Immunocytochemical localization of neuropeptide Y, serotonin, substance P and β-endorphin in optic ganglia and brain of Metapenaeus ensis

1 INTRODUCTION Amines and peptides are important bioactive substance in crustaceans, and they function as hor- mones, neurotransmitters and neuromodulators. By far, bioactive substances that have been detected from nervous system of crustaceans include se…     

Nemerteans of Zhoushan Islands,East China Sea Coast,China I. Enopla: Distromatophynchocoela: Monostilifera: Emplectonemertidae

Two nemerteans of the Zhoushan Islands,Paranemertes sinensis sp. nov. andP. peregrina, are reported in this paper.P. sinensis sp. nov. has the body wall longitudinal musculature anteriorly divided, numerous eyes grouped into four clusters, two pairs of cephalic furrows, rhynchocoel 1/3 to 1/2 of full body length, cephalic glands well developed and posteriorly extending beyond the cerebral ganglia, precerebral septum absent, cerebral sense organs situated in front of brain, three pairs of nephridiopores, and 17–18 proboscis nerves. The project supported by National Natural Science Foundation of China     

Primary molecular basis of androgenic gland endocrine sex regulation revealed by transcriptome analysis in Eriocheir sinensis

In crustaceans, the male sexual differentiation and maintenance are specially regulated by androgenic gland (AG). However, little is known about the genes involved in the regulation process. RNA-Seq was performed on AG with ejaculatory duct (AG_ED) and ejaculatory duct (ED) as control in Eriocheir sinensis, one of the most important economic and fishery crabs with typically sex dimorphism. A total of 925 unigenes were identified as differentially expressed genes (DEGs) and the expression of nine genes randomly selected was confirmed by qRT-PCR. 667 unigenes were up-regulated in AG_ED, being supposed to be AG preferential genes. Among them, the full length of insulin-like androgenic gland factor (IAG) cDNA named as Es-IAG was obtained as a logo gene of AG, which together with the genes insulin-like receptor (INR), and single insulin binding domain protein (SIBD), might constitute the sex regulation pathway. Several sex related genes were identified, and their function will have to be investigated. Also, the identification of juvenile hormone epoxide hydrolase 1 (JHEH1), ecdysteroid 22-hydroxylase (DIB) and ecdysone receptor (ECR) preliminarily clarified the molecular regulation mechanism of eyestalk-AG-testis axis, which plays important roles in molting and reproduction. The results will enhance our understanding for the molecular basis of the AG involved in male sex regulation in crabs.

     

Toxic effects of several phthalate esters on the embryos and larvae of abalone Haliotis diversicolor supertexta

As the most widely used plasticizers in the world, phthalate esters (PAEs) are potential endocrine disruption compounds (EDCs). In the present study, the toxicity of dimethyl phthalate (DMP), diethyl phthalate (DEP), dibutyl phthalate (DBP), di (2-ethylhexyl) phthalate (DEHP) on embryogenesis and larvae development of the marine univalve Haliotis diversicolor supertexta was examined in laboratory. The results show that the malformation of embryos appeared during the experiment, such as embryos died or lysed, small transparent flocculent rings studded on the periphery of the embryo, and the larvae could failed to hatch. In embryo toxic test, embryos incubated at the highest concentration of DMP, DEP and DBP solutions showed significantly high abnormal rate compared with the control, while DEHP solutions displayed no significant difference. In larval toxic test, in all concentrations of DMP, DEP and DBP solutions, larval settlement rates were low significantly than that of the control. Similarly, DEHP solutions show nearly no effect on the larval settlement. The order of toxicity on embryos and larvae is DBP>DEP>DMP>DEHP. Being a simple and easy stimulation to indoor spawn, sensitive to environmental factors, and short culture time, the embryos of H. diversicolor supertexta can be used to indicate toxicity of the PAEs. Supported by National Natural Science Foundation of China (No. 40476057)     

Effect of lead on cytoskeletal protein stability in crucian carp <Emphasis Type="Italic">Carassius auratus</Emphasis>

Inorganic lead （Pb） is one of the most common environmental pollutants. Much evidence indicates that Pb exposure could directly affect fish growth and development. In this study, we investigated the cytotoxic effects of Pb on cytoskeletal protein stability at both protein and mRNA level in crucian carp Carassius auratus. Pb（NO3）2 treatment in concentration of 100 μmol/L resulted in decreased expression of both α- and β-tubulin but γ-tubulin as assayed with SDS-PAGE, Western Blot, and ELISA. In vivo and in vitro analyses on protein expression of tubulins are consistent. The effect of Pb on mRNA expression varied among different tissues. Our results suggest that cytotoxicity of Pb at protein translation level is stronger than at mRNA expression level.     

