蛋白磷酸酶PP-1c在不同倍性鱼6种组织中的分化表达模式

以异源四倍体鲫鲤及其二倍体父/母本(湘江野鲤/红鲫)和子代三倍体湘云鲫等为实验材料,运用Western-blotting技术及荧光免疫组织化学技术等实验手段,分析了PP-1的催化亚基在上述不同倍性鱼体内的表达模式：蛋白水平检测发现PP-1c在不同倍性鱼的大脑、心脏、肌肉、肾脏、肝脏和性腺6种主要器官组织中均有表达,且不同的组织中显示了明显的差异表达模式,而PP-1c在这4种不同鱼的肌肉组织中的表达差异更显著,其中在异源四倍体鲫鲤中表达最低,在父/母本红鲫中的表达水平相对较高,子代三倍体湘云鲫中的表达最高,这种差异性可能从生化的角度说明了子代与父母本之间的变异性。免疫荧光组化实验结果显示,从整体水平来看,4种不同鱼的同一组织中,PP-1c的表达模式是非常相似的,这可能从蛋白和细胞的水平说明了异源四倍体鲫鲤与其二倍体父/母本及子代三倍体湘云鲫之间的遗传相似性。但对于同一组织的不同细胞的具体表达部位是有差异的,具有细胞特异性。     

6 种胃肠激素样内分泌细胞在细鳞鲑消化道的定位

采用免疫细胞化学ABC方法,选择5-羟色胺(5-hydroxytryptamine,5-HT)、生长抑素(somatostatin,SS)、胰多肽(pancreatic polypeptide,PP)、胃泌素(gastrin,GAS)和P物质(substance P,SP)6种特异性哺乳类胃肠激素抗血清,对不同年龄段(1龄、2龄和3龄)细鳞鲑(Brachymystax lenok)的消化道内分泌细胞进行了免疫细胞化学定位研究.结果表明,仅在食管、胃贲门、胃体和胃幽门检出有5-HT、SS和PP阳性细胞的分布(除1龄鱼食道),且这3种内分泌细胞均大量定位于胃部;在前肠、中肠、后肠和直肠中均未检测到这3种内分泌细胞的阳性反应.在3个年龄段的细鳞鲑胃肠各部位均未检测到GAS、GLU和SP阳性细胞.细鳞鲑的5-HT、SS和PP与其他有胃鱼类的内分泌细胞一样,可分为2种类型,即开放型和闭合型,这类细胞主要通过腔分泌和旁分泌两种方式释放激素.5-HT、SS和PP这3种细胞在1龄幼鱼消化道内就已经发育成熟,其对胃肠道活动的调节作用已经与成鱼没有差别,细胞的分布密度随着细鳞鲑的年龄增长不断增加.本研究揭示了不同生长阶段细鳞鲑消化道中这6种胃肠激素内分泌细胞的发育特征,并阐明了这些胃肠激素细胞在细鳞鲑幼鱼消化道的分布、形态以及生理作用.     

锯缘青蟹脑内5-HT和NPY的免疫细胞化学定位

为探讨5-羟色胺和神经肽Y在锯缘青蟹脑内的存在与否和分布状况,应用免疫细胞化学技术,在光学显微镜下观察5-羟色胺和神经肽Y阳性细胞和神经髓质的形态和分布。结果表明：在锯缘青蟹脑中共12个胞体群和11个神经髓质中,前脑和中脑中有4个细胞群和6个神经髓质检出5-HT免疫阳性反应;中脑和后脑中有7个细胞群和3个神经髓质具有NPY免疫阳性反应。5-羟色胺和神经肽Y在锯缘青蟹脑内的特异性分布,为其参与神经牛理活动提供了形态学证据。     

PHGPx在体外共培养山羊睾丸生精细胞中的定位研究

为探讨磷脂氢谷胱甘肽过氧化物酶(PHGPx)在雄性动物生殖机能中的作用及其在体外培养山羊睾丸生精细胞的定位,在已成功构建的山羊睾丸支持细胞-生精细胞共培养的基础上,制作细胞爬片,采用免疫细胞化学技术定位PHGPx的表达。结果显示,PHGPx在支持细胞、精原细胞中不表达,在圆形精子细胞细胞质检测到阳性表达产物。表明PHGPx在精子发生后期圆形精子的变态发育中起特定的调节作用,但作用机制有待深入研究。     

