Inhibition of Fumonisin B1 Cytotoxicity by Nanosilicate Platelets during Mouse Embryo Development

Nanosilicate platelets (NSP), the form of natural silicate clay that was exfoliated from montmorillonite (MMT), is widely used as a feed additive for its high non-specific binding capacity with mycotoxins such as fumonisin B₁ (FB₁), and has been evaluated its safety for biomedical use including cytotoxicity, genotoxicity, and lethal dosage (LD). In the study, we further examined its toxicity on the development of CD1 mouse embryos and its capacity to prevent teratogenesis-induced by FB₁. In vitro cultures, NSP did not disturb the development and the quality of intact pre-implantation mouse embryos. Further, newborn mice from females consumed with NSP showed no abnormalities. NSP had an unexpected high adsorption capacity in vitro. In contrast to female mice consumed with FB₁ only, a very low residual level of FB₁ in the circulation, reduced incidence of neutral tube defects and significantly increased fetal weight were observed in the females consumed with FB₁ and NSP, suggesting a high alleviation effect of NSP on FB₁ in vivo. Furthermore, FB₁ treatment disturbed the gene expression of sphingolipid metabolism enzymes (longevity assurance homolog 5, LASS 5; sphingosine kinase 1, Sphk1; sphingosine kinase 2, Sphk2; sphingosine 1- phosphate lyase, Sgpl1; sphingosine 1-phosphate phosphatase, Sgpp1) in the maternal liver, uterus, fetus, and placenta, but NSP administration reversed the perturbations. Based on these findings, we conclude that NSP is a feasible and effective agent for supplementary use in reducing the toxicity of FB₁ to animals.     

发育中玉米胚结合蛋白(BiP)的动态变化及定位

Western blot检测表明,在玉米胚发育过程中结合蛋白(BiP)含量与胚可溶性蛋白含量变化一致,在授粉16d后BiP含量随发育而增加;对热激不敏感.组织化学免疫定位表明,在玉米胚发育的不同时期,BiP主要定位在胚芽端、初生维管组织和糊粉层中,提示胚在构建器官的同时,也为其功能执行准备了条件;热激不影响其定位.     

Mitochondrial Changes in Platelets Are Not Related to Those in Skeletal Muscle during Human Septic Shock

Platelets can serve as general markers of mitochondrial (dys)function during several human diseases. Whether this holds true even during sepsis is unknown. Using spectrophotometry, we measured mitochondrial respiratory chain biochemistry in platelets and triceps brachii muscle of thirty patients with septic shock (within 24 hours from admission to Intensive Care) and ten surgical controls (during surgery). Results were expressed relative to citrate synthase (CS) activity, a marker of mitochondrial density. Patients with septic shock had lower nicotinamide adenine dinucleotide dehydrogenase (NADH)/CS (p = 0.015), complex I/CS (p = 0.018), complex I and III/CS (p<0.001) and complex IV/CS (p = 0.012) activities in platelets but higher complex I/CS activity (p = 0.021) in triceps brachii muscle than controls. Overall, NADH/CS (r² = 0.00; p = 0.683) complex I/CS (r² = 0.05; p = 0.173), complex I and III/CS (r² = 0.01; p = 0.485), succinate dehydrogenase (SDH)/CS (r² = 0.00; p = 0.884), complex II and III/CS (r² = 0.00; p = 0.927) and complex IV/CS (r² = 0.00; p = 0.906) activities in platelets were not associated with those in triceps brachii muscle. In conclusion, several respiratory chain enzymes were variably inhibited in platelets, but not in triceps brachii muscle, of patients with septic shock. Sepsis-induced mitochondrial changes in platelets do not reflect those in other organs.     

Types of Neural Tissues Induced through the Presumptive Notochord of Newt Embryo

Neural induction through the presumptive notochord was tested by means of the sandwich method. The result disclosed that the notochord was a potent inducer of neural tissues not only in the ectoderm of gastrula but also in the ventral ectoderm of neurula and early tail-bud embryos. Structures formed by the induced neural tissue varied greatly. They can be classified into three types. (1) Tubular: the neural tissues induced in explants containing abundant mesenchymes always formed long tubular structures. The shapes of these neural tubes showed considerable variation; moreover, they were atypical and none formed the regular structure of the spinal cord. This type was most frequent, being found in about 50% of the explants. (2) Inverted: this type was produced when the explant contained mesenchymal component. Consequently, the epithelium of explants was missing. Nevertheless, a considerable mass of neural tissue was always induced. It was noticed that the induced neural tissues were invariably inside out; this type was found in about 30% of the explants. (3) Archencephalic: this was the only type to form the regular structure, i.e., the archencephalon. Formation of the archencephalon was limited solely to those explants containing only a few mesenchymes; this type was found in about 20% of the cases. As described above, it was found that the neural tissues induced by the same inducer of the notochord were not uniform but varied in type. Further, it was shown that the types of neural tissue differed according to different quantities of the surrounding mesenchyme. Based on these facts, it is to be concluded that it is not the inducer of notochord, but the surrounding mesenchyme that is of primary importance for the determination of the types of neural tissue.     

