金鱼Prox1基因cDNA的克隆及其在眼睛发育中的表达分析

脊椎动物的Prox1基因,与果蝇的转录因子prospero同源。为了探讨Prox1基因在金鱼眼睛发生过程中的表达图式,我们从金鱼眼睛SMART库中克隆了Prox1cDNA。它全长共2851bp,编码739个氨基酸。组织分布研究表明,Prox1主要分布于眼、脑、心、肝、脾和肾中。整体原位杂交显示,Prox1mRNA首先是在晶体期的晶体原基中有转录,心跳期则在未成熟晶体的细胞中和视网膜的幼芽区可以检测到。晶体纤维形成后,它主要定位于视纤维层和内网织细胞层。免疫组化显示,心跳期Prox1蛋白的定位与mRNA相同,晶体纤维形成以后,Prox1蛋白主要定位在晶体上皮细胞内侧的晶体纤维上一个环状区域,与Prox1mRNA的定位不同。这说明,Prox1基因在晶体发生过程中有重要作用,且在晶体的不同发育时期起的作用可能有所不同。另外,Prox1在晶体发育过程中有一个从内向外的变化过程。     

眼睛发育与骨形成蛋白信号通路的关系

在早期胚胎发育过程中,眼睛是由起源于不同胚层的几个部分经过一系列的诱导作用以及时间和空间上的相互协调作用形成的复杂而又具有精确功能的器官。在眼睛的形成发育过程中,许多信号通路及其相关的调控因子发挥着重要作用。本文主要关注眼睛发育过程与骨形成蛋白(BMP)信号通路的关系,BMP信号的激活能够诱导晶状体的再生和CLT(角膜到晶状体的分化转移)进程,维持睫状体的功能,促进视网膜的发生,影响巩膜的重塑和泪腺的发育。很多眼部疾病的发生与BMP信号通路的调节紊乱密切相关,因此可以将BMP信号通路作为一个潜在的药物靶点来探究治疗眼部疾病的方法。本文就BMP信号通路对眼睛发育的影响作一综述。     

中华鳖眼睛发育的形态及组织学观察

在孵化基质沙粒径为0.3～0.6mm、孵化温度为(33.0±0.5)℃、孵化基质的湿度为7%～10%、相对湿度为70%～85%的条件下孵化中华鳖(Trionyx sinensis)卵,孵化周期35～36 d.破壳取不同发育时期的胚胎并制作切片,观察眼睛发育的形态学和组织学特征.孵化第4 d头部两侧出现眼泡的突起;第6d眼睛开始出现色素,第14 d色素由褐色变为黑色;第7 d瞳孔出现,透过瞳孔可见晶状体;虹膜于第14d出现,第18、19 d瞳孔周围呈放射状;巩膜突自第19 d出现,第21 d增至最多,第23 d消失;上、下眼睑分别在第19 d和22 d出现,第32 d眼睑可覆盖瞳孔,眼睛形态与成体眼睛相似.表皮外胚层于第3 d形成角膜原基和晶体泡,第32 d角膜发育完成;第34 d晶状体发育完成;神经外胚层于44～48 h由前脑的两侧分化形成视泡,第3 d由视泡分化形成视杯,并逐步分化形成视网膜;第23 d视网膜的八层结构基本形成;第34 d视网膜发育完成.     

Vax基因与视觉神经系统的早期发育

视觉神经系统的发育与形成是一个相当复杂的过程,其基因调控机理是神经发育生物学领域的研究热点.Vax基因家族是新近发现的一类与视觉神经系统发育密切相关的同源异型盒基因,调控前脑、眼原基、视泡、视柄以及视网膜的发育.Vax-1参与色素上皮和视柄的分化;Vax-2则在视网膜及视神经背腹轴建立方面起重要作用.Vax基因的研究将对阐明视觉神经系统发育调控机制提供新的认识.     

