Developmental toxicity and alteration of gene expression in zebrafish embryos exposed to PFOS

Perfluorooetanesulfonate (PFOS) is a persistent organic pollutant, the potential toxicity of which is causing great concern. In the present study, we employed zebrafish embryos to investigate the developmental toxicity of this compound. Four-hour post-fertilization (hpf) zebrafish embryos were exposed to 0.1, 0.5, 1, 3 and 5 mg/L PFOS. Hatching was delayed and hatching rates as well as larval survivorship, were significantly reduced after the embryos were exposed to 1, 3 and 5 mg/L PFOS until 132 hpf. The fry displayed gross developmental malformations, including epiboly deformities, hypopigmentation, yolk sac edema, tail and heart malformations and spinal curvature upon exposure to PFOS concentrations of I mg/L or greater. Growth (body length) was significantly reduced in the 3 and 5 mg/L PFOS-treated groups. To test whether developmental malformation was mediated via apoptosis, flow cytometry analysis of DNA content, acridine orange staining and TUNEL assay was used. These techniques indicated that more apoptotic cells were present in the PFOS-treated embryos than in the control embryos. Certain genes related to cell apoptosis, p53 and Bax, were both significantly up-regulated upon exposure to all the concentrations tested. In addition, we investigated the effects of PFOS on marker genes related to early thyroid development (hhex and pax8) and genes regulating the balance of androgens and estrogens (cyp19a and cyp19b). For thyroid development, the expression of hhex was significantly up-regulated at all concentrations tested, whereas pax8 expression was significantly up-regulated only upon exposure to lower concentrations of PFOS (0.1, 0.5, 1 mg/L). The expression of cyp19a and of cyp19b was significantly down-regulated at all exposure concentrations. The overall results indicated that zebrafish embryos constitute a reliable model for testing the developmental toxicity of PFOS, and the gene expression patterns in the embryos were able to reveal some potential mechanisms of developmental toxicity. (C) 2008 Elsevier Inc. All rights reserved.

Developmental expression of CagMdkb during gibel carp embryogenesis

Midkine (Mdk) genes have been revealed to have different expression patterns in vertebrates and therefore, additional studies on Mdk expression patterns are required in more species. In this study, CagMdkb has been cloned and characterized from a SMART cDNA library of 10-somite stage embryos of Carassius auratus gibelio. Its full length cDNA is 1091 bp and encodes a sequence of 147 amino acids, which shows 97.3% identity to zebrafish Mdkb on the amino acid level. RT-PCR analysis reveals that CagMdkb is first transcribed in gastrula embryos and maintains a relatively stable expression level during subsequent embryogenesis. Western blot analysis reveals a 19 kDa maternal CagMdkb protein band and the zygotic CagMdkb protein is expressed from gastrula stage. At around 10 somite stage, the 19 kDa CagMdkb is processed to another protein band of about 17 kDa, which might be the secreted form with the 21-residue signal peptide removed. With immunofluorescence analysis, maternal CagMdkb protein was found to be localized in each blastamere cell of early embryos. The zygotic CagMdkb positive fluorescence signal was detected from a pair of large neurons at 18-somite stage. At the later stages, CagMdkb protein was also extended to numerous small neurons in the forebrain, midbrain and hindbrain, as well as to nerve fibers in the spinal cord. Co-localization with 3A10 antibody revealed CagMdkb immunoreactivity on developing Mauthner neurons, a member of reticulospinal neurons. In addition, ectopic expression of CagMdkb in early embryos of gibel carp and zebrafish suppressed head formation and CagMdkb function was found to depend on secretory activity. All these findings indicate that CagMdkb plays an important role in neural development during gibel carp embryogenesis and there is functional conservation of Mdkb in fish head formation.

Crustacean zooplankton distribution patterns and their biomass as related to trophic indicators of 29 shallow subtropical lakes

A comparative limnological study was carried out to present a snapshot of crustacean zooplankton communities and their relations to environmental factors to test whether there is a consistent relationship between crustacean biomass and trophic indicators among lake groups with similar trophic conditions. The study lakes showed a wide range of trophic status, with total phosphorus (TP) ranging from 0.008 to 1.448mgL(-1), and chlorophyll a from 0.7 to 146.1 mu g L-1, respectively. About 38 species of Crustacea were found, of which Cladocera were represented by 25 taxa (20 genera), and Copepoda by 13 taxa (I I genera). The most common and dominant species were Bosmina coregoni, Moina micrura, Diaphanosoma brachyurum, Cyclops vicinus, Thermocyclops taihokuensis, Mesocyclops notius and Sinocalanus dorrii. Daphnia was rare in abundance. Canonical correspondence analysis showed that except for four species (D. hyalina, S. dorrii, C. vicinus and M. micrura), almost all the dominant species had the same preference for environmental factors. Temperature, predatory cyclopoids and planktivorous fishes seem to be the key factors determining species distribution. TP was a relatively better trophic indicator than chlorophyll a to predict crustacean biomass. Within the three groups of lakes, however, there was no consistent relationship between crustacean biomass and trophic indicators. The possible reason might be that top-down and bottom-up control on crustaceans vary with lake trophic state. The lack of significant negative correlation between crustacean biomass and chlorophyll a suggests that there was little control of phytoplankton biomass by macrozooplankton in these shallow subtropical lakes. (c) 2007 Elsevier GmbH. All rights reserved.

