与其它3组之间存在显著差异（P<0.05）。结论:20 μmol/L米诺环素可显著减轻谷氨酸诱导的视网膜神经细胞凋亡，其作用机制与稳定线粒体膜电位，以及抑制NOS活性有关。     

针刺对脑性瘫痪模型大鼠神经细胞凋亡的影响

BACKGROUND:Acupuncture has been shown to impact cerebral blood flow, relieve cerebral edema, improve microcirculation and reduce neural cell apoptosis in animals with cerebral palsy. OBJECTIVE:To further verify the effect of acupuncture on neuronal apoptosis in cerebral palsy rats. METHODS:Forty-five Sprague-Dawley rats were randomly divided into sham group, model group and the acupuncture group, with 15 rats in each group. In the model and acupuncture groups, rat models of cerebral palsy were established by ligating the left common carotid artery. In the sham surgery group, the left common carotid artery was not ligated. Rats in the acupuncture group received acupuncture from 2 days after model induction, once a day, for 20 consecutive days. Rats in the sham surgery and model groups were left intact. At 21 days after surgery, neurological functions, pathomorphological changes and neuronal apoptosis were observed.  RESULTS AND CONCLUSION:(1) Motor function score was significantly lower in the model group than in the sham surgery group (P < 0.05). Motor function score was significantly higher in the acupuncture group than in the model group at 21 days (P < 0.05). (2) Hematoxylin-eosin staining: In the model group, cells were scattered. Inflammatory cell infiltration and cystis degeneration were found. The number of nerve cells was reduced. Degeneration, necrosis, gliosis and karyopyknosis were detected. Cavitation was visible in some cells. Cell bodies became small. The structure was not distinct or disappeared. In the sham surgery group, the morphology and structure of nerve cells were normal. In the acupuncture group, the changes in morphology and structure of nerve cells were found between the sham surgery and model groups. (3) The number of apoptotic cells was significantly greater in the model group than in the sham surgery group (P < 0.05). The number of apoptotic nerve cells was significantly higher in the acupuncture group than in the sham surgery group (P < 0.05), but significantly lower than in the model group (P < 0.05). (4) Results confirmed that acupuncture therapy can improve neurological function, reduce nerve cell apoptosis, and play a protective effect on nerve cells of rats with cerebral palsy.     

RCS大鼠视网膜感光细胞的凋亡

为研究遗传性视网膜变性中感光细胞组织结构的时程变化及调亡，对ＲＣＳ大鼠脑ＳＤ大鼠视网膜进行光镜观察和凋亡细胞ＴＵＮＥＬ检测。结果表明，与同龄ＳＤ大鼠相比，ＲＣＳ大鼠视网膜感光细胞从出生后１５ｄ开始，出现外节膜盘堆积；２０ｄ时，内节排列紊乱，消失，３０ｄ，细胞核固缩，细胞消失，到出生后６０ｄ，仅少许感光细胞保留；１００ｄ，几乎所有感光细胞消失。ＴＵＮＥＬ检测，从出生后２５ｄ开始，ＲＣＳ大鼠视网膜有Ｔ     

菩人丹超微粉对糖尿病大鼠视网膜神经细胞凋亡及相关基因表达的影响

目的: 探讨菩人丹超微粉(PRD)对糖尿病大鼠视网膜神经细胞凋亡及相关基因表达的影响。方法: 36只Wistar大鼠随机分为3组,正常对照组、糖尿病模型组和PRD治疗组,每组12只。糖尿病模型组和PRD治疗组大鼠均采用链脲佐菌素连续腹腔注射建立2型糖尿病大鼠模型。模型成功建立后,PRD治疗组大鼠给予PRD灌胃3个月。采用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法(TUNEL)检测大鼠视网膜神经细胞的凋亡;SP免疫组织化学染色法检测视网膜B细胞白血病/淋巴瘤相关抗原2(Bcl-2)、B细胞白血病/淋巴瘤相关抗原相关X蛋白(bax)和半胱氨酸天冬氨酸蛋白酶3(caspase-3)蛋白的表达;逆转录聚合酶链反应(RT-PCR)检测bcl-2、bax和caspase-3 mRNA的表达。结果: 糖尿病模型组与正常对照组比较,大鼠视网膜神经细胞凋亡指数、Bax、 caspase-3蛋白及mRNA的表达均明显升高(P＜0.01),Bcl-2蛋白及mRNA的表达、Bcl-2/Bax比值显著降低(P＜0.01);PRD治疗组与模型组比较,大鼠视网膜神经细胞凋亡指数、bax、caspase-3蛋白及mRNA的表达均明显降低,Bcl-2 蛋白及mRNA的表达、Bcl-2/Bax比值显著升高(P＜0.01)。结论: PRD可通过上调Bcl-2的表达及下调Bax及caspase-3的表达,抑制糖尿病大鼠视网膜神经细胞的凋亡,发挥对糖尿病视网膜的保护作用。     

