聚乳酸/神经生长因子缓释导管修复周围神经缺损实验研究

目的：观察聚乳酸／神经生长因子(PDLLA／NGF)缓释导管对大鼠坐骨神经缺损再生的促进作用及其性能、降解过程。方法：SD大鼠60只，随机分为4组：自体神经移植组(A组)，单纯PDLLA导管组(B组)，单纯导管加导管内一次性给药组(C组)，缓释导管组(D组)，每组15只。制作坐骨神经10mm缺损模型，分别用自体神经移植、单纯PDLLA导管、PDLLA／NGF缓释导管桥接修复。于第1、2、3月后行大体观察、三头肌湿重恢复率测量、电生理检测、组织学和图象分析对比。结果：同时间段组间比较，除三头肌恢复率低以外，D组再生神经取得了和A组相似的效果，明显比B组和C组恢复好。结论：PDLLA／NGF缓释导管桥接修复大鼠坐骨神经缺损，能够有效促进神经再生，效果接近自体神经移植。     

复合FK506/NGF/RGD缓释膜应用于同种异体神经移植的实验研究

目的:探讨复合FK506/NGF/RGD缓释膜对冷冻处理下的同种异体神经的移植效果.方法:选择SD大鼠作为动物模型.在建市动物模型3周前取8只SD大鼠作为供体,切取双侧坐骨神经,冷藏于-196℃液氮中.另取40只SD大鼠随机分为:A单纯冷冻异体神经移植组;B冷冻异体神经联合应用FK506/RGD缓释膜移植组;C自体神经移植组;D冷冻异体神经联合应用FK506/NGF/RGD缓释膜移植组.术后12周进行大体观察、电生理、小腿三头肌恢复率、组织学、超微结构等测定.结果:术后12周内,4组大鼠均未发生急性移植排斥反应.D组运动神经传导速度和肌肉湿重恢复率明显优于A、B、C组(P<0.05).组织学测定发现D组移植段神经纤维形态最为饱满,神经纤维直径大,髓鞘厚,成熟度高.结论:复合FK506/NGF/RGD缓释膜能更有效地促进神经生长.     

bFGF复合膜对自体神经移植的影响

目的：探讨ｂＦＧＦ复合膜对自体神经移植的影响。方法：在自体神经缝接缺损神经处,分别置入含ｂＦＧＦ复合剂的非降解膜和仅含ｂＦＧＦ复合剂的非降解膜和仅含ｂＦＧＦ的非降解腊,并通过组织学和图像分析等方法对兔的正中神经缺损后的再生情况进行比较观察。结果：①实验侧再生神经纤维的数量多于对照侧且有显著性差异。②实验侧再生的有髓神经纤维的平均直径大于对照侧,但没有显著性差异。③再生神经纤维的髓鞘较薄、着色较淡,     

不同方法处理异种神经移植后再生神经纤维的比较研究

探讨没方法先期处理对异种神经移植后神经再的影响。方法将兔的胫神经，分别经反复冻融，或＾６０Ｃｏ辐射，或刀豆球蛋白Ａ灌注先期处理后移植至大的从地骨神经；或末经径处理移植后服用雷公藤甲素。     

外消旋聚乳酸/神经生长因子缓释导管的体内降解研究

目的：研究外消旋聚乳酸／神经生长因子缓释导管（PDLLA／NGF）的降解性能，为修复神经缺损提供依据。方法：制作PDLLA／NGF缓释导管并用于桥接大鼠坐骨神经缺损，术后1、2、3月分别行导管的大体观察和电镜检查。结果：术后大体观察及电镜检查发现，PDLLA／NGF缓释导管能在大鼠体内随时间延长逐步降解，内层降解比外层速度快，到3月时导管外形仍保持完整，再生神经已顺利通过导管腔，无压迫及疤痕形成。结论：PDLLA／NGF导管在大鼠体内降解时间超过3月，能提供缺损神经再生所需的时间，为神经再生提供良好的微环境。     

