环孢素A对异种脱细胞神经移植的影响

目的研究脱细胞异种面神经移植中环孢素A（cyclosporin A,CSA）的作用效果。方法 48只Wistar大鼠随机选择一侧作为实验侧,解剖面神经后于颊支制造1cm的神经缺损。按神经移植的种类分为4组,A组：异种面神经移植组;B组：异种面神经移植应用CSA免疫抑制组;C组：异种脱细胞神经移植组;D组：异种脱细胞神经移植应用CSA免疫抑制组。各组实验动物分别于术后5周,12周进行电生理学测试（潜伏期、运动神经传导速度）,并进行光镜、电镜观察形态学变化。结果术后5周和12周时,C,D组动物在电生理指标及光镜、电镜指标上均优于A,B组,术后5周和12周,D组面神经颊支传导速度均高于其它各组。结论 1.0 cm缺损的异种脱细胞面神经移植动物实验中,应用CSA5mg/（kg.d）5周可以降低排斥反应,提高神经再生能力。     

胰腺癌裸鼠皮下异种移植动物模型建立研究

目的:用人体胰腺癌细胞株,建立小鼠胰腺癌细胞株(pGHAM-1)移植性胰腺癌模型,并研究其生物学特性。方法:将pGHAM-1细胞培养后配成1×107/ml,取0.2mL接种于小鼠皮下。于接种后20、30、40天观察肿瘤的生长、转移及腹水量。接种40天后处死动物,解剖取出移植瘤,用游标卡尺测量肿瘤大小,再进行HE染色的组织病理学检查,免疫组织化学检测血管内皮细胞生长因子(VEGF)及肿瘤转移相关蛋白(nm23-H1)的表达。结果:分别于20、30、40天测量肿瘤体积为84.1±21.9 mm3,413.7±208.4 mm3,2187.3±1882.8 mm3,后者与10 d前比较差异均具有统计学意义(P<0.01)。HE染色结果显示肿瘤组织呈条索状或小梁状排列,肿瘤组织与正常组织交界处有少量淋巴细胞浸润,肿瘤组织中血管生成罕见。免疫组织结果显示VEGF和nm23-H1蛋白在肿瘤组织中均呈阴性表达。结论:异位裸鼠人胰腺癌移植瘤模型易复制,时间短,成功率高,为胰腺癌体内研究提供了理想的动物模型。     

生物血管异种移植的初步研究

目的为了寻求一种新的小口径血管代用品,建立异种移植的动物实验模型,以观察异种移植物的安全性、可靠性、通畅性及组织学改变。方法共采用17只杂种雌性犬,实验组10只,植入经环氧化物处理的猪血管移植物;对照组7只,植入人造血管。手术方法为右侧股动静脉瘘。术后通过超声和血管造影方法来观察移植血管的通畅性,并在术后3月将移植物取出,进行病理学检查,观察移植前后移植物的组织学改变。结果术后第一周、二周行Doppler超声检查结果,两组动静脉瘘均通畅,2周内血管通畅率为100%。术后3个月动脉造影检查后,生物血管组(PG)通畅5只,通畅率62.5%,e-PTFE组通畅4只,通畅率66.7%。两组数据统计学处理,差异无显著性(P>0.05)。术后3月对移植物取材,进行光镜及扫描电镜病理学检查,通畅的生物血管吻合口无狭窄,吻合部位有新的内膜覆盖,周围组织无钙化,有新生的内皮细胞覆盖。结论经环氧化物处理的猪的血管移植物(PG)生物血管作为异种移植物,生物相容性好,具有一定的可行性。     

人胎儿卵巢异种移植的组织学研究

目的探讨人胎儿卵巢组织异体移植的最佳条件。方法观察胎儿卵巢进行异种移植后的组织学变化,并计算各组移植卵巢内各期卵泡的发育情况及卵泡构成比。结果新鲜的卵巢组织、体外培养的卵巢组织以及冷冻复苏后经体外培养的卵巢组织移植后,其内有原始卵泡发育,而且可见丰富的血管网;未见明显的淋巴细胞浸润现象。结论人胎儿卵巢组织(包括新鲜的卵巢组织、体外培养的卵巢组织以及冷冻复苏后经体外培养的卵巢组织)异种移植后原始卵泡在受体体内能进一步发育,生长卵泡分泌的雌激素有调节子宫内膜周期性变化的作用。     