氢盐水可抑制脊髓缺血再灌注损伤兔模型运动神经元的凋亡

背景：脊髓缺血再灌注损伤是一种严重的继发性脊髓损伤,其损伤机制是多因素综合作用的结果,其治疗上也有多种措施,但治疗效果不甚理想。 目的：探讨氢盐水对脊髓缺血再灌注损伤兔模型运动神经元的保护作用及机制。 方法：采用ZIVIN法制备脊髓缺血再灌注损伤兔模型,并采用氢盐水治疗(设为氢盐水组),同时设模型组和假手术组为对照。 结果与结论：氢盐水组兔后肢运动功能Tarlov评分于再灌注后6,12,24,72 h明显优于模型组(P < 0.01)。再灌注后72 h,与模型组相比,氢盐水组丙二醛浓度降低(P < 0.05),过氧化氢酶活性升高(P < 0.05)。苏木精-伊红染色显示,假手术组脊髓前角运动神经元细胞结构完整,模型组脊髓前角大量运动神经元细胞坏死,胞浆内颗粒变性和空泡变性。氢盐水组脊髓前角运动神经元细胞结构基本完整,仅有少量运动神经元细胞空泡变性。原位末端标记染色显示,假手术组未见运动神经元细胞凋亡;模型组见大量凋亡的运动神经元及大量炎性细胞浸润;氢盐水组见脊髓前角少量凋亡的运动神经元及少量炎性细胞浸润。结果证实,氢盐水可抑制兔缺血再灌注损伤脊髓运动神经元的凋亡,其机制与其抗氧化作用有关。 中国组织工程研究杂志出版内容重点：肾移植;肝移植;移植;心脏移植;组织移植;皮肤移植;皮瓣移植;血管移植;器官移植;组织工程全文链接：     

外伤性视神经损伤后视神经及视网膜形态的动态观察

目的:了解外伤性视神经损伤后的病理变化、溃变特点与时相间的关系。方法:参照Allen脊髓损伤法,造成视神经眶尖段间接600gcm力冲击、挤压伤。伤后对视神经和视网膜行形态学动态观察。结果:①伤后48h,视神经轻度肿胀和空泡反应;1周时损伤处视神经出现溃变,神经胶质细胞增生,视网膜神经节细胞(retinalganglioncells,RGCs)形态改变不明显;2周时神经纤维轴束间空泡样改变,局灶性坏死,RGCs核固缩和细胞数量减少。术后3月,视神经损伤部位直径缩小,形成胶质疤痕,RGCs数量明显减少,核固缩细胞增多。②RGCs数量于术后48h、1周、2周、1月和3月分别比正常对照组低3.35%、13.23%、19.74%、23.20%、29.28%。③视网膜细胞在48h内出现凋亡。结论:本实验模型可造成明确的视神经和视网膜损伤,神经元的损伤程度从节细胞、中间神经元、感光细胞的次序依次递减。视网膜和视神经损伤的严重程度与时间呈相关性。RGCs数量在48h至1周时下降速率最快。     

视网膜神经胶质细胞及其分泌的细胞因子与视网膜病变

视网膜内常见的神经胶质细胞包括星形胶质细胞、Müller细胞(放射状胶质细胞)和小胶质细胞。最近研究表明,这些神经胶质细胞能释放多种细胞因子,包括VEGF、bFGF、TGF、IGF、TNF及IL等,这些细胞因子不仅在调节视网膜神经细胞生长发育中起着重要作用,而且是参与视网膜病变的重要因素。     

黄嘌呤抗视神经切断后视网膜细胞凋亡的作用

目的　探讨眼球玻璃体注射 3 异丁基 1 甲基 (IBMX)对切断视神经后视网膜细胞凋亡的作用。方法　用荧光核染料 332 5 8和琼脂糖凝胶电脉技术观察切断视神经 5、7和 1 4d后成年金黄地鼠视网膜细胞凋亡的变化。结果　 ( 1 )视神经切断 5、7、1 4d后 ,节细胞层 (GCL)细胞凋亡密度 (cell/mm2 )分别为 94 60± 2 9 40、81 2 0± 1 5 1 9和 31 2 0± 1 1 43,给予IBMX组在上述时间点凋亡密度为 67 80± 1 3 83、60 2 0± 9 68和 2 7 0 0± 9 5 7。IBMX 7d组与对照组比较有显著差异 (P <0 0 5 )。 ( 2 )琼脂糖凝胶电脉显示切断 7d的视网膜细胞DNA抽提物显示典型的DNA梯形条带。其它各间间点 ( 5d和 1 4d)以及玻璃体内注射IBMX均未见典型DNA梯形条带。结论　IBMX有抗视神经切断后视网膜细胞凋亡的作用     