补阳还五汤和NGF对异种去细胞神经支架移植后神经再生和修复的影响

为了探讨补阳还五汤对异种去细胞神经支架移植后神经再生及功能恢复的影响以及与NGF的效果比较。我们将36只健康成年SD大鼠随机分为3组:补阳还五汤(BuYangHuanWu Decotion,BYHWD)治疗组、NGF治疗组和生理盐水(NS)对照组。动物分别存活4周和8周,不同时段各组为6只。以上3组大鼠分别以10mm长度的兔异种去细胞神经支架桥接大鼠坐骨神经缺损。术后补阳还五汤组和生理盐水对照组分别用补阳还五汤水煎剂和生理盐水按10ml/kg灌胃,持续1月。各组分别在术后第4周、8周行大体观察、坐骨神经功能指数(SFI)测定、辣根过氧化物酶逆行示踪、小腿三头肌湿重恢复率测定、移植体的组织学观察。结果显示,在同一时相点补阳还五汤组SFI、术侧L3～L5脊神经节和L3～L5脊髓节段前角运动细胞的HRP标记细胞数均优于生理盐水对照组(P<0.05)。补阳还五汤组与NGF组比较没有显著性差异(P>0.05)。在同一时相点形态学观察结果:各组神经移植体粗细基本正常,表面大量血管分布,与周围组织轻度粘连;补阳还五汤组再生神经纤维更加密集、排列规则整齐,移植体雪旺细胞(SC)大量增殖,接近于正常。上述结果提示,应用补阳还五汤可促进异种去细胞神经支架移植后神经再生与功能恢复,与NGF的作用效果没有差别。     

NGF和GM1联合应用对去细胞异种神经支架移植后神经再生和修复的影响

为了探讨神经生长因子(NGF)和单唾液酸四己糖神经节苷脂(GM1)联合应用对去细胞异种神经支架移植后神经再生及功能恢复的影响,本研究将SD大鼠随机分为生理盐水对照组、NGF治疗组、GM1治疗组、NGF+GM1联合治疗组,选取兔胫神经进行化学萃取,形成去细胞异种神经支架桥接大鼠10mm坐骨神经缺损,移植前分别用等渗盐水(NS)、NGF液、GM1液或NGF+GM1液浸泡去细胞异种神经支架,术后各组大鼠术侧小腿肌内分别注射NS液、NGF液、GM1液或NGF+GM1液。术后4、8周行大体观察并用神经电生理、肌湿重、免疫组织化学等方法测定神经纤维再生及功能恢复情况。结果显示:在同一时间点,NGF+GM1联合治疗组坐骨神经运动传导速度恢复率、小腿腓肠肌复合动作电位波幅恢复率、小腿三头肌湿重恢复率均优于单独用药组(P<0.05);免疫组织化学结果显示NGF+GM1联合治疗组有大量再生有髓神经纤维顺畅地通过远端吻合口。本研究结果提示联合应用NGF和GM1可明显促进去细胞异种神经支架移植后的神经纤维再生与功能恢复。     

血浆冷存的异体神经移植对神经再生影响的实验研究

目的 用经动物血浆处理后冷存的异体神经移植体,缝接神经缺损,提高神经再生。方法 用15只兔左、右肢正中神经30条,分别切除2.5 cm为实验材料,动物分两组:A组15条神经用经受体动物血浆浸泡冷冻处理的异体神经移植为实验组,B组15条神经只经冷冻处理的异体神经缝接,术后不同时间,采用光、电镜组织学,图像分析仪测定,神经外周粘连定量测定和电生理指标等观察。结果(1)A组再生神经纤维在数量上和直径上均明显较B组的多而粗(p<0.03);(2)A组神经缝合段的结缔组织增生和与其外周组织粘连程度均较B组轻(p<0.01)。结论 经受体血浆处理后冷存的异体神经移植体,有提高神经再生的作用。     

胎儿神经加自体雪旺氏细胞桥接周围神经缺损的实验研究

目的 探讨自体雪旺氏细胞植入胎儿神经后修复周围神经缺损的效果,寻找替代自体神经移植的材料。方法 用Wistar大鼠80只切断左侧大腿坐骨神经,造成15mm缺损,分别用自体神经(A组),液氮冷冻胎儿神经(B组),液氮冷冻胎儿神经加自体雪旺氏细胞粗制品(C组)进行桥接。于术后4、12、24周取桥体、桥体远端坐骨神经,分别行电镜、光镜观察、图像分析和电生理检测,所得数据经单因素方差分析和q检验。结果 A组和C组间有髓纤维数目、无髓纤维数目、复合动作电位峰值恢复率、传导速度恢复率均无显著差异,两组神经再生效果相近,而B组的再生效果则不及A、C两组;B组与A组,B组与C组分别作两两比较,部分指标存在显著差异(P<0.01)。结论 作者认为植入自体雪旺氏细胞的胎儿神经桥接周围神经缺损优于单纯胎儿神经桥接,是一种良好的替代自体神经的桥接物,有进一步研究价值和临床应用前景。     

Promoting peripheral nerve regeneration with biodegradable poly (DL-lactic acid) films