异种移植的病毒安全性研究进展

猪-人异种移植有望解决人源器官短缺的严重问题。然而,以前病毒（provirus）形式整合入猪基因组中的猪内源性反转录病毒（porcine endogenous retrovirus,PERV）难以去除,PERV有可能通过异种移植传播给人类,甚至产生新的病毒性疾病。本文回顾了PERV与异种移植病毒安全性及我国特有小型猪中PERV的相关研究。     

小鼠脊髓损伤模型的建立及神经干细胞移植后运动功能恢复情况的研究

目的制作小鼠脊髓损伤打击模型,观察神经干细胞(NSCs)移植对脊髓损伤小鼠运动功能恢复及Nestin表达的影响。方法将50只小鼠随机分为空白组(5只)、模型组(15只)、对照组(15只)、治疗组(15只),运用改良Allen's法制备小鼠T10脊髓损伤模型并立即在损伤节段进行NSCs移植,于损伤后1、3、7、14、21d进行BBB评分,并通过免疫荧光法及荧光定量PCR检测Nestin的表达情况。结果所有脊髓打击后小鼠均出现双后肢瘫痪,但随时间延长运动功能可有不同程度恢复,NSCs移植14d后治疗组较模型组及对照组BBB评分显著增高(P0.05),且治疗组Nestin表达量也高于模型组及对照组。结论成功建立了小鼠脊髓损伤打击模型;移植的外源性神经干细胞在脊髓损伤处存活并促进损伤后小鼠运动功能的恢复。     

异种移植中Galα(1,3)Gal抗原的研究近况

同种异体组织和器官移植物供体来源有限,使得异种移植再度成为移植领域的研究热点。异种移植的主要障碍是人体内存在的天然抗体与移植物表面含有α1,3半乳糖残基[Galα(1,3)Gal,αGal]的抗原结合,激活补体系统和炎症反应,导致超急性移植排斥反应(HAR)的发生,使移植物失活。除人类和旧世纪猴外,其它所有哺乳动物的体内都含有αGal抗原,该抗原是由一组具有Galα(1,3)Gal双糖末端的糖蛋白或糖脂组成的,它的形成依赖于α1,3半乳糖基转移酶(αGT)的催化。目前,针对αGal抗原克服超急性移植排斥反应的方法主要有如下几种：(1)酶处理去除内皮细胞表面的αGal抗原;(2)物理化学方法去除人体血浆中存在的特异性天然抗体;(3)基因工程方法改造表达催化αGal抗原形成的相关酶基因,从而影响该抗原的表达。     

异种移植中转基因动物技术应用的现状

超急性排斥反应一直是异种移植中的主要障碍,转基因技术为跨越这一障碍发挥了重要作用,转基因技术克服超急性排斥反应的主要技术策略有：利用抗补体基因插入技术创建转基因猪,供体抗原遗传修饰和受体基因治疗。     

再生神经中微管,神经丝与轴突截面积的变化

用电镜及图象分析的方法研究了再生轴突中微管、神经丝与轴突截面积的变化,发现神经再生过程中微管及神经丝的密度增加,并与轴突截面积呈相关关系,而且微管的变化更早,更明显。由于微管参与了轴浆转运的机制,微管的增加提示其在神经再生中起了重要的作用。     

神经退变和再生的构筑变化

将夹伤的大鼠坐骨神经分离成单根纤维,观察98d内轴突和许旺细胞的构筑变化过程发现,损伤既使轴浆转运阻断、积累的细胞器退变,也使髓鞘板层,特别是斯兰氏切迹撕裂、变形或侵入轴突。轴突或髓鞘虽可各呈单一的退变,但以两者并存多见。伤后1d即出现富含微管的再生芽,它被增殖的许旺细胞突起及其基底膜包绕,并逐步发育成熟。根据再生的特征性构筑变化,提出了再生芽、无髓和有髓纤维、斯兰氏切迹、朗氏结与神经小束的初见、发育和成熟高峰期的时间顺序。无髓纤维的发育成熟早于有髓纤维。     