骨髓间充质干细胞调节大鼠视神经损伤后小胶质细胞的活化

背景:外伤性视神经损伤是引起视力丧失的重要原因,治疗方法也比较局限,为探求更好的治疗方法,该实验从小胶质细胞方向入手进行探究。在神经病理条件下,激活小胶质细胞能够维持中枢神经系统的稳定,但小胶质细胞过度活化会产生大量的炎症因子,使损伤进行性加重。目的:探讨视神经损伤后小胶质细胞活化情况以及骨髓间充质干细胞对其过度表达的调节作用。方法:选取8周龄SD大鼠18只,将其随机分为移植组、模型组和假手术组,每组6只,其中模型组、移植组选取左眼进行视神经钳夹造模后分离视网膜和视神经,假手术组只分离视网膜和视神经,不进行钳夹,移植组在损伤后立即向左眼玻璃体内注入第3代骨髓间充质干细胞(注入细胞1×108,细胞量为2μL),模型组、假手术组玻璃体内注入等量的PBS,术后15 d全部处死,在灌流固定后取视网膜连带视神经用于苏木精-伊红染色和免疫组织化学检测。结果与结论:模型组视神经及视网膜小胶质细胞活化标记物Ox-42以及炎症因子TNF-α的表达量均高于假手术组(P<0.05),移植组视神经及视网膜中Ox-42和TNF-α表达量降低(P<0.05)并且接近假手术组水平。结果表明,小胶质细胞的过度活化与视神经损伤相关,骨髓间充质干细胞可以抑制小胶质细胞过度活化及炎症因子的释放,从而在一定程度上保护视网膜和视神经免受损伤。     

晶状体损伤促进视神经再生的作用及机制

目的：研究晶状体损伤对视神经再生的促进作用,并探讨巨噬细胞所起的作用。方法：成年 SD 大鼠钳夹造成视神经损伤模型(NC),戳伤晶状体(LP),玻璃体内注射酵母多糖(ZI)或注射体外活化的单核/巨噬细胞 (MI),动物以此分组。用 Nissl 染色法显示存活的视网膜节细胞(RGCs),用抗 GAP-43抗体标记轴突再生 RGCs,用抗 ED-1抗体标记活化的单核/巨噬细胞。结果：NC LP 组存活的 RGCs 比 NC 组明显增加,再生的 RGCs 及活化的单核/巨噬细胞都有明显增多的结果。NC ZI 组与 NC MI 组也有类似结果。结论：晶状体损伤具有显著促进视神经再生的作用,其机制可能与趋化并激活巨噬细胞有关。     

Impacts of Rho kinase inhibitor Fasudil on Rho/ROCK signaling pathway in rabbits with optic nerve injury

Objective: The aim of this study was to study the impacts of Rho kinase inhibitor Fasudil on expressions of Rho/ROCK signaling pathway associated genes in rabbits with optic nerve injury (ONI), and to explore the therapeutic mechanisms towards ONI. Methods: The rabbit ONI model was established, then the rabbits were divided into model group (treated with saline), control group (treated with dexamethasone, Dex), and intervention group (treated with Fasudil, Fas). The eyeball and optic nerve were sampled at 3, 7, 14 and 21 days after injury. The morphological changes of retina and optic nerve were observed. The expressions of RhoA, Caspase-3, Rock 2 and Nogo-A gene were determined by immunohistochemistry and real-time polymerase chain reaction (RT-PCR) methods. Results: At different time after injury, there were significant differences of RhoA, Caspase-3, Rock 2 and Nogo-A gene expression among three groups (P < 0.05). Conclusions: After ONI, Fas can decrease the expression of Caspase-3 gene, and down-regulate the expressions of Nogo-A and Rock 2 gene. Therefore, it can treat ONI through affecting the Rho/ROCK signaling pathway.     

黄芪甲苷对成年大鼠视神经切断后视网膜节细胞存活的影响

目的:探讨黄芪甲苷(AST)在成年大鼠视神经切断后对视网膜节细胞(RGCs)存活的影响。方法:动物分为正常组、单纯切断视神经组、AST处理组和生理盐水对照组。用荧光金(FG)逆行示踪标记法及定量解剖学技术观察正常和经AST处理的SD大鼠于视神经切断后5、7、14 d的RGCs的密度。结果:正常组RGCs平均密度为(2 230±156)/mm~2。单纯视神经切断组RGCs平均密度与生理盐水对照组相比较,无显著性差异;AST处理组与单纯切断视神经组和生理盐水对照组相比较,在各个时间点上均存在显著性差异。结论:黄芪甲苷可提高成年大鼠视神经切断后视网膜节细胞短期存活。     