在非洲爪蛙胚胎中过表达atf4影响早期神经发育及眼睛的形成

越来越多的证据表明转录激活因子4(atf4)是一个与胚胎发育相关的基因.该文研究了非洲爪蛙atf4在胚胎发育过程中的表达和功能.atf4特异性地表达在非洲爪蛙胚胎的脑部、眼睛、血岛、原肾、肝脏、胰腺以及胃和十二指肠的部分细胞.在非洲爪蛙胚胎的动物极半球过表达适量(不影响胚胎整体形态发生的剂量)的atf4,对神经上皮细胞中sox3的表达无明显影响,也不引起细胞凋亡;但是对原始神经元的标记基因以及预定形成前脑、中脑、视网膜和晶状体的前体细胞的标记基因表达都有不同程度的抑制,最终导致无晶状体小眼的表型.该研究结果首次提示对正常的早期神经发育及眼睛形成而言,atf4的活性需受到严格的调控.     

金鱼Prox1基因cDNA的克隆及其在眼睛发育中的表达分析

脊椎动物的Prox1基因，与果蝇的转录因子prospero同源。为了探讨Prox1基因在金鱼眼睛发生过程中的表达图式，我们从金鱼眼睛SMART库中克隆了Prox1 cDNA。它全长共2 851bp，编码739个氨基酸。组织分布研究表明，Prox1主要分布于眼、脑、心、肝、脾和肾中。整体原位杂交显示，Prox1 mRNA首先是在晶体期的晶体原基中有转录，心跳期则在未成熟晶体的细胞中和视网膜的幼芽区可以检测到。晶体纤维形成后，它主要定位于视纤维层和内网织细胞层。免疫组化显示，心跳期Prox1蛋白的定位与mRNA相同，晶体纤维形成以后，Prox1蛋白主要定位在晶体上皮细胞内侧的晶体纤维上一个环状区域，与Prox1 mRNA的定位不同。这说明，Prox1 基因在晶体发生过程中有重要作用，且在晶体的不同发育时期起的作用可能有所不同。另外，Prox1在晶体发育过程中有一个从内向外的变化过程。     

光敏核不育水稻叶片中钙调素mRNA原位杂交分析

采用组织ＲＮＡ原位杂交技术研究钙调素基因在光敏核不育水稻接受光周期信号后时空表达特征的结果表明,钙调素基因在不同光周期诱导后,吉片在育性转换对不周期敏感的不同发育时期中,ｍＲＮＡ表达量和空间分布上没有明显的差异,与光敏核不育水稻育性不存在直接的关系,可能与光敏色素信号系统调节叶绿体发育及光合作用有关。     

光敏核不育水稻叶片中钙调素mRNA原位杂交分析(简报)

采用组织RNA原位杂交技术研究钙调素基因在光敏核不育水稻接受光周期信号后时空表达特征的结果表明,钙调素基因在不同光周期诱导后,叶片在育性转换对光周期敏感的不同发育时期中,mRNA表达量和空间分布上没有明显的差异,与光敏核不育水稻育性不存在直接的关系;叶肉细胞中钙调素mRNA的大量表达,可能与光敏色素信号系统调节叶绿体发育及光合作用有关。     