Non-specific immune response of bullfrog <Emphasis Type="Italic">Rana catesbeiana</Emphasis> to intraperitoneal injection of bacterium <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>

Non-specific immune response of bullfrog Rana catesbeiana to pathogenic Aeromonas hydrophila was studied to 60 individuals in two groups. Each bullfrog in bacterium-injected group was injected intraperitoneally （i.p.） with 0.2 ml bacterial suspension at a density of 5.2 × 10^6 CFU/ml, while each one in control group injected i.p. with 0.2 ml sterile saline solution （0.85%, w/v）. Three bullfrogs in both groups were sampled at 0, 1, 3, 7, 11, 15 and 20 days post-injection （dpi） for the evaluation of non-specific immune parameters. It was observed that intraperitoneal injection of A. hydrophila significantly increased the number of leucocytes and that of NBT-positive cells in peripheral blood. Significant increases in serum bactericidal activity and serum acid phosphatase activity were also observed in the bacterium-injected frogs when compared with those in the control group. However, a significant reduction was detected in vitro in phagocytosis activity of peripheral blood phagocytes. No significant difference in changes in the number of peripheral erythrocytes, serum superoxide dismutase （SOD） activity, and lysozyme activity was detected between the two groups. It is suggested that bullfrogs may produce a series of non-specific immune reactions in response to the A. hydrophila infection.     

Molecular identification,expression pattern,and in-vitro bioactivity analysis of insulin-like growth factor 2 in olive flounder Paralichthys olivaceus

Insulin-like growth factors (IGFs) are key regulators of development and growth. Here, we characterized the igf2 gene from olive flounder (Paralichthys olivaceus) and determined its temporal and spatial expression. We set up an in-vitro protein expression system in eukaryotic human embryonic kidney (HEK293T) cells and explored its effects on cell proliferation. The flounder igf2 cDNA contained a 648-bp open reading frame (ORF) encoding a protein of 215 amino acids (aa), which spanned the complete signal peptide (47 aa), mature peptide (70 aa), and E domain (98 aa). In adult flounder, igf2 mRNA was detected in all selected tissues. In early development, igf2 mRNA was detected throughout development from unfertilized eggs to hatching-stage embryos. In-situ hybridization analysis indicated that igf2 mRNA was specially expressed in the brain region, floor plate, hypochord, otic vesicle, and pectoral fin during embryogenesis. Western blotting analysis indicated that the soluble recombinant flounder IGF2 protein was successfully produced through eukaryotic expression in HEK293T cells. In addition, the recombinant IGF2 protein significantly promoted the proliferation of human cervical carcinoma (HeLa) and HEK293T cells. These results provide new information about the structural and functional conservation, expression patterns, and biological activity of the igf2 in teleosts.

     

The polypeptide in Chlamys farreri can protect human dermal fibroblasts from ultraviolet B damage

To investigate the effect of polypeptide fromChlamys farreri (PCF) on NHDFin vitro, we modeled oxidative damage on normal human dermal fibroblasts (NHDF) exposed to ultraviolet B (UVB). In this study, 3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) were tested to measure cell viability. Enzymes including superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), catalase (CAT) and xanthine oxidase (XOD) were determined biochemically. Total antioxidative capacity (T-AOC) and anti-superoxide anion capacity (A-SAC) were also determined. Ultrastructure of fibroblasts was observed under transmission electron microscope. The results showed that: UVB (1.176×10⁻⁴ J/cm²) suppressed the growth of fibroblasts and the introduction of PCF (0.25%–1%) before UVB reduced the suppression in a concentration-dependent manner. PCF could enhance the activities of SOD, GSH-PX and T-AOC as well as A-SAC. Also PCF could inhibit XOD activity, while it did not affect CAT activity. Ultrastructure of fibroblasts were damaged after UVB irradiation, concentration-dependent PCF reduced the destructive effect of UVB on cells. These results indicated that PCF can protect human dermal fibroblasts from being harmed by UVB irradiation via its antioxidant proerty. This work was supported by the National Natural Science Foundation of China (No. 39970638) and the Science and Technology Bureau of Qingdao (No. 2001-28-50).     