夹伤与冰冻损伤大鼠股神经的选择性再生

背景：促进周围神经损伤后神经功能的恢复,一方面要尽量加快损伤处神经轴突的再生,另一方面需要提高近端与远端神经对接的精确度。 目的：观察机械挤压伤与冰冻损伤2种损伤模式下周围神经损伤后神经轴突的选择性再生情况。 方法：取健康8周龄雄性Sprauge-Dawley大鼠110只,分为3组,对大鼠行股神经主干夹伤、冰冻损伤或空白对照处理。造模后分别于第2,3,6,12周进行大体行为学检查,另外分别用纯蓝和荧光红标记错向长入的隐神经和正确长入的股神经肌支,逆行示踪标记运动神经元观测脊髓前角中示踪剂的分布及数目;造模后8周行电生理检查,并进行统计学分析。 结果与结论：夹伤组与冰冻损伤组大鼠术侧后腿活动范围均缩小,伸腿功能均受限,但随时间的延长,功能有所恢复。股四头肌处可记录到运动诱发电位,2组之间差异无显著性意义(P > 0.05)。冰冻损伤组与夹伤组在荧光显微镜下均观察到脊髓前角红染神经元逐渐增加,造模后不同时间点夹伤组红染神经元数目显著高于冰冻损伤组(P < 0.05), 蓝色与紫色神经元数量逐渐减少。结果提示,保持神经束膜的完整性,即使损伤范围较大也能得到轴突再生中准确对接以及损伤肢体功能的恢复。     

不同剂量氯化锂对成年大鼠视神经切断后视网膜神经节细胞的保护作用

目的:研究不同剂量氯化锂(LiCl)对成年大鼠视神经切断后视网膜神经节细胞(RGCs)存活的作用。方法:眶内切断72只成年雌性SD大鼠左侧视神经且残端留置荧光金(FG)后,随机分为生理盐水对照组和低剂量(30 mg/kg/d)、中剂量(60 mg/kg/d)、高剂量(85 mg/kg/d)氯化锂实验组。术前1 d及术后每天腹腔注射生理盐水或不同剂量的氯化锂溶液,直至术后2 d、7 d或14 d处死动物。平铺视网膜后取样计数FG逆行标记的存活节细胞,并由此计算出每一视网膜内节细胞的平均密度。结果:术后2 d各剂量氯化锂组节细胞平均密度与对照组相比无显著性差异(P>0.05)。当存活时间增至7 d时,各剂量氯化锂组节细胞密度均明显高于对照组(P<0.01),且中剂量组节细胞密度增高最为显著。术后14 d时,低剂量与中剂量组节细胞密度仍显著高于对照组(P<0.01),但高剂量组节细胞密度与对照组相比无统计学差异(P>0.05)。结论:腹腔内注射氯化锂可显著促进成年大鼠视神经切断后节细胞的存活,这种神经保护作用为剂量依赖性。     

大鼠视神经吸断伤后相关分子表达的变化

目的：研究视神经损伤后神经胶质细胞去分化的程度及相关分子的表达。方法：成年雄性大鼠视神经吸断伤后,采用免疫组织化学、原位杂交组织化学结合计算机图像分析,检测视神经巢蛋白、胶质纤维酸性蛋白 (GFAP)、髓鞘碱性蛋白(MBP)、神经纤维丝(NF)以及 Nogo-A mRNA 在伤后3、7、14和28 d 4个不同时相点的表达。结果：视神经伤后,巢蛋白表达上调,在28 d 时表达最高;GFAP 表达先下调,7 d 时最低,随后逐渐上调; MBP 表达上调,呈先上升后降低的趋势;NF 表达呈明显下降趋势;Nogo-A mRNA 表达呈上升趋势,3 d 到7 d 的变化最显著。结论：视神经损伤后相关分子表达提示其神经胶质细胞可去分化为神经前体细胞或神经干细胞,但这些前体细胞或干细胞呈不利于神经再生的状态。     

An insight on established retinal injury mechanisms and prevalent retinal stem cell activation pathways in vertebrate models

Implementing different tools and injury mechanisms in multiple animal models of retina regeneration, researchers have discovered the existence of retinal stem/progenitor cells. Although they appear to be distributed uniformly across the vertebrate lineage, the reparative potential of the retina is mainly restricted to lower vertebrates. Regenerative repair post‐injury requires the creation of a proliferative niche, vital for proper stem cell activation, propagation, and lineage differentiation. This seems to be lacking in mammals. Hence, in this review, we first discuss the many forms of retinal injuries that have been generated using animal models. Next, we discuss how they are utilized to stimulate regeneration and mimic eye disease pathologies. The key to driving stem cell activation in mammals relies on the information we can gather from these models. Lastly, we present a brief update about the genes, growth factors, and signaling pathways that have been brought to light using these models.