一氧化氮研究方法评析

本文介绍目前国内外研究一氧化氮(NO)的主要方法,包括电子顺磁共振成像技术(EPR Imaging technique)、液内分光光度检测法、NADPH-d 组织化学法、一氧化氮合酶—Ⅰ(NOS-Ⅰ)免疫细胞化学法及 NOS-Ⅰ免疫荧光技术等。     

Ultrastructural and immunocytochemical investigation of pathogen development and host responses in resistant and susceptible wheat spikes infected by Fusarium culmorum

Pathogen development and host responses in wheat spikes of resistant and susceptible cultivars infected by Fusarium culmorum causing Fusarium head blight (FHB), were investigated by means of electron microscopy as well as immunogold labelling techniques. The studies revealed similarities in the infection process and the initial spreading of the pathogen in wheat spikes between resistant and susceptible cultivars. However, the pathogen’s development was obviously more slow in the resistant cultivars as in comparison to a susceptible one. The structural defence reactions such as the formation of thick layered appositions and large papillae were essentially more pronounced in the infected host tissues of the resistant cultivars, than in the susceptible one. β -1,3-glucan was detected in the appositions and papillae. Furthermore, immunogold labelling of lignin demonstrated that there were no differences in the lignin contents of the wheat spikes between susceptible and resistant cultivars regarding the uninoculated healthy tissue, but densities of lignin in host cell walls of the infected wheat spikes differed distinctly between resistant and susceptible cultivars. The lignin content in the cell walls of the infected tissues of the susceptible wheat cultivar increased slightly, while the lignin accumulated intensely in the host cell walls of the infected wheat spikes of the resistant cultivars. These findings indicate that lignin accumulation in the infected wheat spikes may play an important role in resistance to the spreading of the pathogen in the host tissues. Immunogold labelling of the Fusarium toxin DON in the infected lemma showed the same labelling patterns in the host tissues of resistant and susceptible cultivars. However, there were distinct differences in the toxin concentration between the tissues of the susceptible and resistant cultivars. At the early stage of infection, the labelling densities for DON in resistant cultivars were significantly lower than those in the susceptible one. The present study indicates that the FHB resistant cultivars are able to develop active defence reactions during infection and spreading of the pathogen in the host tissues. The lower accumulation of the toxin DON in the tissues of the infected spikes of resistant cultivars which results from the host’s defence mechanisms may allow more intensive defence responses to the pathogen by the host.     

GTH-cells in the pituitary of the African catfish,Clarias gariepinus,during gonadal maturation: an immuno-electron microscopical study

In an ultrastructural immunocytochemical study we investigated the development of the gonadotropic cells in the pituitary of two to six months old male African catfish in relation to testicular development. In this period, pituitary and testicular tissue samples were collected on five occasions (groups I–V). Blood samples could only be taken from the fish in groups III–V. The testicular development was divided in three stages i.e., immature (only spermatogonia, group I), early (spermatogonia and spermatocytes, groups II and III) and advanced (all germ cell stages including spermatozoa, groups IV and V) spermatogenesis. 11-Ketotestosterone blood levels were low, except for the last group. Antisera were raised against the complete catfish α,βGTH-II, as well as to the separate α- and β-subunits of catfish GTH-II. In the proximal pars distalis of immature fish, undifferentiated cells, somatotrops, putative thyrotrops (pTSH) and putative gonadotrops (pGTH) were found. In the two latter, secretory granules were labeled with anti-αGTH, but not with anti-βGTH-II. pTSH- and pGTH-cells were distinguished on the basis of the size of their secretory granules. During early spermatogenesis, two classes of putative gonadotrops could be distinguished. One type had the same immunocytochemical and ultrastructural characteristics as in immature fish; the secretory granules in the second cell type, which was more abundant, were also immunopositive for anti-βGTH-II. The mean volume of the secretory granules in these GTH-II cells was three times larger than that in the early appearing pGTH-cells. In addition, the later appearing GTH-II cells contained large inclusions, known as globules. These structures labeled with anti-αβGTH-II and with anti-βGTH-II, but not with anti-αGTH. It is assumed that the globules are involved in a differential storage and/or breakdown of the GTH-II subunits. During advanced spermatogenesis the two gonadotropic cell types could still be distinguished, but the early appearing pGTH-cell type was scarce. The present observations permit the conclusion that the early appearing cells may be GTH-I cells. However, definitive proof about their identity depends on the availability of antibodies or cDNA probes specific for GTH-I.