Transmission of Homoiogenetic Induction in Presumptive Ectoderm of Newt Embryo

Using newt, Cynops pyrrhogaster , embryos, the production and/or transmission of a homoiogenetic effect of the induction within a presumptive ectoderm was investigated. The primary inductor was the swimbladder of a crucian carp, Caracius auratus. In the first experiment, a piece of presumptive ectoderm was isolated from an early gastrula and one-third of its inner surface was placed in contact with the swimbladder for 30 min. After removal of the inductor, the ectodermal piece was allowed to stand by itself for several hours (pre-cultivation), and then it was divided into three parts of equal size, i.e. the end that had been placed in contact with the swimbladder (P1), the next middle (P2) and the end part (P3).
The tissues produced in each part were examined after 10 days' cultivation in Holtfreter's solution. The induction was evoked not only in P1, but also in P2 and P3 which had been free from the inductor. The incidences of tissue differentiation in P2 and P3 increased with the lapse of the pre-cultivation time, and the rise in P3 came after P2. These results suggested that the mesodermal tissues in the parts not placed in contact with the swimbladder (P2 and P3) were evoked by the homoiogenetic stimulus which came from P1 and P2, respectively.
In the next experiment, P2 of the ectodermal piece was substituted by an aged ectoderm which had lost its primary competence. In this system, the mesodermal induction was not evoked in P3. This suggested that the production of the homoiogenetic activity of the ectoderm might be associated with its competenece.     

Localization and Characterization of Lectins in Yolk Platelets of Xenopus Oocytes

The localization and characteristics of yolk platelet lectins (YLs) in Xenopus laevis oocytes were studied with antiserum against cortical granule lectins (CGLs) as a probe. In oocytes at stages I, II and III-IV, specific, immunofluorescent staining for the lectins was observed on the cortical cytoplasm extending about 2, 4 and 20 μm, respectively, from the egg surface. In stage III-IV oocytes, the superficial layer of the yolk platelets was also stained. The cortical cytoplasm included cortical granules, coated pits, coated vesicles, multivesicular bodies and primordial yolk platelets. The YLs were incorporated into the oocytes by endocytosis as demonstrated using gold-labeled YLs. On PAGE, native YLs gave two bands of CGL-like proteins and proteins that appeared as a single diffuse band. The YLs and the CGLs shared antigenicity and hemagglutination activity specific to D-galactoside residues. However, the proteins of the diffuse band had little or no activity for either hemagglutination or jelly-precipitation, suggesting that they were monomers with a single reactive site. These results indicate that the YLs are supplied to the oocytes, presumably from extracellular sources, polymerized to CGL-like molecules in the cortical cytoplasm and accumulated in the superficial layer of the yolk platelets.     

杉木小孢子叶球发育过程中形态结构变化

以扬州地区生长的成年杉木为实验材料,通过采用数码相机拍摄、体视镜、扫描电镜以及半薄切片等方法,对杉木小孢子叶球发育、小孢子囊发育及其散粉规律进行详细观察。结果发现,10月中旬,杉木小孢子叶球形成并着生于新枝顶端。翌年3月中下旬,小孢子叶球成熟,进入散粉期,散粉首先从小孢子叶球基部开始依次向上部扩散。小孢子叶在孢子叶球主轴上螺旋排列,单个小孢子叶通常由3个小孢子囊组成,构成三角形。小孢子囊壁由1层表皮细胞、1~2层中间层细胞和1层绒毡层细胞组成,表皮细胞首先形成,并向内分化出2~3层细胞,分别分化形成中层和绒毡层,最后中层和绒毡层消失。杉木的小孢子囊通过开裂口控制其开裂,20℃室温下整个小孢子叶球散粉时间持续约18 h,单个小孢子叶的散粉时间为8~10 h。以上结果显示,杉木小孢子叶球的发育过程经历了体积增大、鳞片开张及散粉等阶段,这些形态和结构上的变化利于提高散粉效率,这说明杉木在长期的演化过程中,形成了许多有利于风媒传粉的结构特征。     