灵长类动物胫神经和腓总神经损伤再生规律的研究

目的:研究灵长类动物胫神经和腓总神经再生能力差异。方法:健康成年恒河猴16只,分为A、B两组,每组8只,使用刀片切割完全损伤胫神经和腓总神经,后立即予神经外膜缝合,在术后3周、8周分别取A、B组胫神经和腓总神经吻合口远、近端神经组织行Luxol Fast Blue染色,观察胫神经和腓总神经远端、近端轴突数目,计算轴突密度,远端轴突密度/近端轴突密度为神经再生通过率。结果:术后3周和8周时,胫神经和腓总神经相比,胫神经在远端轴突密度、神经通过率等指标上,胫神经愈后优于腓总神经(P0.05)。结论:坐骨神经神经损伤修复后,胫神经轴突通过吻合口的通过率较腓总神经高,吻合口远端有更多的神经轴突,其靶器官有更多的神经纤维支配,这是导致坐骨神经损伤修复后胫神经功能恢复较腓总神经功能恢复好的重要原因之一。     

周围神经再生过程中靶器官的诱向性作用机制

以细胞培养技术与自然凝胶电泳系统方法证明,在周围神经再生过程中,损伤的坐骨神经远侧端,即起衍生的靶器官诱导神经突起的定向生长。分析探讨与神经诱向性再生相关的活性因子,其结果提示在远侧端神经组织中出现的90kDa蛋白组分具有很强的诱向性作用,诱导神经突起在神经再生过程中能够准确地到达靶器官。由此说明雪旺细胞在神经再生过程中扮演着重要角色。     

Changes in Rapid Transport of Phospholipids in the Rat Sciatic Nerve During Axonal Regeneration

Abstract: Axonal transport of phospholipids in normal and regenerating sciatic nerve of the rat was studied. At various intervals after axotomy of the right sciatic nerve in the midthigh region and subsequent perineurial sutures of the transected fascicles, a mixture of 60 μCi [Me-^HC]choline and 15 μCi [2-³H]glycerol in the region of the spinal motor neurons of the L5 and L6 segments was injected bilaterally. The amount of radioactive lipid (and in certain cases its distribution in various lipid classes) along the nerve was determined as a function of time. Three days after fascicular suture and 6 h after spinal cord injection of precursors, there was an accumulation of labeled phospholipids and sphingolipids in the transected sciatic nerve in the region immediately proximal to the site of suture. Nine days after, there was a marked increase in the accumulation of radioactivity in the distal segments of the injured nerve, which increased up to 14 days after cutting and disappeared as regeneration proceeded (21–45 days). In all segments of both normal and regenerating nerve fibers, as well as in L5 and L6 spinal cord segments, only phosphatidylcholine and sphingomyelin were labeled with [¹⁴C]choline. These results suggest that the regeneration process in a distal segment of a peripheral neuron, following cutting and fascicular repairing by surgical sutures, is sustained in the first 3 weeks by changes in the amount of phospholipids rapidly transported along the axon towards the site of nerve fiber outgrowth.     

碱性成纤维细胞生长因子对大鼠晚期周围神经再生作用的实验研究

目的探讨外源性碱性成纤维细胞生长因子(bFGF)对晚期周围神经再生的作用.方法50只SD大鼠随机分治疗组、对照组各25只,切断右侧坐骨神经,12周后予以修复,修复术后每日分别给予bFGF和生理盐水,行神经电生理和组织学检查.结果治疗组和对照组修复处远段神经均有不同程度再生,4周时已可见到再生轴突,且治疗组多见.计量分析治疗组运动神经传导速度、神经肌肉动作电位幅值、髓鞘厚度、再生轴突直径和截面积明显优于对照组.治疗组与对照组相比,差异有显著性.结论bFGF能促进晚期周围神经再生.     

Early structural changes in the axoplasmic cytoskeleton after axotomy studied by cryofixation

Summary Alterations in the cytoskeleton were studied in the axoplasm of neurites at the tips of proximal stumps of transected chicken sciatic nerves. The studies were carried out using cryofixation with a nitrogen-cooled propane jet. The most immediate effect is the almost complete disassembly of axoplasmic microtubules. This consequently causes the axonal transport of membrane-bounded organelles to cease and results in an accumulation of mitochondria and vesicles of the smooth endoplasmic reticulum. The neurofilament network is partially disorganized. Neurofilaments become shorter and fragmented, and are linked by a large number of anastomosed cross-linkers. The neurofilaments become newly aligned to the axis of the axoplasm and are of normal length 48–72 h after the transsection. At this stage the newly formed neurofilament bundles are in close proximity to the anastomosed cisternae and profiles of the smooth endoplasmic reticulum. The axonal sprouts always show a normally organized cytoskeletal network. These studies support the idea that the rapid remodelling of the neurofilament network is apparently a local event, not dependent on the slow transport of cytoskeletal materials to the tip of the proximal stump. The repair of the degraded cytoskeleton may be in accordance with the function of the endoplasmic reticulum as Ca²⁺-sequestering membrane system, which may be involved in restoring the physiological conditions of the axoplasm.     