Expression pattern and developmental behaviour of cellular nucleic acid-binding protein (CNBP) during folliculogenesis and oogenesis in fish

In vertebrates, folliculogeneis establishes an intricate system for somatic cell-oocyte interaction, and ultimately leads to the acquisition of their respective competences. Although the formation process and corresponding interactions are strikingly similar in diverse organisms, knowledge of genes and signaling pathways involved in follicle formation is very incomplete and the underlying molecular mechanisms remain enigmatic. CNBP has been identified for more than ten years, and the highest level of CNBP transcripts has been observed in adult zebrafish ovary, but little is known about its functional significance during folliculogeneis and oogenesis. In this study, we clone CNBP cDNA from gibel carp (Carassius auratus gibelio), and demonstrate its predominant expression in gibel carp ovary and testis not only by RTPCR but also by Western blot. Its full-length cDNA is 1402 bp, and has an ORF of 489 nt for encoding a peptide of 163 aa. And its complete amino acid sequence shared 68.5%-96.8% identity with CNBPs from other vertebrates. Based on the expression characterization, we further analyze its expression pattern and developmental behaviour during folliculogeneis and oogenesis. Following these studies, we reveal an unexpected discovery that the CagCNBP is associated with follicular cells and oocytes, and significant distribution changes have occurred in degenerating and regenerating follicles. More interestingly, the CagCNBP is more highly expressed in some clusters of interconnected cells within ovarian cysts, no matter whether the cell clusters are formed from the original primordial germ cells or from the newly formed cells from follicular cells that invaded into the atretic oocytes. It is the first time to reveal CNBP relevance to folliculogeneis and oogenesis. Moreover, a similar stage-specific and cell-specific expression pattern has also been observed in the gibel carp testis. Therefore, further studies on CNBP expression pattern and developmental behaviour will be of significance for understanding functional roles of CNBP during gametogenests. (c) 2005 Elsevier B.V. All rights reserved.

Retention of the developmental pluripotency in medaka embryonic stem cells after gene transfer and long-term drug selection for gene targeting in fish

Embryonic stem (ES) cells provide a unique tool for introducing random or targeted genetic alterations, because it is possible that the desired, but extremely rare recombinant genotypes can be screened by drug selection. ES cell-mediated transgenesis has so far been limited to the mouse. In the fish medaka (Oryzias latipes) several ES cell lines have been made available. Here we report the optimized conditions for gene transfer and drug selection in the medaka ES cell line MES1 as a prelude for gene targeting in fish. MES1 cells gave rise to a moderate to high transfection efficiency by the calcium phosphate co-precipitation (5%), commercial reagents Fugene (11%), GeneJuice (21%) and electroporation (>30%). Transient gene transfer and CAT reporter assay revealed that several enhancers/promoters and their combinations including CMV, RSV and ST (the SV40 virus early gene enhancer linked to the thymidine kinase promoter) were suitable regulatory sequences to drive transgene expression in the MES1 cells. We show that neo, hyg or pac conferred resistance to G418, hygromycin or puromycin for positive selection, while the HSV-tk generated sensitivity to ganciclovir for negative selection. The positive-negative selection procedure that is widely used for gene targeting in mouse ES cells was found to be effective also in MES1 cells. Importantly, we demonstrate that MES1 cells after gene transfer and long-term drug selection retained the developmental pluripotency, as they were able to undergo induced differentiation in vitro and to contribute to various tissues and organs during chimeric embryogenesis.

Lipid biomarkers in the Zoige-Hongyuan peat deposit: Indicators of Holocene climate changes in West China

IEECAS SKLLQG

Compound specific D/H ratios and molecular distributions of higher plant leaf waxes as novel pale oenvironmental indicators in the Chinese Loess Plateau

IEECAS SKLLQG

Dinoflagellate Cysts as Indicators of Cultural Eutrophication in the Oslofjord, Norway

Dinoflagellate cyst records were analysed from four sediment cores from the inner Oslofjord. The cores covered the pre-industrial period, and the most important period of human population growth associated with industrial development of the region, from the mid-1800s to the present, including the reported development of cultural eutrophication. Comparisons between the cyst records and the known history of eutrophication suggest cyst signals that should prove useful for tracing the development of eutrophication. The eutrophication signal consisted of a doubling of total cyst concentration, and a marked increase in one species in particular,Lingulodinium machaerophorum(from <5 to around 50% of the assemblages) with increased eutrophication. In the core considered most representative of general water quality in the inner fjord, these trends reversed back to pre-industrial levels during the 1980s and 1990s when improved sewage treatment took effect.