Regeneration and repair of peripheral nerve injury has always been a major problem in the clinic. The conventional technique based on suturing the nerve ends to each other coupled with the implantation of nerve conduits outside is associated with postoperative adhesions and scar problems. Recently, a novel biodegradable poly (DL-lactic acid) (PDLLA) film has been introduced. This novel anti-adhesion film has a porous structure with better mechanical properties, better flexibility, and more controllable degradation as compared to traditional non-porous nerve conduits. However, little is known about the effects of such PDLLA films on regeneration and repair of peripheral nerve injury in vivo. In this study, we evaluated the effects of PDLLA films implantation after sciatic nerve transection and anastomosis on subsequent sciatic nerve regeneration in vivo, using a rat sciatic nerve injury model. Sciatic nerve transection surgery coupled with direct suturing only, suturing and wrapping with traditional nerve conduits, or suturing and wrapping with PDLLA films was performed on adult Wistar rats. The additional wrapping with PDLLA films inhibited the nerve adhesion after 12 weeks recovery from surgery. It also increased the compound muscle action potentials and tibialis and gastrocnemius muscle wet weight ratio following 8 weeks recovery from surgery. Regenerated nerve fibers were relatively straight and the aligned structure was complete in rats with implantations of PDLLA films. The results suggested that PDLLA films can improve the nutritional status in the muscles innervated by the damaged nerves and promote nerve regeneration in vivo.     

片仔癀对局灶性脑梗死大鼠神经生长因子表达的影响

目的:探讨片仔癀对局灶性脑梗死大鼠NGF表达的影响及其作用机制。方法:制备MCAO大鼠模型,将SD大鼠70只随机分为7组:假手术组,模型组,片仔癀大、中、小剂量组,尼莫地平组和脑得生组,造模3 d后取材,采用免疫组织化学法观察各组大鼠大脑皮层NGF的表达情况。结果:与假手术组比较,各造模组大鼠大脑皮层NGF的表达均显著升高(P<0.01);与模型组比较,各给药组大鼠大脑皮层NGF的表达也均显著升高(P<0.01);与尼莫地平组和脑得生组比较,片仔癀各剂量组大鼠大脑皮层NGF的表达有所升高(P<0.05);结论:片仔癀可促进局灶性脑梗死大鼠大脑皮层NGF的表达,从而起到脑保护的作用。     

Developmental changes in nerve growth factor (NGF) binding and NGF receptor proteins trkA and p75 in the facial nerve

This study characterizes the temporal-spatial distribution of nerve growth factor (NGF) low (p75) and high-affinity (trkA) receptors in the facial nerve and geniculate ganglion (GG) of developing quail embryos (E-3 to E-14). We used ¹²⁵I-labeled NGF (¹²⁵I-NGF) to study binding dynamics in a temporal series of isolated primordia and an autoradiographic series of staged specimens to characterize the occurrence and distribution of NGF receptors in this cranial nerve and its ganglion. In addition, expression of trkA and p75 protein-like immunoreactivity in the facial nerve and GG was studied by Western blot, in order to distinguish between high- and low-affinity NGF receptors respectively. The quantitative study of binding show that isolated facial primordia ranging from E-3 to E-14 exhibit different levels of specific binding. High initial binding levels were observed on E-3 specimens, then an initial decrease on day 4 (E-4) followed by a steady increase from days E-4 to E-7. Maximum ¹²⁵I-NGF binding was achieved on E-7, followed by a steady decline in binding on days 8 (E-8) and 9 (E-9), reaching near background levels on day 10 (E-10) of development and until the oldest stage assayed (E-14). Most of the cells bearing NGF receptors appeared to be nonneuronal crest-derived cells, but some placode-derived neurons and motor fibers of the VIIth cranial nerve transiently expressed the ability to bind ¹²⁵I-NGF. The temporal pattern of p75 expression matches the pattern of quantitative binding of NGF, while the trkA expression is restricted to a few stages mainly E7 and E9, implying that most of the binding detected is via low-affinity receptors, except for a proportion of high-affinity receptors present at stages of maximum binding. This temporal pattern of NGF binding sites suggests that cells within the VIIth cranial nerve are responsive to and/or dependent upon NGF in vivo, so NGF may play a biological role during normal development of the facial nerve. In view of the developmental events that parallel the occurrence and type of NGF binding sites, we suggest that this role may be to modulate from earlier chemotaxis and cell proliferation to much later events, such as neuronal differentiation and neuron-glia interactions. The significance of these findings in regeneration during adult life remain to be investigated.     

NGF对兔脑缺血／再灌注损伤半胱氨酸蛋白酶-12的影响

目的探讨神经生长因子(NGF)对兔脑缺血/再灌注损伤半胱氨酸蛋白酶-12(caspase-12)的影响。方法健康雄性新西兰大白兔26只,随机分成假手术组(Sham组,n=6)、缺血/再灌注组(I/R组,n=10)和NGF治疗组(NGF组,n=10)。免疫组化方法检测组织caspase-12及caspase-3的表达;用流式细胞术(FCM)及DNA原位末端缺口标记法(TUNEL法)检测神经细胞凋亡。结果与Sham组相比,I/R组caspase-12及caspase-3表达增加(P<0.01),凋亡细胞数增加(P<0.01);而NGF处理后显著缓解caspase-12与caspase-3的表达增加及凋亡细胞数的增加(P<0.01),但仍高于Sham组。结论抑制caspase-12介导的caspase级联反应性凋亡途径的激活是NGF减少缺血/再灌注损伤细胞凋亡的机制之一。     