Isotopic Labeling of Embryo, Yolk Sac, and Intracellular Parasites of the Embryonated Hen''s Egg by Yolk Replacement

A technique is described which allows the replacement of 50% of the yolk of the embryonated hen's egg with large volumes of diverse but chemically defined solutions. By using an electrosurgical unit and a polyethylene tunnel, the procedure was performed on eggs from days 3 through 7 with greater than 90% surgical success and viability for the short term. More than 50% of the eggs replaced showed viability for 2 weeks, and a significant proportion went full term. ³²PO₄ and amino acids (³H and ¹⁴C) added to the replaced eggs were incorporated into the macromolecules of the embryo and yolk sac as well as into parasitic rickettsiae cultivated in the replaced eggs. The incorporated ³²PO₄ was shown to be assimilated into a variety of biochemical species.     

Biochemical Studies during the Development of the Chick Embryo

The presence and the change of deoxyribosidic compounds in the acid extract of the embryo with the incubation were examined with an aid of the organism, L. leichmannii. The main deoxyribosidic compounds in the extract prepared from the 18 th day embryo were identified as uracil, cytosine and thymine deoxyribosides and deoxyribotides of cytosine and thymine from the behaviour on paper chromatographic and paper electrophoretic separation. A small amount of purine deoxyribosyl compound which was assumed as hypoxanthine deoxyriboside was detected, and the content of which per 1 g of fresh embryo changed with the lapse of the incubated day; especially, the content was minimum at the period from the 10 th to 15 th day incubation. At this period, the total growth promoting compounds contained 50% of deoxyribotide though deoxyribosides was lower than that of the other days. This period is the most significant stage of the embryo growth and the most active time of synthesis of DNA through the embryo growth.     

Biochemical Studies during the Development of the Chick Embryo

The devised method consists of the enzymatic hydrolysis, separation of deoxyribosides in the hydrolysate by paper chromatography and paper electrophoresis, and the estimation of the separated fractions with L. leichmannii. This method permits the determination of deoxyriboside composition of the smaller amounts of DNA or the related compounds with relatively higher accuracy even under the presence of some other compounds when nucleic acids and acid insoluble fractions of the chick embryo were analyzed.

The change of each deoxyriboside composition in acid insoluble fraction prepared from the 3 to 19 day old embryos investigated by the method. Among the major four deoxyribosides, the contents of deoxyguanosine and of deoxycytidine was nearly constant during the development of the embryo, whereas that of thymidine and of deoxyadenosine appeared to undergo the change slightly at the periods from 10 to 15 days incubation. It seems that this periods is also the most active time of the synthesis of DNA and of the changes of deoxyribosyl compounds in acid soluble fraction through the embryo growth.     

Isotopic Labeling of Embryo,Yolk Sac,and Intracellular Parasites of the Embryonated Hen's Egg by Yolk Replacement

A technique is described which allows the replacement of 50% of the yolk of the embryonated hen''s egg with large volumes of diverse but chemically defined solutions. By using an electrosurgical unit and a polyethylene tunnel, the procedure was performed on eggs from days 3 through 7 with greater than 90% surgical success and viability for the short term. More than 50% of the eggs replaced showed viability for 2 weeks, and a significant proportion went full term. ³²PO₄ and amino acids (³H and ¹⁴C) added to the replaced eggs were incorporated into the macromolecules of the embryo and yolk sac as well as into parasitic rickettsiae cultivated in the replaced eggs. The incorporated ³²PO₄ was shown to be assimilated into a variety of biochemical species.     

Biochemical Changes Associated with Zygotic Pine Embryo Development

Johnson, M. A., Carlson, J. A., Conkey, J. H. and Noland, T.L. 1987. Biochemical changes associated with zygotic pine embryodevelopment.—J. exp. Bot. 38: 518–524. At intervals during the period from pre-fertilization to nearmaturation, pine (Pinus resinosa Ait. and Pinus strobus L.)ovules were analysed for several biochemical constituents, andthe results were expressed on a fresh weight basis. Lipid accumulatedin parallel with the growth of the developing seeds. Solubleprotein also accumulated but only in the initial stages of development.ATP content peaked approximately 2 weeks after fertilization,followed about one week later by the energy charge; these peakswere associated with maximal growth stages of the developingembryos. Likewise, peaks of glutathione (GSH) and ascorbic acid(AA), two water-soluble reductants, preceded or coincided withthe ATP maximum. At late stages of seed development, dissectionof the more mature ovules into embryos and gametophytes foranalysis revealed that the ATP, GSH, and AA were more concentratedin the embryonic tissue. On the other hand, this segregationshowed that virtually no proanthocyanidin was located in thedeveloping embryos proper, although they contained other reductants,some of which were probably phenolics. Also, general stainingwith reagents for phenolics and thiols indicated that the formeroccurred primarily in non-embryonic tissue, whereas GSH wasin the embryo per se. These findings are consistent with rolesfor ATP, GSH, and AA in the growth and development of zygoticpine embryos; however, it would appear that lipid and proteinare being stored for subsequent germination events and thatmuch of the phenolic component is segregated from the developingembryo. Key words: Pine, embryogenesis, biochemistry     

