哺乳动物视网膜节细胞中神经营养因子的主要生物学作用

哺乳动物视网膜节细胞的存活和再生受许多神经营养因子的调节。本文综述了神经营养素家族、细胞因子家族及玻璃体内移植外周神经节段等因素促进视网膜节细胞存活的作用 ;同时也分析了它们对视网膜节细胞的轴突生长和再生的影响 ,并提出这些神经营养因子可能通过升高cAMP水平起作用。     

睫状神经营养因子的基因结构及受体

睫状神经营养因子最初由于能促进鸡胚胎睫状神经节副交感神经元存活而被发现,现已知道它对神经系统的细胞具有更广泛的作用。最近研究表明睫状神经营养因子与成血细胞因子的一个亚家族成员在结构上相似并共用受体成分。     

神经营养因子与周围神经再生

人们对神经营养因子在促进周围神经元的存活 ,维持周围神经元的数目方面的作用进行了大量研究。神经损伤后 ,给予外源性神经营养因子 ,能够产生与靶器官源性的神经营养因子相似的作用。周围神经再生过程中 ,神经营养因子NGF、NT 3和BDNF对背根节初级感觉神经元和脊髓运动神经元的存活及其表型的表达具有明确的选择性作用。其他神经营养因子 ,如CNTF、GDNF、LIF、GGF等 ,对神经元细胞也具有不同作用。本文将就神经营养因子对治疗周围神经损伤及周围神经疾病的潜在作用作一综述     

神经营养因子对视网膜缺血再灌注损伤的影响

视网膜缺血再灌注损伤是由于自由基、炎性细胞因子等细胞微环境的改变引起的病理过程,能造成视网膜及神经组织严重损伤。微环境包括对细胞产生影响的周围结构和成分,如附近的神经细胞、基质细胞和结合在胞外基质上的各种生长因子和细胞因子等。这些因子有的具有保护视网膜作用,如神经生长因子、睫状神经营养因子、脑源性神经营养因子、碱性成纤维细胞生长因子等;有的具有损伤作用,如血管内皮生长因子等。本文结合文献以这些因子在视网膜缺血再灌注损伤中的作用作一综述。     

鸡胚背根神经节感觉神经元体外培养研究

本文对背根神经节感觉神经元的体外培养条件进行了研究。结果表明利用差速贴壁法可使神经元纯度达到95％以上;在血清和适宜的细胞外基质存在条件下，神经元分散良好。本研究还通过外源性脑源性神经营养因子对神经元纤维再生的作用进一步证实了该方法的可靠性。     

重组睫状神经营养因子对周围神经再生中多种细胞的作用

为了解重组睫状神经营养因子 ( CNTF)对周围神经再生中多种细胞的作用 ,用硅管套接切断的成年大鼠坐骨神经 ,在受损神经局部一次性给予重组睫状神经营养因子 :用免疫组织化学 ABC法结合计算机图像分析观测再生神经中 GAP-4 3、S10 0、CD68、MHC- 免疫反应阳性物质的变化。与生理盐水对照组相比 ,证明 CNTF组再生神经中上述四种物质显著增多。结果提示重组睫状神经营养因子能促进轴突的再生、Schwann细胞的迁入、单核细胞的渗出和活化     

神经营养因子与周围神经再生

周围神经损伤后 ,成功的神经再生取决于很多因素。这其中神经当时所处的微环境起着很重要的作用。大量的实验证明 ,该微环境中的神经营养因子对周围神经的维持和存活有着不可缺少的作用。本文拟对神经营养因子与周围神经再生的关系作一综述。1　神经营养因子　　神经营养因子 (ＮｅｕｒｏｔｒｏｐｈｉｃＦａｃｔｏｒｓ,ＮＴＦ)是一类在靶组织内合成 ,并逆行运输至效应神经元的能对中枢和周围神经发挥营养作用的小分子肽类物质和蛋白质。神经营养因子种类繁多 ,大致可分为生长因子类 (ＧｒｏｗｔｈＦａｃｔｏｒｓ,ＧＦ)和细胞因子类 (Ｎｅｕ…     

睫状神经节神经营养因子在大鼠坐骨神经生后发育中的表达—免疫组…

用免疫组织化学ＡＢＣ技术和计算机图像处理系统研究了１ｄ龄，１月龄、２月龄和３月龄大鼠坐骨神经中睫状神经节神经营养因子样免疫反应的分布及其强度的变化。结果表明，在４个年龄组中均存在阳性免疫的。阳性免疫反应见于雪旺细胞胞浆，轴突和髓鞘呈阴性。睫状神经节神经营养因子免疫反应以１月龄鼠最高，２月龄次之，３月龄最低。本文的结果提示，睫状神经节神经营养因子在出后即存在于雪旺细胞中，其含量随周围神经的发育而有所     