猫视网膜多巴胺能无长突细胞发育的中央—周边梯度

本文报道用酪氨酸羟化酶(TH)免疫细胞化学方法标记猫视网膜多巴胺(DA)能无长突细胞发育的中央—周边梯度。TH阳性反应的I型DA能无长突细胞在发育成熟过程中呈现时空顺序的中央—周边梯度:(1)P_1时期分化较高的细胞,即染色深,胞体大,具有2—4支树突,分枝分布于内网状层(IPL),最外缘的星状Ⅰ_1类细胞大都集中于视网膜的中央部位;而分化较低的细胞,即染色淡,胞体小,具有1—2支树突,分枝分布于IPL外层和中层的不规则形Ⅰ_3类细胞大都集中于视网膜的周边部位;介于两者之间的Ⅰ_2类细胞散在分布于整个视网膜。它们形成了空间分布上的中央—周边分化成熟梯度。(2)随着发育进程,Ⅰ_1类细胞数增多,分布区逐渐从中央向四周扩展,由占视网膜总面积的30%(P_1时)增至65%(P_6时),P_(13)时达97%。开眼后P_(13)时,由于Ⅰ_1类细胞分布已扩展至周边,中央区和周边区间细胞平均直径和树突发育成熟程度的差别逐渐缩小,Ⅰ型DA能无长突细胞发育成熟的中央—周边梯度明显减弱。至P_(23)时,周边区细胞对TH抗体免疫反应强度以及形态学上成熟程度均相似中央区者,上述中央—周边梯度特征则完全消失。I型DA能无长突细胞发育成熟过程中呈现时空顺序的中央—周边梯度特征是视网膜个体发育过程中的暂时现象,它与视网膜中一些神经发生过程存在平行关系。它在视网膜神经发生中的作用,文中进行了讨论。     

低亲和性神经营养素受体p75在人胚胎视网膜中的表达

p75神经营养素受体在视网膜的发育以及再生过程中发挥着重要的作用,而在人类视网膜中的分布状况尚未被研究. 利用免疫组织化学方法,在光镜水平下确定了p75在人胚胎发育5、6和7个月的视网膜中的分布情况. 在视网膜神经节细胞层出现最强的p75免疫阳性反应,在其他各层也有较弱的免疫阳性反应. 在胚胎6、7月的视网膜中,主要由Müller细胞的终足构成的内界膜上出现了比较强的p75表达. p75在人胚胎视网膜中的分布情况与大鼠视网膜中很类似,主要表达在Müller细胞, 在神经节细胞上也可能有表达.     

Alpha-Fetoprotein in Retina and Lens of the Human Eye at Early Stages of Prenatal Development

The presence of alpha-fetoprotein (AFP) was shown in the retina and lens of the human fetal eye at different stages of prenatal development. PCR analysis revealed AFP mRNA neither in the retina nor in the lens, whereas in the fetal liver (control) AFP mRNA was found to be expressed. The data obtained indicate that AFP is not synthesized in retinal and lens cells of the human fetal eye but is imported from elsewhere to be taken up by these cells. The presence of AFP in the retina and lens implies its involvement in early morphogenesis and differentiation of these ocular tissues during prenatal human development.     

Cavefish as a model system in evolutionary developmental biology

The Mexican tetra Astyanax mexicanus has many of the favorable attributes that have made the zebrafish a model system in developmental biology. The existence of eyed surface (surface fish) and blind cave (cavefish) dwelling forms in Astyanax also provides an attractive system for studying the evolution of developmental mechanisms. The polarity of evolutionary changes and the environmental conditions leading to the cavefish phenotype are known with certainty, and several different cavefish populations have evolved constructive and regressive changes independently. The constructive changes include enhancement of the feeding apparatus (jaws, taste buds, and teeth) and the mechanosensory system of cranial neuromasts. The homeobox gene Prox 1, which is expressed in the expanded taste buds and cranial neuromasts, is one of the genes involved in the constructive changes in sensory organ development. The regressive changes include loss of pigmentation and eye degeneration. Although adult cavefish lack functional eyes, small eye primordia are formed during embryogenesis, which later arrest in development, degenerate, and sink into the orbit. Apoptosis and lens signaling to other eye parts, such as the cornea, iris, and retina, result in the arrest of eye development and ultimate optic degeneration. Accordingly, an eye with restored cornea, iris, and retinal photoreceptor cells is formed when a surface fish lens is transplanted into a cavefish optic cup, indicating that cavefish optic tissues have conserved the ability to respond to lens signaling. Genetic analysis indicates that multiple genes regulate eye degeneration, and molecular studies suggest that Pax6 may be one of the genes controlling cavefish eye degeneration. Further studies of the Astyanax system will contribute to our understanding of the evolution of developmental mechanisms in vertebrates.     