Fish Apolipoprotein-A-I Has Heparin Binding Activity: Implication for Nerve Regeneration

This study provides evidence that apolipoprotein-A-I (apo-A-I), derived from fish plasma and nerve, has heparin binding activity. We have shown previously that injury in a regenerative CNS, such as that of fish optic nerves, leads to increased levels of apo-A-I in media conditioned by these nerves, as compared with media conditioned by noninjured nerves. In the present study, we have purified and characterized apo-A-I from both fish plasma and optic nerves. Sequence analysis of the 15 N-terminal amino acids revealed that at least 14 amino acids are identical in these two purified apo-A-I samples. The purified apo-A-I derived from both fish plasma and optic nerves binds to heparin. Binding measurements using [3H]heparin followed by Scatchard analysis revealed that apo-A-I binds to heparin with relatively low affinity (KD = 2.8 x 10(-6) M). Results are discussed with respect to the possibility that accumulation of apo-A-I in the extracellular matrix of fish optic nerves is made possible via heparin binding, like that to apolipoprotein-E in mammals.     

[PEAD:肝素:NGF]生物材料促进大鼠坐骨神经损伤恢复

目的:探讨新型材料poly(ethylene argininylaspartate diglyceride)(PEAD)结合肝素包裹神经生长因子组成的三元复合体比单纯运用NGF治疗大鼠坐骨神经损伤效果明显,为临床治疗外周神经损伤提供实验依据。方法:24只200g左右Wistar大鼠,分成生理盐水组,NGF组,NGF凝聚体三组,每组各8只,距梨状肌下缘远侧约1.5cm处运用静脉夹夹紧坐骨神经2min,采用无创细线(5/0)缝合肌肉和皮肤,并用碘伏进行消毒,NGF组每天沿坐骨切迹肌注80ngNGF,持续30天;NGF凝聚体组仅在造模时肌注复合体(内含2.4μg的NGF);生理盐水组给予等体积的生理盐水。术后每周运用脚步印迹法评价动物的行为学,并于30天后灌流、收集各组损伤侧坐骨神经,运用HE染色及投射电镜观察坐骨神经结构恢复情况,免疫荧光标记MBP,观察其蛋白的表达。结果:NGF组,NGF凝聚体组在行为学、病理结构及蛋白的表达远高于生理盐水组,并且NGF凝聚组的治疗效果优于NGF组。结论:新型凝聚体包载NGF具有明显的促进周围神经损伤后的修复与再生作用,能够在一定程度上提高单纯运用NGF治疗大鼠坐骨神经损伤的不足,达到更加理想和显著的促恢复效果。     

补阳还五汤口服和药浴对损伤的大鼠坐骨神经传导速度的影响

目的探讨补阳还五汤口服加药浴对坐骨神经传导速度的影响。方法60只SD大鼠暴露左侧坐骨神经。对照组只钳夹;实验组钳夹并加用补阳还五汤口服及药浴治疗。观察钳夹前和钳夹切除后大鼠坐骨神经传导速度（SNCV）。结果于2、4、6周分别测对照组、实验组的坐骨神经传导速度（SNCV）。各时间段实验组坐骨神经传导速度恢复快于对照组,P〈0．01。结论补阳还五汤口服加药浴对坐骨神经传导速度有明显的促进作用。     

In Vivo Phosphorylation of Axonal Proteins in Goldfish Optic Nerve During Regeneration

In vivo phosphorylation of axonal proteins was investigated in normal and regenerating optic nerves of goldfish by two-dimensional gel electrophoresis. By 6-24 h after intraocular injection of H3(32)PO4, approximately 20 optic nerve proteins ranging in size from 19 to 180 kilodaltons and in pI from 4.4 to 6.8 were seen to have incorporated radiolabel. Five of these proteins showed a robust increase in incorporation of phosphate during regeneration. Among the latter was an acidic (pI 4.5) 45-kilodalton protein, which has previously been shown to be conveyed by fast axonal transport and to increase dramatically in its rate of synthesis during regeneration of goldfish optic axons.