胶质细胞源性神经营养因子缓释微球及NogoA,ChABC缓释微球联合应用损伤脊髓再生的形态学变化

背景：促进轴突再生的原则是改善抑制再生的环境和提高轴突生长能力,措施主要有轴突生长抑制因子阻滞剂和神经营养因子应用。用可降解微球加载药物是一种在局部提供持续药物释放的方法。 目的：探讨胶质细胞源性神经营养因子、NogoA、ChABC 缓释微球联合应用促进大鼠损伤脊髓再生病理形态学修复的作用。 方法：建立SD大鼠T10 脊髓完全横断伤模型,分别在损伤局部给予生理盐水、胶质细胞源性神经营养因子、胶质细胞源性神经营养因子缓释微球、NogoA缓释微球、ChABC 缓释微球及3种微球联合治疗,并设立未造模的正常组及假手术组。损伤后10周,每组行四甲基若丹明葡聚糖胺顺行示踪,及神经丝蛋白200、生长相关蛋白43、胶质细胞源性神经营养因子免疫组化检查,并采用免疫组化图像分析系统进行定量分析。 结果与结论：胶质细胞源性神经营养因子、NogoA、ChABC 缓释微球联合能提高脊髓损伤局部神经丝蛋白200、生长相关蛋白43、胶质纤维酸性蛋白的表达水平,显示局部脊髓再生修复加强,其效果优于单用胶质细胞源性神经营养因子缓释微球。提示,胶质细胞源性神经营养因子缓释微球及NogoA,ChABC 缓释微球联合促大鼠损伤脊髓再生修复其效果优于单用胶质细胞源性神经营养因子缓释微球。     

HSV介导的神经营养因子-3体外表达拮抗顺铂的耳毒性作用

目的　观察单纯疱疹病毒 (HSV)扩增子型表达载体介导的神经营养因子 3(NT 3)体外表达对顺伯耳毒性的拮抗作用。　方法　以HSV扩增子型表达载体为基础 ,构建由CMV作启动子 ,表达c myc标记的大鼠NT3或鼠的小肠碱性磷酸酶 (MIAP)的HSVnt3和HSVmaip扩增子型表达载体。通过无辅助病毒的包装体系包装后 ,感染体外培养的耳蜗器官 ,经不同浓度的顺铂处理 4 8h后 ,观察转导NT 3或报告基因 (MIAP)后的耳蜗螺旋神经节细胞存活和神经突起的再生。　结果　显示在病毒滴度 (MOI)为 0 2 5时 ,HSVnt3感染体外培养的耳蜗 4 8h后 ,能够刺激耳蜗分泌较高水平的NT 3(3μg L)。用不同浓度的顺铂处理 4 8h ,HSVnt3转导的耳蜗螺旋神经节细胞可以在一定范围内 ,耐受顺铂的耳毒性作用 ,与HSVmiap转导的耳蜗比较 ,神经元残留的数量明显高于对照组 (P <0 0 0 1) ,耳蜗螺旋神经节细胞神经突起的长度和密度也与对照组有明显的差别 (P <0 0 0 1)。　结论　提示无辅助病毒的HSV所介导的神经营养因子 3表达 ,能够在体外拮抗顺铂的耳毒性作用。     

Intravitreal injections of neurotrophic factors and forskolin enhance survival and axonal regeneration of axotomized beta ganglion cells in cat retina

