人胎视网膜超氧化物歧化酶和波形蛋白免疫阳性细胞的发育

目的：观察人胎视网膜超氧化物歧化酶（SOD）和波形蛋白（VIM）免疫阳性细胞的分布发育。方法：不同孕龄的人胎16例,ABC免疫细胞化学方法显示视网膜SOD和VIM免疫阳性细胞,结果：（1）SOD免疫阳性细胞：E15w节细胞层开始出现SOD免疫阳性细胞;D20W和E28W SOD免疫阳性细胞排列较整齐,分布于视网膜的外核层,内核层,节细胞层,其数量增多,其中内核层SOD免疫阳性细胞增多明显。（2）VIM免疫阳性细胞的发育：E15w内界膜开始出现Muller细胞的VIM免疫阳性终足,并见VIM免疫阳性突起伸向外界膜;E20 人VIM免疫阳性物质集中于内界膜,并见VIM免疫阳性突起伸向外界膜;E28wVIM免疫阳性物质的数量较E20w以前各孕龄明显增多,除色素上皮和视杆视锥层外均有VIM免疫阳性物质出现,除伸向外界膜的VIM免疫阳性突起外,还内网层,内核层,节细胞层和神经纤维层还可见水平走行的细胞突起,结论：（1）视网膜发育基本成熟后,视网膜SOD可能主要来源于内核层的SOD免疫阳性细胞。（3）视网膜神经纤维髓鞘是从内向外逐渐形成的。     

人胚胎视网膜5—羟色胺样免疫反应神经元的发育

本文应用免疫细胞化学技术研究了妊娠第8周至第28周人胚胎视网膜5—羟色胺样免疫反应(5—Hydroxytryptamine like immunoreac-tion,5—HT—LI)神经元的定位和发育,观察了切片和平铺片共40例。结果表明,第10周人胚胎视网膜开始出现5—HT—LI神经元,它们分散在内成神经细胞层,胞体圆形,卵圆形,直径6—7μm。人胎第11至第13周5—HT—LI神经元数量增加,细胞大小比较一致,并处于迁移之中;此时约49.5%的5—HT—LI细胞位于节细胞层,其余位于节细胞层以外。第17至第18周位于节细胞层的5—HT—LI细胞达88.5%,位于内核层约11.5%;其中位于内核层最内缘的细胞约7.3%,这些细胞大致可分为大、中、小3型,其直径分别是8—10μm,6—7μm,和5—6μm,每平方毫米细胞数为8.1±2.19细胞(平均数±标准差)。第21周时为10.7±2.16细胞/mm~2。第14周以前可见5—HT—LI纤维分布在神经纤维层,自此之后,未见5—HT—LI纤维存在,上述结果提示,5—HT—LI神经元是发育中的无长突细胞,这些细胞在第10周人胎视网膜出现,至第17至18周已发育到相当阶段,但未见细胞间的纤维联系,此后神经元继续发育至成熟。     

ENK与CaBPs在PPE-GFP转基因小鼠视网膜细胞中的共存

目的:以PPE-GFP转基因小鼠为研究工具,观察绿色荧光蛋白(GFP)阳性的脑啡肽(ENK)能神经元与钙结合蛋白D28K(CB)、钙视网膜蛋白(CR)和小白蛋白(PV)等钙结合蛋白(CaBPs)成员在视网膜的分布及共存情况。方法:利用免疫组织化学和免疫荧光双标染色的方法。结果:GFP阳性的ENK能细胞主要分布在视网膜内核层内缘,少量分布在节细胞层。所有的GFP阳性细胞均与神经元标志物NSE共存,但不与星形胶质细胞标志物GFAP共存。GFP与CB、CR和PV均有部分共存,其中GFP/CB共存神经元占GFP阳性细胞的8.65%,占CB阳性细胞的5.84%;GFP/CR共存神经元占GFP阳性细胞的18.18%,占CR阳性细胞的14.28%,且共存细胞仅见于内核层;GFP/PV共存细胞占GFP阳性细胞的68.75%,占PV阳性细胞的91.67%,共存细胞主要位于内核层,少量见于节细胞层。结论:ENK能神经元在视网膜内具有板层特异性的分布特点和与钙结合蛋白成员有不同的共存模式,上述结果为深入研究小鼠视网膜ENK能神经元的功能意义提供了形态学依据。     