Dinoflagellate cysts as potential indicators of industrial pollution in a Norwegian Fjord

Variation in dinoflagellate cyst assemblages through the last approximately 300 years was studied in two sediment cores, one from the heavily polluted Frierfjord, and one from the adjoining, relatively unpolluted Brevikfjord, in order to docu1ent possible dinoflagellate responses to pollution. Changes in the cyst-flora were compared with historical information on the development of industry and also with geochemistry of the sediments, reflecting aspects of pollution. In the Frierfjord core, increasing pollution was accompanied by a decrease in cyst concentration, possibly reflecting reduced production, at least of dinoflagellates, and a shift toward more heterotrophic species, possibly reflecting reduced light penetration in the euphotic zone, or increased production of prey for the heterotrophs. These trends seem to have reversed as pollution decreased after about 1975, suggesting that cyst assemblages contain signals that may prove useful for tracing the development of pollution. Cyst assemblages in the Brevikfjord core only showed minor changes.

Polymeric pH indicators immobilized PVA membranes for optical sensors of high basicity based on a kinetic process

Two kinds of polymeric pH indicators PPF (phenolphthalein-formaldehyde product) and CPF (o-cresolphthalein-formaldehyde product) immobilized cross-linked poly(vinyl alcohol) membranes (PPF-PVA and CPF-PVA) for optical intermittent determination of high basicity ([OH-] = 1-8 M) based on a kinetic process were developed. In our previous work, we had demonstrated that PPF-PVA and CPF-PVA could perform the determination of high pH values from pH 10.0 to 14.0. Here the discoloring kinetic behaviors of PPF-PVA and CPF-PVA were compared with those of free phenolphthalein, o-cresolphthalein and thymolphthalein. Experimental results and theoretical analysis indicated that the response behaviors of the optodes' membranes in concentrated NaOH solutions were diffusion-independent and still complied with the pseudo-first-order kinetics. In addition, two data analysis methods for determination were presented. One was directly based on the reduced absorbance: the other was based on the discoloring kinetic constant. It was found that the latter could perform a rapid (60 s) and reliable (relative standard deviation: 2.6%) determination for high basicity.

Transporting nutrients from sediments: nitrogen and carbon as indicators to measure scavenging ability of a detritivorous fish, Liza haematocheila

The redlip mullet, Liza haematocheila, is a common species in polyculture as a scavenger in China. Feeding on detritus, redlip mullet transports nutrients from sediments up into the water column and converts them into forms that can be utilized by phytoplankton and affects the relative abundance of detritus and dissolved inorganic nutrients to phytoplankton, zooplankton and other fishes. We used nitrogen and carbon as the indicators in this study to measure the scavenging ability, which means intake of nitrogen or carbon by redlip mullet, and the loss of nutrients. Temperature and body weight significantly affected growth nitrogen, faecal nitrogen and faecal carbon. At a certain temperature, the proportion of growth nitrogen or growth carbon increased while the proportion of excretion nitrogen or respiration carbon decreased with increasing body weight.

汉语发展性阅读障碍的神经心理学研究The Neuropsychological Basic of Chinese Developmental Dyslexia by ERP Study

本论文实验结果发现汉语阅读障碍在视听觉功能上有非语言特异性的感知觉加工缺陷：他们在完成视觉任务时成绩稍差，反应时延长，ERP波形成分的峰潜伏期也延长，波幅降低，尤其是左侧枕叶视皮层对右侧视野投射信号的加工效能差，这是首次报道他们枕叶视皮质的左右差异；他们完成听觉分辨任务时，对听觉分辨的效能比正常儿童低；尤其是在听觉相关电位中的表现，峰潜伏期延长，波幅降低。他们在视听觉双通道信号整合的加工上仍表现出ERP波形成分的峰潜伏期延长，波幅较低的现象；综合这三方面的结果，可以推论阅读障碍者可能对一般的信息进行感知觉加工的效能弱。 他们在同音字判断任务中表现出有语音意识缺陷：完成同音字判断任务的正确率与正常儿童有显著的差异；对假字的加工和形似同音字的加工产生的ERP波形均与正常儿童有显著差异，这种差异尤其在左脑更明显。 结合Franck的理论，推论汉语阅读障碍形成原因：汉语阅读障碍行为表现多种多样，这些变异性可能有一些共同的神经心理基础；由于出生前神经的异常发育，遗传的异常或环境的影响而导致的大脑左右两半球功能的差异，尤其左半球引起其对语言加工的缺陷，可能伴随出现视听觉等基本的感知觉功能障碍。