Retrieval of chlorophyll a and suspended solid concentrations by hyperspectral remote sensing in Taihu Lake, China

Chlorophyll α（ch1-α） and suspended solid concentrations are two frequently used water quality parameters for monitoring a lake. Traditional measurement of ch1-α and suspended solids, requiring laborious laboratory work, which is often expensive and time consuming. Hyperspectral remote-sensing measurement provides a fast and easy tool for estimating water trophic status. In situ hyperspectral data on March 7-8, July 6-7, September 20 and December 7-8, 2004 and the corresponding water chemical data were used to regress the algorithm of water quality parameters. Results showed that the peak of water leaving radiance around 700 nm （R700） varied proportionally with ch1-α concentration, and moved to infrared when algal bloom occurred. The reflectance ratio of R702/R685 was well correlated with ch1-α when water surface in no algal bloom case and the correlation coefficient was better if absorption of phycocyanin was considered. The reflectance ratio R620/R531 was highly correlated to the concentration of suspended solids. The relationship between suspended solids and other band groups were also compared. Secchi disk depth could be calculated by non-linear correlation with suspended solids concentration.     

The hydrogen peroxide impact on larval settlement and metamorphosis of abalone <Emphasis Type="Italic">Haliotis diversicolor supertexta</Emphasis>

Abalone Haliotis diversicolor supertexta is an important economic mollusk. The settlement and metamorphosis are two critical stages during its development period, which has direct influence on abalone survival and production. The influence of reactive oxygen species （hydrogen peroxide） on abalone embryo and juvenile development were examined in this study. Larvae of Haliotis diversicolor supertexta were induced to settlement and metamorphose by exposure to seawater supplemented with hydrogen peroxide. They had the best performance at 800 μmol/L. The concentration of 1 000 μmol/L or higher was toxic to the larvae, as the larvae could settle down only at benthic diatom plates without complete metamorphosis. In addition, H2O2 adding time was critical to the larval performance. 24h after two-day post-fertilization was proved to be the optimal adding time. In this paper, two action mechanisms of hydrogen peroxide are discussed： （1） hydrogen peroxide has direct toxicity to ciliated cells, thus cause apoptosis; （2） hydrogen peroxide, as a product from catecholamines＇ autoxidation process in vivo, can reverse this process to produce neuro-transmitters to induce abalone metamorphosis.     

Antiangiogenic activity of low-temperature lysozyme from a marine bacterium <Emphasis Type="Italic">in vivo</Emphasis> and <Emphasis Type="Italic">in vitro</Emphasis>

We extracted marine low-temperature lysozyme (MLTL), a novel lysozyme, from a marine microorganism through fermentation. Our previous study suggested that a low molecular weight (16 kDa) may exert anti-tumor activity through antiangiogenesis. In this study, we extracted a high weight (39 kDa) and investigated its antiangiogenic activity in vivo and in vitro. Using zebrafish embryos as an in vivo study model, we found that treatment with MLTL significantly inhibited the growth of subintestinal vessels (SIVs) in a dose-dependent manner and that 400 μg/ml MLTL was sufficient to block the growth of SIVs. An in vitro study conducted using human umbilical vein endothelial cells (HUVECs) revealed that MLTL suppressed the proliferation, migration and tube formation of HUVECs in a dose-dependent manner. Interestingly, assays by flow cytometry and DNA electrophoresis indicated that MLTL was able to induce apoptosis of HUVECs. Moreover, further study demonstrated that the disruption of intracellular Ca²⁺ homeostasis may play an important role in MLTL induced apoptosis of HUVECs. Taken together, the results of this study demonstrate for the first time that MLTL inhibits angiogenesis through its pleiotropic effects on vascular endothelial cells and induces apoptosis through regulation of cellular Ca²⁺ levels. The results of this study also revealed a possible mechanism underlying the antiangiogenic effect of MLTL and suggested that MLTL may be a promising new antiangiogenic agent for use in cancer therapy.     