大鼠视网膜发育过程中细胞增殖与凋亡的形态学

目的:观察视网膜发育过程中增殖细胞核抗原(PCNA)的表达变化及细胞凋亡状态。方法:14~20 d胎鼠(E14-20 d)、生后0~15 d幼鼠(P0-15)及成年鼠(P36),取出眼球组织,免疫组织化学显色及原位末端标记(TUNEL)观察PCNA的表达和细胞凋亡变化。结果:发育早期E14~P7,可观察到有较强PCNA阳性表达,P9时PCNA阳性细胞数目明显减少,P15开始未见明显PCNA阳性表达。E18可观察到凋亡细胞,随着发育的进行,凋亡细胞逐渐增多,P7时凋亡细胞数目最多,之后又逐渐减少,成熟的视网膜组织内未见明显凋亡细胞。结论:在视网膜神经细胞发育过程中,细胞的增殖和凋亡呈现了有序的动态变化,正是这种有序的增殖和凋亡的平衡才使视网膜最终发育成了正常的组织结构及功能。     

人胎海马结构小白蛋白免疫反应性神经元的分布

目的 观察人胎海马结构小白蛋白（PV）免疫反应性神经元的分布。方法 取孕龄为30周的人胎尸体,用ABC免疫细胞化学方法显示PV免疫反应性神经元。结果 海马结构的各区域内均有丰富的PV免疫反应性神经元分布,以锥体细胞怪最为密集。CA1、CA2、CA3始层PV免疫反应性神经元呈散在分布,胞体形态多样,细胞的突起伸向浅怪的始层和深层的分子层;分子层PV免疫反应性神经元较稀少。门区PV免疫反应性神经元分布密集,但细胞分层不明显,可见部分细胞的突起伸向齿状回;齿状回PV免疫反应性神经元集中分布于颗粒细胞层,其余各层在有少量散在PV免疫反应性神经元,细胞染色浅谈,无明显突起,下托复合体PV免疫反应性神经元主要分布于锥体细胞层,始层和分子层较稀少,细胞淡染,突起不明显。结论 海马结构的各区域均有丰富的PV免疫反应性神经元分布,主要分布于锥体细胞层和齿状回的颗粒层。各区域PV免疫反应性神经元发育成熟的时间可能并不同步,CA1－3和门区PV免疫反应性神经元发育成熟早于齿状回和下托复合体。     

CD20免疫反应阳性细胞在人胎阑尾的发育

目的从形态学的角度探讨人胎阑尾内CD20阳性B细胞在各胎龄段的发育。方法收集13～28周人胎22例,测量各胎儿顶臀长（CRL）。按Patten法确定胎龄,取其阑尾,经HE染色、免疫组织化学sP法染色,光镜观察。结果13～16周阑尾上皮已经完全演变成为单层柱状,胎儿阑尾绒毛出现,结构分层逐渐明显,淋巴细胞散在分布,淋巴小结雏形逐渐形成;14周CD20免疫反应阳性细胞出现,数量少,单个分布于结缔组织内;17～20周CD20阳性反应细胞增多,成群分布;23—28周粘膜固有层内CD20阳性细胞参与构成淋巴小结。统计学处理证实随胎龄增加,CD20阳性细胞数量逐渐增多（P〈0．01）。结论14周CD20阳性细胞出现,随胎龄增加数量增多,主要参与构成淋巴小结。     

28周人胎视皮质含钙结合蛋白神经元的区域分布

用免疫细胞化学方法研究了28周人胎树皮质17区和18区含CALBINDN(CB)及含PARVALBUMIN(PV)神经元的分布.结果发现,17区会CB及台PV神经元的数量均较18区者多得多,而且与18区比较,它们还可见于皮质较浅的层次,使 17/18区交界处明显可辨.结合我们对人类新皮质含CB及含PV神经元的发育研究结果,本实验结果提示,人类视皮质17区神经元的发育和成熟早于18区神经元.     