Some retinal ganglion cells in adult cats survive axotomy for two months and regenerate their axons when a peripheral nerve is transplanted to the transected optic nerve. However, regenerated retinal ganglion cells were fewer than 4% of the total retinal ganglion cell population in the intact retina. The present study examined the effects of intravitreal injections of neurotrophic factors (brain-derived neurotrophic factor, ciliary neurotrophic factor, basic fibroblast growth factor, glial cell-derived neurotrophic factor, neurotrophin 4), first on the survival of axotomized cat retinal ganglion cells within 2 weeks, and then on axonal regeneration of the retinal ganglion cells for 2 months after peripheral nerve transplantation. We tested first enhancement of the survival by one of the factors, and then one or two of them supplemented with forskolin, which increases intracellular cAMP. Single injections of 0.5 microg or 1 microg brain-derived neurotrophic factor, 1 microg ciliary neurotrophic factor, or 1 microg glial cell-derived neurotrophic factor significantly increased total numbers of surviving retinal ganglion cells; 1.6-1.8 times those in control retinas. Identification of retinal ganglion cell types with Lucifer Yellow injections revealed that the increase of surviving beta cells was most conspicuous: 2.5-fold (brain-derived neurotrophic factor) to 3.6-fold (ciliary neurotrophic factor). A combined injection of 1 microg brain-derived neurotrophic factor, 1 microg ciliary neurotrophic factor, and 0.1 mg forskolin resulted in a 4.7-fold increase of surviving beta cells, i.e. 50% survival on day 14. On the axonal regeneration by peripheral nerve transplantation, a combined injection of brain-derived neurotrophic factor, ciliary neurotrophic factor, and forskolin resulted in a 3.4-fold increase of beta cells with regenerated axons. The increase of regenerated beta cells was mainly due to the enhancing effect of neurotrophic factors on their survival, and possibly to a change of retinal ganglion cell properties by cAMP to facilitate their axonal regeneration.     

Ciliary neurotrophic factor promotes the regrowth capacity but not the survival of intraorbitally axotomized retinal ganglion cells in adult hamsters

Ciliary neurotrophic factor has recently been shown to promote the axonal regrowth of retinal ganglion cells into a peripheral nerve graft following an intracranial transection of the optic nerve (approximately 7 mm from the optic disc). It is unclear whether the enhancement of axonal regrowth by ciliary neurotrophic factor application correlates with the enhancement of survival of retinal ganglion cells and/or the up-regulation of expression of growth-associated protein-43 messenger RNA in retinas. The present study evaluated the regenerative capacity of retinal ganglion cells following intraorbital transection of the optic nerve (approximately 1.5 mm from the optic disc) and the attachment of a peripheral nerve to the ocular stump of the optic nerve. In addition, we have determined the survival of retinal ganglion cells and the expression of growth-associated protein-43 messenger RNA in ciliary neurotrophic factor-treated retinas following optic nerve transection. The results showed that in the ciliary neurotrophic factor-treated retinas, the number of retinal ganglion cells which had regrown axons into a peripheral nerve is about four times more than the control. In the axotomized retinas, ciliary neurotrophic factor initiated sprouting of axon-like processes at 14 and 28 days post-axotomy and up-regulated the expression level of growth-associated protein-43 messenger RNA at 7, 14 and 28 days post-axotomy. However, ciliary neurotrophic factor did not prevent the death of axotomized retinal ganglion cells. We suggest that one possible mechanism for the axonal regeneration of axotomized retinal ganglion cells by ciliary neurotrophic factor could be mediated by the up-regulation of growth-associated protein-43 gene expression and not by increasing the pool of surviving retinal ganglion cells after axotomy.     

Ciliary neurotrophic factor is an axogenesis factor for retinal ganglion cells

Although mature mammalian retinal ganglion cells normally fail to regrow injured axons, exposure to the molecular environment of the peripheral nervous system stimulates regenerative growth. The present study used dissociated rat retinal ganglion cells purified by immunopanning to identify peripheral nervous system-derived factors that promote axonal outgrowth. Of the multiple growth factors investigated, only ciliary neurotrophic factor and the related cytokine, leukemia inhibitory factor, had striking neuritogenic activity, with half-maximal effects at 1-2 ng/ml. Brain-derived neurotrophic factor stimulated retinal ganglion cell survival nearly as well as ciliary neurotrophic factor, but had only minor effects on outgrowth. Thus, the neuritogenic effects of ciliary neurotrophic factor are not a simple consequence of increased survival. Ciliary neurotrophic factor-stimulated outgrowth was correlated with increased expression of the growth-associated membrane phosphoprotein, GAP-43, a hallmark of optic nerve regeneration in vivo. A high molecular weight fraction from media conditioned by rat optic or sciatic nerve mimicked the effect of ciliary neurotrophic factor in inducing axonal outgrowth. Ciliary neurotrophic factor was detected in the conditioned media on western blots, and the biological activity of the conditioned media was neutralized with an anti-ciliary neurotrophic factor antibody. These results indicate that ciliary neurotrophic factor has specific effects on axon outgrowth in retinal ganglion cells that are dissociable from its effects on cell survival, and that ciliary neurotrophic factor accounts for most of the axon-promoting activity for retinal ganglion cells present in either the sciatic or optic nerve.     