Effects of carboxymethyl-chitosan on wound healing in vivo and in vitro

Effects of carboxymethyl-chitosan with different molecular weights on wound healing were investigated in vivo and in vitro.A second degree burn model was performed in rats and the accelerative effects of carboxymethyl-chitosan on wound repair were observed.Contents of transforming growth factor(TGF)-β1,interleukin(IL)-6 and matrix metalloproteinase(MMP)-1 in wounds were determined by enzyme-linked immunosorbent assay(ELISA).In vitro study evaluated the influence of carboxy-methyl-chitosan on cytokines secretion of fibroblasts and macrophages.In vivo results showed that carboxymethyl-chitosan effec-tively accelerated the wound healing process in burned rats(P<0.05).Levels of TGF-β1,IL-6 and MMP-1 in carboxymethyl-chitosan groups were significantly elevated,compared with control group(P<0.05).In vitro results indicated that carboxymethyl-chitosan significantly promoted the proliferation of fibroblasts and stimulated its IL-8 and IL-10 secretion at different incubation time,but it did not affect collagen secretion of fibroblasts.Carboxymethyl-chitosan enhanced phagocytosis ability of macrophages,and in-creased its tumor necrosis factor(TNF)-α secretion.In conclusion,carboxymethyl-chitosan promoted wound healing by activating macrophages,accelerating fibroblasts growth and exerting considerable effects on the secretion of a series of cytokines.     

Safety analysis of water resources and eco-environment in Shiyang River Basin

The research on the present situation of soil and water development and utilization in Shiyang River Basin shows that water resources and eco-environment situation in this area are near the edge of collapse. Since the water crises occurred in the 1970s, problems caused by continuous decrease of water resources have been becoming serious year by year and eco-environment crisis occurred as a consequence. Up to now, 10 380ha of irrigated lands have been abandoned due to sand coverage and water shortage in the basin. Ground water was over exploded in Wuwei and Minqin because of water shortage. Ground water table in many places dropped under 5m (which is the ecology water table level), thus about 3000ha of Elaeagnus angustifolia forest come to dead and another 5800ha become feeble, and wind-drift sand near the oasis become alive. According to the current situation, if water utilization scope was not enlarged, a water transfer volume of 600×10⁶m³/a from other areas will be suitable to keep water resources and eco-environment safety in the basin, and also 70×10⁶m³/a will be left as spare water. Under this condition the water resources and eco-environment of the basin can reach the critical safety line of 2.032×10⁹m³/a; or if 180×10⁶m³ of water can be transferred from other areas, the water resources can reach the safety warning line of 1.732×10⁹m³/a. Foundation item: Under the auspices of the National Natural Science Foundation of China (No. 40235053) and Lanzhou Jiaotong University "Qinglan" Foundation Biography: ZHANG Ji-shi (1963-), male, a native of Tongwei of Gansu Province, senior engineer, specialized in water resources and climate change in Northwest China. E-mail: zjs1963@yahoo.com.cn; hangjs@mail.lzjtu.cn     

The insemination of goldfish (Carassium auratus) oocyte maturedin vitro

Full maturation of goldfish oocyte was inducedin vitro by 17 α-hydroxy-20β-dihydroprogesterone. The oocyte maturation involves GV migration to the periphery of the oocyte and germinal vesicle breakdown (GVBD). In the experiment, incubation duration for GVBD varied in different broods of oocytes. Generally, if the duration for GVBD was shorter than 6 h, oocytes would have a better chance to survive after maturation and insemination. The maturation of nucleus (GV) and cytoplasm are not synchronous. Cytoplasm maturation occurs several hs after GVBD. Oocytes inseminated 8–9 h after GVBD have the highest fertilizing and hatching rate. Fertilized ova maturedin vitro can develop to sexually mature adults capable of reproduction. Contribution No 1731 from the Institute of Oceanology, Academia Sinica.     

Chemical constituents of marine medicinal mangrove plant <Emphasis Type="Italic">Sonneratia caseolaris</Emphasis>