人胎儿和新生儿脊髓和DRG内nNOS阳性神经元的表达和分布

目的：观察24～27w人胎儿和新生儿的脊髓和背根神经节(DRG)内神经元型一氧化氮合酶(nNOS)阳性神经元的表达和分布。方法：ABC免疫细胞化学方法。结果：(1)24～27w人胎儿胸髓和腰1～3节段的中间带外侧核和前角Ⅷ层和Ⅸ层内可观察到nNOS阳性神经元。颈、胸、腰各段DRG内nNOS免疫阳性神经元占DRG细胞总数的84％～88％。(2)新生儿脊髓和DRG内nNOS免疫阳性神经元的分布情况与上述胎儿相似。但DRG内nNOS免疫阳性神经元的体积有所增加，数量明显减少，约占DRG细胞总数的70％～73％。结论：人胎儿脊髓和DRG在发育的24～27w至出生时，nNOS阳性神经元的表达在定位分布上无差异，但随胚胎发育阳性神经元的数量明显减少。     

大鼠视网膜神经元NOS与Parvalbumin的共存研究

用NADPH脱氢酶组化及Parvalbumin免疫组化双标记技术观察了正常大鼠视网膜一氧化氮合酶(NOS)与Parvalbumin(PV)的分布,结果显示NOS阳性神经元主要位于内核层内缘带第二列,少数位于节细胞层,胞体圆形/卵圆形,直径8～12μm,细胞一侧发出突起伸向内网层1、3、5亚层,以第3亚层最为明显,PV免疫反应(PV—Ⅰ)神经元位于内核层最内缘第一列,少数位于第二列、中间部及节细胞层,胞体卵圆形,直径6～10μm,由胞体一端发出突起伸向内网层第1、5亚层.神经纤维层可见PV～Ⅰ纤维.内核层内缘第二列可见少数双标阳性细胞,在它们的PV免疫反应胞质内散布有NOS颗粒.实验结果表明 NOS阳性神经元与PV～Ⅰ神经元均为无长突细胞,分属不同的亚型,少效PV～Ⅰ神经元属节细胞,个别双标细胞可能为另一种亚型的无长突细胞,提示NOS与PV在视觉信息传递中可能存在某些联系.     

人胎视网膜发生的光镜和电镜观察

本文在光镜下观察40例人胎视网膜的发生,在电镜下观察15例人胎视网膜视细胞、双极细胞、节细胞的发育。结果表明:胚胎第9周时神经上皮可分内、外成神经细胞层。第10周时内、外成神经细胞层之间的Chievitz带消失;第11周时节细胞从内成神经细胞层内迁;第13周节细胞与内成神经细胞之间出现内网层;第16周始双极细胞从外成神经细胞层中内迁形成外网层和内核层。第20周后视网膜各层形成。而视细胞、双极细胞、节细胞的超微结构于胎儿8个月后才发育完善。其结构与成人基本相同。     

Developmental expression of calretinin immunoreactivity in the human retina and a comparison with two other EF-hand calcium binding proteins.

This paper reports the localization pattern of calretinin, a calcium-binding protein, in the human retina during development, as studied by immunohistochemistry. A comparison is made of the cellular distribution of calretinin with two other calcium-binding proteins, calbindin and parvalbumin, recently reported by us in the human retina, and by parallel labeling with both antisera in the same tissues. At 11-12 weeks of gestation, calretinin immunoreactivity was expressed in many prospective ganglion cells of the central inner neuroblastic zone. At 16-17 weeks of gestation, the immunoreactivity was localized in the ganglion cell layer, inner plexiform layer, and in most differentiated amacrine, horizontal and cone cells located in the central (1-2 mm temporal from optic disc) to midperipheral parts of the retina. By midgestation (20-21 weeks), calretinin immunoreactivity was strongly developed in the cone photoreceptors. Parallel labeling with calbindin and parvalbumin antisera revealed that the calretinin-positive horizontal cells were somewhat smaller and less frequent and less intense than the calbindin- and parvalbumin-positive counterparts, at 16-21 weeks of gestation. No horizontal cells were calretinin immunopositive in the postnatal (four-month-old infant) and adult retinas examined. Also, at both stages, a few bipolar and cone cells were weakly immunoreactive. These observations suggest a critical role for calretinin in the development and maturation of a select class of horizontal cells. The widespread expression of immunoreactivity in the early ganglion cells indicates that calretinin may be involved in their differentiation. The weak immunoreactivity pattern noted in the adult photoreceptor and bipolar cells, and an apparent lack of immunoreactivity in the mature horizontal cells, tends to indicate that, unlike calbindin and parvalbumin, calretinin plays little role in the transport and physiological buffering of Ca2+ in these neurons of the human retina. It appears, however, that calretinin is predominantly involved in both processes in amacrine cells.     

