中国鲎成体血细胞的发生

应用透射电镜技术研究中国鲎成体血细胞发生。成体鲎血细胞于与生殖细胞发生相邻部位的结缔组织,其间以基膜为界。血细胞发生可分为原始血细胞、浆细胞和颗粒细胞３个阶段。在雄体,在雄体,血细胞在由基膜形成的网状结构中发育,并以此为界与生精小管相邻。早期原始血细胞核以常染色质为主,异染色质仅在晚原始血细胞核内边缘出现,胞质充满小不一的膜囊。浆细胞比原始血细胞大,细胞及核的形状多样,核异染色质呈粗网状分布,胞质     

中国鲎变形细胞的超微结构观察

鲎属于节肢动物门螯肢动物亚门肢口纲剑尾目,现存三属五种,分布于北美、中美和东南亚沿海一带。变形细胞是鲎血液中最主要的细胞成分。 被称为活化石的鲎是一种古老的动物,在近四亿年的漫长过程中其形态几乎没有变化。自1956年Bang等给美洲鲎注射格兰阴性杆菌引起血管内凝血以来,鲎变形细胞溶解物用于微量内毒素的检测,受到普遍重视。近年来国内从中国鲎(Tachyplelus tridentatus     

中国鲎幼体阶段(黄皮鲎)的形态特点

经研究发现,中国鲎幼体阶段（黄皮鲎）生殖盖板中叶不长于侧叶,尾呈三角形,上有一列小刺,腹甲后端尾上有三个棘突,尾两个侧边无小刺,腹甲背面无小刺分布,但其头胸甲和腹甲背面棘突较长,明显呈幼体状,中国鲎的胚后幼体发育从小到大,身体背部颜色处于黑色→土黄色→褐色的一种渐变过程,当中国鲎幼体头胸甲宽度处于10-25cm时,也就是胚后发育的第12至第16次蜕皮期间时,其体色呈土黄色,在粤西、海南、北部湾一带俗称“黄皮鲎”。所以,黄皮鲎为中国鲎一定阶段未成熟的幼体,而不是一个新种或亚种。标本采自广东省湛江市,保存于湛江海洋大学水生生物博物馆。     

中国鲎变形细胞超微结构的进一步观察

中国鲎的变形细胞可在血液中发育成熟,根据其超微结构,可分为原始变形细胞、幼变形细胞和变形细胞三个阶段。 变形细胞受到外环境或内毒素的刺激后,第一型颗粒出现一系列变化,最终把内容物释放到细胞外。与此同时,细胞表面形成若干胞质性突起,边缘带分散断裂,质膜下和胞质性突起中出现微丝。第二型颗粒无变化,提示它们和血液凝胶化过程无直接关系。     

烟草与蓝猪耳胚胎发生中细胞表面三种凝集素受体的动态分布

以异硫氰酸甲酯(FITC)标记的三种凝集素(伴刀豆凝集素, 麦芽凝集素和大豆凝集素)为荧光探针,对烟草及蓝猪耳各发育时期胚细胞表面的凝集素受体进行了定位.结果显示胚柄基部荧光信号最强,沿胚柄单列细胞向胚体方向渐次减弱.以后随着胚柄功能的逐渐丧失而改变.同时,三种凝集素受体集中分布于胚柄细胞间的分裂面;凝集素受体在原胚中分布的另一个特点是聚集于新形成的细胞壁上.随着胚胎发育至分化阶段,凝集素受体则主要分布在胚体细胞的外切向壁上;三种凝集素受体的动态分布显示了凝集素受体的分布与细胞分裂之间的密切关系及其调控胚胎发育的作用.     