视网膜干细胞移植对青光眼视网膜神经节细胞的保护

BACKGROUND:Stem cell transplantation is a new method for blinding eye disease. But there is a lack of research about the protective effect of retinal stem cell transplantation on retinal ganglion cells in glaucoma. OBJECTIVE:To explore the protective effect of retinal stem cell transplantation on retinal ganglion cells of rats with glaucoma. METHODS:Forty-five Sprague-Dawley rats were randomly divided into three groups (n=15 per group) including control, model and retinal stem cell transplantation groups. Rat models of glaucoma were prepared in the latter two groups, and at 7 days after modeling, rats in the three groups were given intravitreal injection of 1 mL retinal stem cells (5x10⁶ cells), the same amount of PBS, and no treatment, respectively. Subsequently, relative indicators were detected at 2 weeks after transplantation. RESULTS AND CONCLUSION:The expressions of brain-derived neurotrophic factor and insulin-like growth factor I protein as well as the number of retinal ganglion cells were the highest in the control group, followed by the retinal stem cell transplantation group model group, and the lowest in the model group (P < 0.05). The number of apoptotic retinal ganglion cells in model group was significantly higher than that of control group (P < 0.05), and which in the retinal stem cell transplantation group was significantly lower than that in the model group (P < 0.05), but higher than that in the control group (P < 0.05). These results suggest that retinal stem cell transplantation for rat glaucoma can exert a protective effect on retinal ganglion cells.     

Administration of brain-derived neurotrophic factor suppresses the expression of heat shock protein 27 in rat retinal ganglion cells following axotomy

Optic nerve transection results in the apoptotic cell death of the majority of retinal ganglion cells by 14 days. The neurotrophin brain-derived neurotrophic factor (BDNF) enhances survival of retinal ganglion cells. In addition, the small heat shock protein Hsp27, with its anti-apoptotic effects, may be important for neuron survival following axotomy or trophic factor withdrawal. We recently reported the induction and expression of Hsp27 in a subset of retinal ganglion cells following axotomy. Here we have examined the effect of BDNF administration on the expression of Hsp27 in axotomized adult rodent retinal ganglion cells. Retinal ganglion cells were pre-labeled with Fluorogold prior to optic nerve transection and concomitant intraocular injection of BDNF or vehicle. Hsp27 immunofluorescence was examined in retinal sections from 4 to 28 days following injury. Consistent with previous survival studies, the number of Fluorogold-labeled retinal ganglion cells declined from 100% at 4 days to approximately 15% by 14 days following axotomy and vehicle injection. In contrast, with BDNF administration, retinal ganglion cell survival was maintained at 100% to 7 days following axotomy. We report that the number of Hsp27-positive injured retinal ganglion cells, as detected by immunohistochemical staining, was decreased by 50% in BDNF-treated retinas, when compared with vehicle-treated controls. This decreased expression of Hsp27 in response to BDNF treatment was seen both at early (4 days) and delayed (14 days) times. BDNF following optic nerve transection significantly reduced the expression of Hsp27 in retinal ganglion cells. These results indicate that BDNF may down-regulate alternate cell survival pathways, including the stress-induced expression of Hsp27, and may help to explain the failure of chronic neurotrophin treatment to maintain long-term retinal ganglion cell survival.     

Axonal regeneration of cat retinal ganglion cells is promoted by nipradilol, an anti-glaucoma drug

Neurons in the CNS can regenerate their axons in an environment of the peripheral nervous system, but this ability is limited. Here we show that an anti-glaucoma drug, nipradilol, at low concentration led to a four-fold increase in the number of cat retinal ganglion cells regenerating their axons into a transplanted peripheral nerve 4 and 6 weeks after axotomy. Nipradilol also increased the number of three main regenerating retinal ganglion cell types (alpha, beta, not alpha/beta), and enhanced the rate of axonal regeneration of these retinal ganglion cells. Nipradilol is a donor of nitric oxide and an antagonist of alpha-1, beta-1 and -2 adrenoreceptors, and we therefore examined whether one of these pharmacological effects might be more important in promoting axon regeneration. A nitric oxide donor increased the number of regenerating retinal ganglion cells, but not the rate of axonal regeneration. Denitro-nipradilol (nitric oxide-deprived nipradilol) or a nitric oxide scavenger injected before nipradilol increased the number of regenerating retinal ganglion cells but did not promote regeneration rate. Blockade of individual alpha- and beta-adrenoreceptors did not increase the number of regenerating retinal ganglion cells or the rate of regeneration. From these results, it is suggested that nitric oxide plays a crucial role in mediating the effects of nipradilol on axon regeneration and neuroprotection, and the metabolite of nipradilol supports the effects.     