Ontogeny of two calcium-binding proteins (calbindin D-28K and parvalbumin) in the human inferior olivary complex and their distribution in the adults

The inferior olivary complex (IOC) is a prominent nuclear relay system of the medulla oblongata. Anatomically, it is connected to the cerebellum for coordination of motor activities. Calbindin D-28K (CALB) and parvalbumin (PV) are cytosolic calcium-binding proteins (CBP) that play a role in Ca²⁺ homeostasis. We examined their ontogeny and distribution in the fetal, postnatal and adult human IOC by immunohistochemistry. At 11–12 weeks of gestation (wg), calbindin immunoreactivity was present in the principal olive and the medial accessory olive, it was absent in the dorsal olive. Parvalbumin immunoreactivity developed at 16–17 wg in the ventral lamella and the lateral bulge of the principal olive only. Calbindin expression gradually increased from 20 to 37 wg, whilst by contrast, parvalbumin expression was moderate. By 37 wg, all three IOC subnuclei were immunopositive for both proteins. In a 3-month-old infant, parvalbumin was intensely developed in the olivary axons. In the adults (40- to 59-year-old), calbindin was distributed in most neurons, and olivocerebellar fibres, whereas parvalbumin was present in some neurons and few fibres. Parvalbumin expressed till 51 years, and disappeared by 59 years of age. Calbindin immunoreactivity in the olivary axons was declined at 70 years of age. The data suggest a differential distribution and requirement of these proteins in the human IOC maturation. It may be that the IOC utilizes mainly calbindin for Ca²⁺ buffering. The loss of parvalbumin with ageing might influence the excitability of the spared IOC neurons.     

The role of nitric oxide in the apoptosis of neurons in the retina of the human fetal eye

The locations of NADPH-diaphorase (NADPH-d), inducible NO synthase (iNOS), and TUNEL-immunoreactive neurons in the retina of human fetuses collected during the first to third trimesters of pregnancy were studied. High levels of NADPH-d activity were seen in the inner segments of light-sensitive cells, amacrine cells, and ganglion cells. The population of NADPH-d-positive amacrine cells included three types of neuron. Type 1 neurons were large and had sparse dendritic fields occupying the inner nuclear and outer retinal layers. Small type 2 neurons were located in the inner retinal layer. Ectopic amacrine cells, type 3, were located in the outer part of the ganglion layer. A high density of NADPH-d-positive neurons was seen in the central part of the retina, surrounding the central fovea and optic disk area. NADPH-d activity increased progressively during ontogenesis and correlated with the appearance of immunoreactive iNOS in neurons. iNOS labeled a subpopulation of amacrine and ganglion cells, which appeared at 20–21 weeks of development and reached a peak of immunoreactivity by the end of the third trimester. TUNEL-immunopositive neuron nuclei with signs of apoptotic destruction were seen at 30–31 weeks of pregnancy. The greatest apoptotic index was seen in the ganglion and amacrine cell populations. These data identify NO as a factor mediating apoptosis of neurons during the critical period of differentiation of interneuronal connections in the human retina. Director: Doctor of Biological Sciences M. A. Vashchenko Director: Doctor of Biological Sciences S. L. Kondrashov __________ Translated from Morfologiya, Vol. 129, No. 1, pp. 42–49, January–February, 2006.     