中国鲎变形细胞活体观察、活体染色及其对细菌感染反应的研究

目前,国外应用鲎变形细胞提取物(或称溶解物,简称鲎试剂)于医药、卫生和食品等行业检测内毒素和致热原(Jorgensen 1973;Weary,1980),具有灵敏度高,快速和操作简便等优点。近年来,国内某些单位已开始研制应用。     

三刺鲎(Tachypleus tridentatus)血淋巴的研究 1.血蓝蛋白的分离及其免疫原性的研究

鲎在分类学上属于节肢动物门,剑尾目、鲎科。由于它在古生代寒武纪时期业已出现,故有“活化石”之称。目前世界上已发现有三属五种鲎。我国东南沿海盛产三刺鲎。鲎的血淋巴液中含血蓝蛋白,它是一种高分子量含铜呼吸蛋白,有一定构型,又能与其他分子连接,可作为研究细胞表面分子的标志物(刘黎等,1979);由于它的分子量很大又可作为免疫原(Malley,A 1965)(Lamy,J 1979),因此,引起了人们的关注。本文简要报告三剌鲎血蓝蛋白的分离纯化及其免疫原性的研究结果。     

人-牛异种克隆胚构建及其线粒体来源

通过人-牛异种核移植技术获得异种克隆囊胚, 便于在不消耗人类卵母细胞的情况下从异种克隆胚中分离出人类干细胞。通过透明带下注射法将人胎儿成纤维细胞和牛耳成纤维细胞分别注入去核牛卵母细胞中构建异种和同种胚胎, 并比较两者之间的融合率、卵裂率、8-细胞发育率以及囊胚率。并对处于2-细胞、4-细胞、8-细胞、桑椹胚、囊胚阶段的异种克隆胚的线粒体DNA来源进行检测。结果表明, 异种克隆胚体外各个阶段的发育率均低于同种克隆胚, 尤其是8-细胞到囊胚阶段的发育率, 以及囊胚率都显著低于同种克隆胚(P<0.05)。异种克隆胚在2-细胞到桑椹胚阶段检测到人、牛线粒体DNA共存, 囊胚阶段只检测到牛线粒体DNA。结果表明: 牛卵母细胞可以重编程人胎儿成纤维细胞, 完成异种克隆胚植入前的胚胎发育, 异种克隆胚由于核质相互作用的不谐调, 影响其发育能力, 使其囊胚率显著低于同种克隆胚。牛线粒体DNA存在于植入前异种胚胎发育的各个阶段。异种克隆胚胎用于人类胚胎干细胞分离具有可行性。     

Bac-to-Bac/BmNPV杆状病毒在家蚕幼虫中表达重组鲎C因子

鲎C因子是鲎血细胞中一种对内毒素具有高亲和力的丝氨酸蛋白酶,被内毒素激活后可水解人工合成的三肽底物,因而能替代传统的鲎试剂用于内毒素检测。通过RT-PCR从东方鲎血细胞中扩增出鲎C因子基因(factor C)序列,利用Bac-to-Bac/BmNPV杆状病毒表达系统在家蚕幼虫中表达该蛋白,并对其进行活性测定。结果显示:被感染的家蚕血清稀释后能检测到Factor C活性,稀释500倍的血清可以用于内毒素检测,且其灵敏度能达到0.2 EU/mL,有望开发新型的、低成本的内毒素检测试剂。     

鲎源抗菌肽的研究及其潜在应用价值

鲎血细胞来源的抗菌肽,在鲎天然免疫中起至关重要的作用,它对外源病原菌具有抗性作用,降低了外源病原菌对鲎活体的致病性,增强了鲎的天然免疫能力。鲎源抗菌肽有一些不同于其他来源的抗菌肽的优势,对鲎源抗菌肽的研究有很重要的实用意义。概述了鲎源抗菌肽的性质、分子结构、基因序列及其制备,并对它们的潜在应用价值进行了论述。     

Induction of Secondary Embryos by Intra- and Interspecific Grafts of Center Cells under the blastopore in Horseshoe Crabs

When the cell mass (center cells) of the early gastrulae in both American and Asian horseshoe crabs was grafted into the embryo of the homologous species, secondary embryos were formed as a result of these grafts. Secondary embryos were also formed in a similar way when the center cells of heterologous embryos were grafted between the American and Asian horseshoe crab embryos. The characteristics of the secondary embryos were similar to the host embryos in both cases, indicating that the center cells played the roles same as those by the amphibian organizer. The homogenate of center cells also induced the formation of secondary embryos. This is the first published study in which secondary embryos of horseshoe crabs have been induced by grafting. The fact may mean that this type of embryonic induction is widespread in the animal kingdom.     