Fibroblast growth factors promote the survival of adult rat retinal ganglion cells after transection of the optic nerve

Basic and acidic fibroblast growth factors (FGF) were implanted next to the proximal stump of the transected optic nerve of adult rats, in order to assess whether these molecules have neurotrophic activity in vivo. Of the 119,973 +/- 2484 (S.E.M.) retinal ganglion cells present in retinae of unoperated control rats, 11,375 +/- 2413 (S.E.M.) remained at 30 days after transection of the optic nerve in control operated rats. After implantation of gel foam soaked in basic FGF, the number of retinal ganglion cells surviving at 30 days after axotomy tripled (36,387 +/- 3270 (S.E.M.], after acidic FGF, it increased almost 4-fold (40,916 +/- 5405 (S.E.M.]. These results indicate that FGF has neurotrophic activity in the adult central nervous system, and that this molecule is able to rescue adult retinal ganglion cells from axotomy induced cell death. It remains to be shown whether FGF acts directly on retinal ganglion cells or indirectly via glial cells or other cells.     

Roller Organotypic Cultures of Postnatal Rat Retina

Floating retinal sections from 7-12-day-old rats form ball-shaped retinal bodies during roller culturing. Histological studies of serial sections of retinal bodies showed that their outer surface is formed by the retina completely retaining organotypic cytoarchitectonics. Some retinal bodies have laminar structure consisting of several layers of the retina. At the initial stages of culturing some retinal bodies contain a cavity, which later is completely obliterated due to the growth of axons of ganglion cells and migration of glial cells and fibroblasts. This study demonstrated the possibility of long-term survival, differentiation, and in vitro axonal regeneration of ganglion cells, the main retinal efferent neurons, which can provide the basis for investigation of pathology and drug correction of injuries and stimulation of regeneration of these cells in experimental glaucoma models.     

Neuronal susceptibility to damage: comparison of the retinas of young, old and old/caloric restricted rats before and after transient ischemia

Compared to young rats, old age increases susceptibility and caloric restriction decreases susceptibility for the loss of retinal ganglion cells and displaced amacrine cells following retinal ischemia/reperfusion. In retinas of old animals before ischemia, reactive gliosis, including activation of Muller cells, microglia and astrocytes, is increased compared to retinas from young and old/caloric restricted animals. Post-ischemia, the existing reactive gliosis in retinas of old animals is not neuroprotective and the reactive gliosis is even further increased in old animals compared to young or old/caloric restricted animals. In retinas from old/caloric restricted animals, inducible heat shock protein-70 and brain-derived neurotrophic factor increased more markedly after ischemia/reperfusion compared to retinas from young and old animals. Thus, compared to retinas in young animals, neurons of old animals may be more susceptible to cell death by secondary glial mechanisms after retinal ischemia/reperfusion. Caloric restriction in old animals is neuroprotective against damage in the retina following ischemia, perhaps by suppressing glial activity and by the neuroprotective effects of inducible heat shock protein-70 and brain-derived neurotrophic factor.     

Responses of rat trigeminal neurones to dental pulp cells or fibroblasts overexpressing neurotrophic factors in vitro

The adult dental pulp is innervated by sensory trigeminal axons and efferent sympathetic axons. Rat trigeminal ganglia extend neurites when co-cultivated in vitro with pulpal tissue explants, suggesting that pulpal cells secrete soluble molecules that stimulate the growth of trigeminal ganglion axons. In addition, cultured pulpal cells produce mRNAs for neurotrophins and glial cell line-derived neurotrophic factor-family members. These data suggest that neurotrophic factors are involved in the formation of a pulpal innervation. Here, we examine how pulpal cells and 3T3 fibroblasts overexpressing certain neurotrophic factors (nerve growth factor, brain-derived neurotrophic factor, neurotrophin-3, neurotrophin-4, glial cell line-derived neurotrophic factor or neurturin) influence survival and growth of single trigeminal ganglion neurones in vitro in quantitative terms. The results show that most of the neurotrophic factor-overexpressing fibroblasts induce similar neuronal soma diameters, but higher survival rates and neurite lengths compared with pulpal cells. With respect to neurite growth pattern, trigeminal ganglion neurones co-cultured with fibroblasts overexpressing nerve growth factor develop a geometry that is most similar to that seen in co-cultures with pulpal cells. We conclude that none of the fibroblasts overexpressing neurotrophic factors can fully mimic the effects of pulpal cells on trigeminal ganglion neurones, and that nerve growth factor promotes a neurite growth pattern most similar to the picture seen in co-cultures with pulpal cells.