兔胎视网膜内P物质免疫反应神经元的发生

本文用免疫细胞化学ＡＢＣ法，研究了新西兰白兔１８、２２、２５、２６、２８和３０ｄ胎龄视网膜内Ｐ物质免疫反应（ＳＰＩＲ）神经元的发生。在胎龄１８和２２ｄ兔视网膜未见ＳＰＩＲ细胞体和纤维。在胎龄２５ｄ视网膜的节细胞层最先出现ＳＰＩＲ神经元，胞体浅染呈卵圆形，突起不明显，在神经纤维层偶见串珠状ＳＰＩＲ纤维，其平均细胞密度为１０４．６个细胞／ｍｍ￣２。到胎龄２６和２８ｄ时，在节细胞层的ＳＰＩＲ神经元的胞体渐深染，可见个别ＳＰＩＲ神经元发出粗而短的突起伸向内网层，平均细胞密度分别为３８７和７７９．５个细胞／ｍｍ２。到胎龄３０ｄ时ＳＰＩＲ神经元开始出现于内核层的内排细胞，但数量很少，胞体呈卵圆形，发出细突起伸入内同层，在节细胞层的ＳＰＩＲ神经元的突起分支增加。此时ＳＰＩＲ神经元平均细胞密度为３５７．４个细胞／ｍｍ￣２。     

大鼠,金黄地鼠和家兔视网膜内一氧化氮合酶分布的比较

用ＮＡＤＰＨ黄递酶组织化学染色法观察了正常成年大鼠、金黄地鼠和家兔视网膜内一氧化氮合酶的分布，并比较了３种不同动物的区别。结果显示，在视网膜内ＮＯＳ阳性神经元主要为分布于内核层的无长突细胞、节细胞层的移位无长突细胞和少数节细胞，不同种类动物的视网膜内，ＮＯＳ阳性细胞的配布、密度和细胞形态均有差异。大鼠视网膜内ＮＯＳ阳性细胞多尾于内核层无长突细胞和节细胞层移位无长细胞，偶见于视网膜节细胞。金黄地鼠视     

Early expression of recoverin in a unique population of neurons in the human retina

 The calcium-binding protein recoverin has been reported as present in photoreceptors, cone bipolar cells and sparse cells in the ganglion cell layer in the adult retinae of various vertebrate species. The present study was undertaken to clarify the developmental pattern of recoverin-immunoreactive cells in the human retina with particular attention to the cells in the inner retinal layers. In the adult human retina, small populations of recoverin-containing cells are present in the ganglion cell and nerve fiber layers. However, the precursors of these cells are quite numerous on the inner and outer borders of the nerve fiber layer in the fetal retina. By 13 weeks of gestation these cells express recoverin very intensely. By 24 weeks they are mature-looking with relatively large soma sizes (mean=118 μm²) and appear round, oval or multipolar in shape, with varying numbers of short processes. There follows a noticeable reduction of the mean soma size, but little change in morphology and process number during the remaining gestational stages up to and after birth. The mean numerical density of the recoverin-positive cells in the fetal inner retinal layers is gradually reduced from the high level at 13 weeks until birth, when there is a great drop to the adult level. The recoverin-immunoreactive cells in the ganglion cell layer demonstrate distinctively different developmental and morphological features from the principle neurons and glial cells in the retina. They are probably the neurons derived from the marginal zone of the retinal primordium that reside in the inner and outer borders of the nerve fiber layer due to the invasion of ganglion cell axons. The expression of recoverin in the neurons may be significant in maintaining an inside-out and centro-peripheral gradient of calcium concentration in the premature retina, thereby playing a role in determining the polarity of the differentiating ganglion cells and the growth of their axons in a centrifugal spatiotemporal order. Accepted: 3 June 1996     

Vaccination induces major histocompatibility complex class II expression in the Atlantic salmon eye

The aim of the study was to investigate the presence, distribution and density of major histocompatibility complex (MHC) class II+ cells in the ocular tissues of the Atlantic salmon, Salmo salar, prior to and following vaccination. Eyes were collected 14 days prior to and at 4, 11, 25 and 39 days and 4 months subsequent to vaccination with a commercial fish vaccine. A quantitative analysis was performed in sections on the number of immunopositive cells in the retinal layers. In all groups, MHC class II+ cells were detected in the area of the limbus but not in the central parts of the cornea. In the uvea, immunopositive cells were present in unvaccinated and vaccinated fish. Abundant immunopositive cells were identified in the choroid rete (or choroid gland) in all groups as well as in the ventral ciliary cleft, where macrophage-like MHC class II+ cells were seen. Quantitative histology of the retina revealed a significant increase in MHC class II+ cells in the outer plexiform layer (OPL) and the inner nuclear layer (INL) 4 days following vaccination. Positive cells were detected in all layers of the retina with the exception of the photoreceptor layer.