Step-wise specification of retinal stem cells during normal embryogenesis

The specification of embryonic cells to produce the retina begins at early embryonic stages as a multi-step process that gradually restricts fate potentials. First, a subset of embryonic cells becomes competent to form retina by their lack of expression of endo-mesoderm-specifying genes. From these cells, a more restricted subset is biased to form retina by virtue of their close proximity to sources of bone morphogenetic protein antagonists during neural induction. During gastrulation, the definitive RSCs (retinal stem cells) are specified as the eye field by interactions with underlying mesoderm and the expression of a network of retina-specifying genes. As the eye field is transformed into the optic vesicle and optic cup, a heterogeneous population of RPCs (retinal progenitor cells) forms to give rise to the different domains of the retina: the optic stalk, retinal pigmented epithelium and neural retina. Further diversity of RPCs appears to occur under the influences of cell-cell interactions, cytokines and combinations of regulatory genes, leading to the differentiation of a multitude of different retinal cell types. This review examines what is known about each sequential step in retinal specification during normal vertebrate development, and how that knowledge will be important to understand how RSCs might be manipulated for regenerative therapies to treat retinal diseases.     

Retinal homeobox genes and the role of cell proliferation in cavefish eye degeneration

The teleost Astyanax mexicanus exhibits eyed surface dwelling (surface fish) and blind cave dwelling (cavefish) forms. Despite lacking functional eyes as adults, cavefish embryos form eye primordia, which later arrest in development, degenerate and sink into the orbit. We are comparing the expression patterns of various eye regulatory genes during surfacefish and cavefish development to determine the cause of eye degeneration. Here we examine Rx and Chx/Vsx family homeobox genes, which have a major role in cell proliferation in the vertebrate retina. We isolated and sequenced a full-length RxcDNA clone (As-Rx1) and part of a Chx/Vsx(As-Vsx2) gene, which appear to be most closely related to the zebrafish Rx1 and Alx/Vsx2 genes respectively. In situ hybridization shows that these genes have similar but non-identical expression patterns during Astyanax eye development. Expression is first detected in the optic vesicle, then throughout the presumptive retina of the optic cup, and finally in the ciliary marginal zone (CMZ), the region of the growing retina where most new retinoblasts are formed. In addition, As-Rx1 is expressed in the outer nuclear layer (ONL) of the retina, which contains the photoreceptor cells, and As-Vsx2 is expressed in the inner nuclear layer, probably in the bipolar cells. With the exception of reduced As-Rx-1 expression in the ONL, the As-Rx1 and As-Vsx2 expression patterns were unchanged in the developing retina of two different cavefish populations, suggesting that cell proliferation is not inhibited. These results were confirmed by using PCNA and BrdU markers for retinal cell division. We conclude that the CMZ is active in cell proliferation long after eye growth is diminished and is therefore not the major cause of eye degeneration.     

Keeping an eye on retinoic acid signaling during eye development

Retinoic acid is a metabolic derivative of vitamin A that plays an essential function in cell-cell signaling by serving as a ligand for nuclear receptors that directly regulate gene expression. The final step in the conversion of retinol to retinoic acid is carried out by three retinaldehyde dehydrogenases encoded by Raldh1 (Aldh1a1), Raldh2 (Aldh1a2), and Raldh3 (Aldh1a3). Mouse Raldh gene knockout studies have been instrumental in understanding the mechanism of retinoic acid action during eye development. Retinoic acid signaling in the developing eye is particularly complex as all three Raldh genes contribute to retinoic acid synthesis in non-overlapping locations. During optic cup formation Raldh2 is first expressed transiently in perioptic mesenchyme, then later Raldh1 and Raldh3 expression begins in the dorsal and ventral retina, respectively, and these sources of retinoic acid are maintained in the fetus. Retinoic acid is not required for dorsoventral patterning of the retina as originally thought, but it is required for morphogenetic movements that form the optic cup, ventral retina, cornea, and eyelids. These findings will help guide future studies designed to identify retinoic acid target genes during eye organogenesis.