Thermal biology of horseshoe crab embryos and larvae: A role for heat shock proteins

Eggs of the American horseshoe crab, Limulus polyphemus L., develop on sandy estuarine beaches during the spring and summer, and are potentially vulnerable to thermal stress during the 3-4 weeks of development to the first instar (trilobite) larval stage. In many marine taxa, heat shock (stress) proteins (Hsp's) help individuals acclimate to stresses by restoring the proper folding of cellular proteins whose shape has been altered by temperature shock or other forms of environmental stress. We examined the survival of embryos and first instar (trilobite) larvae following heat shock, and compared the levels of Hsp70 in heat shocked and control animals. Animals acclimated to 13 or 22 °C had close to 100% survival when heat shocked for 3 h at 35 or 40 °C, but exposure to 45 °C for 3 h was lethal. To study the effect of heat shock on Hsp70 production under environmentally realistic conditions, animals were acclimated to either 13 or 22 °C, heat-shocked at 35 °C for 3 h, and soluble proteins were extracted following 0, 2, 4, or 6 h recovery at 22 °C. The relative amounts of Hsp70 in horseshoe crab embryos and larvae were examined using SDS-PAGE and Western blotting. Relative to controls animals held at a constant temperature, there was a slight elevation of Hsp70 only among heat shocked trilobite larvae in the 6 h recovery treatment. Hsp70 levels did not differ significantly between control and heat shocked embryos. Horseshoe crabs have adapted to living in a thermally stressful environment by maintaining a high baseline (constitutive) level of cellular stress proteins such as Hsp70, rather than by synthesizing inducible Hsp's when stressful temperatures are encountered. This may be an effective strategy given that the heat shocks encountered by intertidal embryos and larvae occur regularly as a function of diurnal and tidal temperature changes.     

Proteins of Developing Rice Embryos As Characterized by Twodimensional Gel Electrophoresis

Developmental changes of rice (Oryza sativa, subsp, japonica) in embryonic proteins during embryogenesis were investigated by modified two-dimensionalpolyacrylamide gel electrophoresis. The results indicated that there were apparently differences in the embryonic proteins between the embryos at 7th and 13th day after anthesis. Some proteins only appeared in the embryos al 7th day, disappearing at 13th day. Then some new proteinsappeared at 21th day embryos, which were different from that disappeared during differentiation. In concomitant with the completion of embryo differentiation, the number of acidic proteins decreased, while the basic ones showed an increasing trend. Itwas also found that in the 7th day embryos, there were a higher relative percent ofembryonic protein spots in the region of higher molecular weight, while in the 13thday there were higher relative percent of ones in the region of lower molecular weight.The electrophoretic pattern of rice germ lectin (RGL) showed that the synthesis ofRGL was associated with embryo differentiation. According to these results, we propose that some of embryonic proteins which areonly found at early stage of embryogenesis may be important factors for the regulationof embryo differentiation. Although the function of these proteins is still an openquestion, these specific proteins, at least, represent an excellent mark for plant embryodifferentiation.     

Effect of the Glutamine-Analogue "Azaserine" on Embryonic Development of the Horseshoe Crab

Embryos of the horseshoe crab were treated with the glutamine-analogue "azaserine". This compound induced malformed embryos at the stage of enlargement of the germ disc. The embryonic areas of these monsters were separated into two pieces. This is the first report of monsters induced with azaserine.
The effect of azaserine was counteracted by glutamine.
Azaserine repressed cell growth of the embryos, and prevented development of their germ disc. During active morphogenic movement, the embryonic area of treated embryos was separated in the central region of elongation of the embryonic area.     

Induction of multiple embryos with NaHCO3 or calcium free sea water in the horseshoe crab

Summary When horseshoe crab embryos were treated with NaHCO₃ at the developmental stage when the germ disc appears, multiple embryos were formed. NaHCO₃ may effect the formation of multiple embryos by binding Ca²⁺ ions of the embryo since multiple embryos were also formed by treatment with Ca²⁺ free sea water.The treatment caused the blastoderm layer to tear. When the embryos were returned to normal sea water after the treatment, the blastoderm recovered. Some cell masses, probably derived from the germ disc or its prospective cells, formed during the process of the recovery. Each cell mass developed into an embryo.Contributions from the Shimoda Marine Research Center, Nor. 348     

Purification, characterization, and amino acid sequence of an embryonic lectin in perivitelline fluid of the horseshoe crab