Twenty-four compounds including eight steroids (1–8), nine triterpenoids (9–16, 24), three flavonoids (20–22), and four benzenecarboxylic derivatives (17–19, 23) were isolated and identified from stems and twigs of medicinal mangrove plant Sonneratia caseolaris. The structures of the isolated compounds were determined by extensive analysis of their spectroscopic data. Among these metabolites, compounds 1, 4–20 and 22–24 were isolated and identified for the first time from S. caseolaris. In the in vitro cytotoxic assay against SMMC-7721 human hepatoma cells, compound 21 (3′,4′,5,7-tetrahydroxyflavone) exhibited significant activity with IC₅₀ 2.8 μg/mL, while oleanolic acid (14), 3,3′-di-O-methyl ether ellagic acid (18), and 3,3′,4-O-tri-O-methyl ether ellagic acid (19) showed weak activity. None of these compounds displayed significant antibacterial activites. Supported by the National Natural Science Foundation of China (No. 30770234); Knowledge Innovation Program of Chinese Academy of Sciences (KZCX2-YW-211-04); Department of Science and Technology of Shandong Province (No.2006GG2205023)     

Paleo-environmental reconstruction during the period of Nanjing Homo Erectus

Using five-group dating of the fossil by Uranium series age method shows that the age of Nanjing Homo Erectus is between 280–430 ka B. P. Combining with the analysis of fossil mammalian form Stegodon, Sus. Lydekkeri Zdansly, M. pachyosteus Young, P. grayi Zdansky and E. SanmeniensisTeilhard et Pive teau, which are located in the same stratum, the authors thinks that the Nanjing Hmo Erectus may have lived in the late Mid-Pleistocene (about 350 ka B. P.). Based upon the spores and pollen in the cave sediments of the area, we can find that most of spores and pollen are subtropical and warm temperature types. For instance, broad leaf trees which are fond of warm climate, including Cunning hamia, Jsuga and Pinus, as well as Morus, Carpinus and Ptendium. Among the above spores and pollen, only Tsuga, Liquidambar, Betula, Ulmus and Salix are the remainders of Humid-hot environment from the Tertiary period. Above evidences illustrate that the living environment of the Nanjing Homo Erectus not only was not cloder but also was not drier than the Peking Man’s. It could be said that dry climate and the ice age of the Mid-Pleistocene didn’t undergo this area. As for the manner of the Homo Erectus’s making stone artifact and using fire, because now we have not found the evidence of the stone artifact and the ash, the futher excavation to the talus of the Huludong cave should be needed. Foundation item: Under the auspices of the National Natural Science Foundation of China (49771075) and Testing Foundation by the Modern Analysis Center of Nanjing University. Biography: ZHU Cheng (1954–), male, professor. His major research interests include environmentul evolution and global changes.     

Free radical scavenging abilities of polypeptide from Chlamys farreri

1 INTRODUCTION Active oxygen free radicals can damage cell structure, even kill the cells, causing aging and cer- tain serious diseases (Bors et al., 1989). Usually, the production and scavenging of active oxygen free radicals are in balance in healthy hu…     

The preparation and antioxidant activity of glucosamine sulfate

Glucosamine sulfate was prepared from glucosamine hydrochloride that was produced by acidic hydrolysis of chitin by ion-exchange method. Optical rotation and elemental analysis characterized the degree of its purity. In addition, the antioxidant potency of chitosan derivative-glucosamine sulfate was investigated in various established in vitro systems, such as superoxide (O-2)/hydroxyl (·OH) radicals scavenging, reducing power, iron ion chelating. The following results are obtained: first, glucosamine sulfa...     

A sulfotransferase specific to N-21 of gonyautoxin 2/3 from crude enzyme extraction of toxic dinoflagellate Alexandrium tamarense CI01

Sulfotransferase (ST) is the first enzyme discovered in association with paralytic shellfish poisoning (PSP) toxin biosynthesis in toxic dinoflagellates. This study investigates the ST activity in crude enzyme extraction of a toxic dinoflagellate species, Alexandrium tamarense CI01. The results show that crude enzyme can transfer a sulfate group from 3’-phosphoadenosine 5’-phosphosulfate (PAPS) to N-21 in the carbamoyl group of gonyautoxin 2/3 (GTX2/3) to produce C1/C2, but is inactive toward STX to produce GTX5. The crude enzyme is optimally active at pH 6.0 and 15°C. The activity is enhanced by Co2 , Mg2 , Mn2 and Ca2 individually, but is inhibited by Cu2 . Moreover, the activity shows no difference when various sulfur compounds are used as sulfate donors. These results demonstrate that the ST specific to GTX2/3 is present in the cells of A. tamarense CI01 and is involved in PSP toxin biosynthesis. In addition, the ST from different dinoflagellates is species-specific, which explains well the various biosynthesis pathways of the PSP toxins in toxic dinoflagellates.