醮核荔枝胚胎发育相关蛋白质的分离及初步鉴定

通过二维聚丙烯酰胺凝胶电泳以及计算机辅助的图像分析技术,对荔枝开花后40d的正常与败育胚蛋白质图谱进行初步分析。结果表明,100个蛋白质点在表达丰度上有明显差异;选择仅在正常发育胚胎胶上表达的蛋白点15个和仅在败育胚胎胶上表达的蛋白点50个,进行基质辅助激光解吸附电离飞行时间质谱（MALDI—TOFMASS）分析,鉴定出9个与胚发育相关的蛋白,这些蛋白可能参与了胚败育的调节和控制。     

葡萄胚胎发育与败育过程中胚珠的多胺含量变化

花后40~60d期间,种子败育型无核葡萄胚珠的3种多胺含量明显低于有核葡萄,据此认为无核葡萄胚珠中多胺含量急剧下降和较低的多胺水平可能是导致葡萄胚胎败育的重要原因。(Spd Spm)/Put和Spm/PAs比值变化与葡萄胚胎发育密切相关,较低的比值不利于胚胎的正常分化。     

用DDRT-PCR技术克隆小鼠早期胚胎发育相关基因

mRNA差异显示 (DDRT PCR)技术在哺乳动物早期胚胎发育相关基因研究中的应用 ,因获得足够量的早期胚胎材料困难而受到限制 .通过对DDRT PCR技术各种条件参数进行优化组合 ,并对某些环节进行改良 ,以小鼠的MⅡ卵、2 细胞胚胎和 4 细胞胚胎为材料进行差异显示 ,仅以相当于5 0个卵细胞的量为起始材料 ,便得到了理想的差示结果 .从差异条带中挑取感兴趣的差异条带进行回收、阳性鉴定、亚克隆、序列分析、并在反向Northern杂交基础上设计了鉴定实验 .结果发现 ,有一个片段差异显著且是阶段性特异表达 .经GenBank检索 ,发现该片段仅有同源的EST ,其全长及功能尚不清楚 ,是一个功能未知基因 ,将该片段命名为ed1.反向Northern杂交结果表明 ,ed1在 2 细胞期胚胎中有表达 ,而在MⅡ卵及 4 细胞胚胎中均不表达 .     

Morphological Changes of the Inner Surface of the Blastocoelic Wall before and during Gastrulation in the Newt, Cynops pyrrhogaster

Ultrastructural observations were made of morphological changes in the inner surface of the blastocoelic wall (BW) in newt embryos during the period from morula to gastrula. A scanning electron microscopic investigation distinguished three phases of process formation on the inner surface of the BW. First, small blebs and filopodia were found in the morula. These then changed to large blebs in the blastula and early gastrula. Finally, they acquired lamella-like structures (lamellipodia) in the middle and late gastrula. In addition to the changes in the cell surface, cytoplasmic changes such as of the organization of the cortical microfilamentous layer (CML) were found by transmission electron microscopy (TEM). The morphological changes on the inner surface of the BW and those in the cytoplasmic ultrastructures are considered to be closely related, playing some important roles in the mechanisms of morphogenetic cell movements.     

Changes in Isoperoxidases during Cold Treatment of Dormant Pear Embryo

The number of isoperoxidases and the intensity of certain isozymes increased with increasing periods of stratification of pear (Pyrus communis cv. Bartlett) embryos. The presence of GA₃ or kinetin during stratification enhanced the activity of certain isoperoxidases, and these enhancements were blocked in the presence of ABA which by itself had an inhibitory effect. Enhancement in isoperoxidases of pear embryos during stratification was inhibited by 6-methylpurine and cycloheximide; and in the presence of either of these two inhibitors, stratification failed to release the dormancy. Pear embryos germinated for 3 days showed changes in the pattern of isoperoxidases depending on the length of stratification.