Hemagglutinating activity in perivitelline fluid of the horseshoe crab embryo dramatically increases during the third and fourth molt of the embryo. A 27-kDa lectin, which we named tachylectin-P (TL-P), was newly identified in perivitelline fluid of the horseshoe crab Tachypleus tridentatus. TL-P preferentially agglutinated human A-type erythrocytes, and the activity was inhibited by N-acetyl group-containing monosaccharides. The amino acid sequence analysis indicated that TL-P is almost structurally the same as a hemocyte-derived lectin with no hemagglutinating activity, tachylectin-1 (TL-1), and that 218 out of 221 amino acid residues in total were conserved between the two lectins. Despite the high sequence similarity, biological and biochemical characteristics of TL-P differed from those of TL-1: (i) unlike TL-1, TL-P agglutinates several animal-derived erythrocytes; (ii) unlike TL-1, TL-P has no significant affinity for bacterial lipopolysaccharides or antibacterial activity; (iii) Based on apparent molecular masses determined by gel filtration, TL-P forms a dimer in solution, while TL-1 is present as a monomer; (iv) and TL-P interacts with endogenous proteins of 13 and 14 kDa present in the perivitelline fluid; however, neither has any affinity for TL-1. We propose that TL-P may have an important role in completing embryonic development by interacting with endogenous glycoproteins or N-acetylhexosamines.     

'Early' mammalian myoblasts are resistant to phorbol ester-induced block of differentiation

Mesenchymal cells were isolated from somites and limbs of mouse embryos at different developmental stages. When grown in tissue culture, some of the cells underwent muscle differentiation as indicated by synthesis of sarcomeric myosin, acetylcholine receptor and, in the case of limb cells, fusion into multinucleated myotubes. When the tumour promoter 12-O-tetradecanoyl phorbol 13-acetate (TPA) was added to these cultures, it caused differential effects, depending upon the age of the embryo from which cells were isolated. In cultures of somites or limb bud from embryos up to 12 days post coitum, TPA did not interfere with the appearance of differentiated muscle cells. When TPA was added to cultures from older embryos, it inhibited muscle differentiation with an efficiency which increased with the age of the embryo, reaching about 90% inhibition at 15 days. After this period, a new population of myogenic cells appeared in the limb, which were able to differentiate in the presence of TPA and represented the great majority of myoblasts after day 18 of embryonic development. The simplest interpretation of these data can be based on the existence of three major classes of myogenic cell precursors, which appear sequentially during muscle histogenesis: 'early' myoblasts, which appear resistant to tumour promoters; 'late' myoblasts, whose differentiation is inhibited by tumour promoters and 'satellite' cells which, like early myoblasts, show no sensitivity to TPA.     

鸡胚巨噬细胞AcP酶变化及AcP酶与凋亡细胞的关系

本研究采用电镜及酶细胞化学的方法观察了鸡胚脾脏不同胚龄组巨噬细胞溶酶体酸性磷酸酶（AcP酶）的变化、凋亡实验组巨噬细胞及其AcP酶与凋亡细胞的关系。取10天、13天和17天鸡胚脾脏,按Gomori法显示AcP酶,各胚龄脾脏巨噬细胞AcP酶细胞化学反应阳性,按AcP酶染色阳性做溶酶体计数,结果显示随着胚龄的增加溶酶体数随之增加,尤以第17天组溶酶体数增加最为明显,所得数据经统计分析表明各胚龄组间溶酶体数的差异有统计学意义。凋亡实验组采用放线菌酮诱导15天鸡胚脾脏细胞凋亡,结果显示凋亡细胞为各类幼稚血细胞,以幼稚淋巴细胞为主。巨噬细胞未见凋亡,而是吞噬了大量的凋亡细胞和凋亡小体,AcP酶反应颗粒不仅出现在巨噬细胞的溶酶体、吞噬体,还见于高尔基复合体、内质网等。细胞AcP酶反应强度数字化结果表明：凋亡组酶活性显著高于对照组,差别有统计学意义,提示胚胎巨噬细胞在凋亡细胞出现时AcP酶活性增强,说明巨噬细胞吞噬和消化凋亡细胞或凋亡小体是通过AcP酶等活性物质来实现的。