NGF对局灶性脑缺血再灌注大鼠海马CREB表达的上调作用

目的探讨外源性神经生长因子(NGF)对局灶性脑缺血再灌注大鼠海马CREB和磷酸化的CREB(p-CREB)表达的影响。方法用线栓法阻塞大鼠大脑中动脉制作局灶性脑缺血再灌注模型,应用免疫组织化学、W estern B loting和图像分析方法检测大鼠海马CA1区CREB和磷酸化CREB表达。结果假手术组海马CA1区CREB有明显表达,缺血再灌注组CA1区表达较假手术组减少,NGF组CA1区的CREB表达多于缺血再灌注组(P<0.05)。假手术组海马CA1区p-CREB表达很少,缺血再灌注组p-CREB表达多于假手术组,NGF组p-CREB表达多于缺血再灌注组(P<0.05)。结论NGF上调海马CREB和p-CREB的表达,对缺血神经元起保护作用,CREB和p-CREB参与NGF对缺血神经元的保护作用机制。     

高亲和性神经生长因子受体对神经生长因子诱导神经母细胞瘤 …

观察人神经母细胞瘤细胞系ＩＭＲ－３２细胞在恢复高亲和性神经生长因子受体基因ｔｒｋＡ表达后对神经生长因子（ＮＧＦ）诱导分化的反应。方法应用基因重组技术构建含外源ｔｒｋＡ ｃＤＮＡ的逆转录病毒载体,经ＰＡ３１７细胞包装后感染靶细胞系ＩＭＲ－３２细胞,经Ｓｏｕｔｈｅｒｎ ｂｌｏｔ杂交,逆转录－聚合酶链反应（ＲＴ－ＰＣＲ）证实转化细胞系中含有外源基因的整合及稳定表达后,进行神经生长因子诱导分化实验。结果转     

The effect of alpha-latrotoxin on the neurosecretory PC12 cells differentiated by treatment with nerve growth factor

Neurosecretory PC12 cells differentiated in vitro by prolonged (at least 2 weeks) treatment with nerve growth factor were exposed to alpha-latrotoxin and studied by morphological and biochemical techniques. Cell monolayers or suspensions responded to the toxin with a prompt and massive release of neurotransmitter. The dose dependency (Km approximately 5 X 10(-10)M) and the maximum release effect (approximately 60% of the stored [3H]dopamine released within 8 min) were not appreciably different from the values found in non-differentiated PC12 cells. Moreover, the concentration dependency of the release was found to correspond closely to that of the [125I]alpha-latrotoxin binding to its specific sites (the alpha-latrotoxin receptors). The number of these receptors was over two-fold higher in differentiated than in undifferentiated cells. Since, however, differentiation implies a large increase in cell size and surface area, the receptor density (number/unit area) remained virtually unchanged. By radioautography the alpha-latrotoxin receptors were found to remain diffusely distributed at the entire surface of differentiated cells even when these were allowed to form synapses with myotubes. This situation is at variance with that demonstrated recently at the frog neuromuscular junction, where alpha-latrotoxin receptors are exclusively localized at the nerve terminal plasmalemma. Scanning and transmission electron microscopy revealed that the enlargements of neurites where dense granules are preferentially accumulated--the varicosities and terminals--underwent swelling and extensive disorganization within a few minutes after the application of alpha-latrotoxin, whereas the cell bodies and the tracts of neurites occupied primarily by microtubules were less severely affected. The greater sensitivity of varicosities and terminals with respect to the other parts of the differentiated cells, rather than the consequence of a specific addressing of the toxin to these structures, might be due to their vulnerability by toxin-induced events, such as the uncontrolled activation of ion fluxes.     

神经生长液对兔周围神经损伤的修复作用研究

目的:研究神经生长液(nerve growth decoction,NGD)促进兔周围神经损伤的修复作用。方法:大耳白兔40只,雌雄各半,行腓总神经夹伤术。实验分为五组:NGD低、中、高剂量组,弥可保组(阳性对照),空白对照组。口服给药四周后行电生理、组织学检测和超微结构观察。结果:NGD高、中剂量组和弥可保组在腓总神经传导速度恢复率,脊髓前角运动神经元的存活率及再生髓鞘计数均明显好于空白对照组(P<0.01)。超微结构观察给药组有髓神经纤维的髓鞘形态、厚度、成熟度均优于对照组,变性纤维的数目少于对照组。结论:神经生长液能促进损伤周围神经的再生和功能的恢复。