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Toxicity Evaluation of Acrylamide on the Early Life Stages of the Zebrafish Embryos (Danio rerio)  [PDF]
Hattie Spencer, Joseph Wahome, Mary Haasch
Journal of Environmental Protection (JEP) , 2018, DOI: 10.4236/jep.2018.910067
Abstract: Acrylamide is a chemical used mainly in industrial applications and the treatment of drinking and wastewater, making it easy to enter aquatic ecosystems. There are few studies known about the toxicity of acrylamide to aquatic organisms which have shown evidence of a number of histopathological effects. To assess the effects of acrylamide to freshwater fish, Zebrafish (Danio rerio) embryos were exposed to serial concentrations of acrylamide (0, 100, 300, and 500 mg/L) to investigate the acute toxicity effects on teleost embryogenesis. Embryos less than 24 hrs old were exposed under static non-renewal conditions for ten days or until hatching. The toxic endpoints evaluated include: egg/embryo viability, hatchability, and morphological/developmental anomalies during organogenesis. The acute toxicity test resulted in a 48 h-LC50 of 585 mg/L for egg viability. Exposure of embryos significantly reduced hatchability and larval survival, in a concentration dependent manner. Dimethyl sulfoxide (DMSO) was used as a solvent carrier to permeate the uptake of acrylamide through the chorion membrane. No significant damages or complications were observed in embryos exposed to DMSO. At 500 mg/L, the highest test concentration, the survival of embryos was greatly reduced within 24 hrs of exposure. The lower test, 100 mg/L, produced a significant number of developmental anomalies to the Zebrafish that included dorsal tail flexure, severe pericardial edema, facial and cranial defects and decreased heartrate (40 bpm). Premature hatching of embryos and developmental arrest was observed in all concentrations. The severity of these anomalies was concentration-dependent and resulted in low survival rate and high frequency of malformations. These results indicate that acrylamide is teratogenic and provide support for sub-lethal toxicity testing using Zebrafish embryos.
Noninvasive technique for measurement of heartbeat regularity in zebrafish (Danio rerio) embryos
Po Chan, Chun Lin, Shuk Cheng
BMC Biotechnology , 2009, DOI: 10.1186/1472-6750-9-11
Abstract: The heart rate measured by this noninvasive method in zebrafish embryos at 52 hour post-fertilization was similar to that determined by direct visual counting of ventricle beating (p > 0.05). In addition, the method was validated by a known cardiotoxic drug, terfenadine, which affects heartbeat regularity in humans and induces bradycardia and atrioventricular blockage in zebrafish. A significant decrease in heart rate was found by our method in treated embryos (p < 0.01). Moreover, there was a significant increase of the rhythmicity index (p < 0.01), which was supported by an increase in beat-to-beat interval variability (p < 0.01) of treated embryos as shown by Poincare plot.The data support and validate this rapid, simple, noninvasive method, which includes video image analysis and frequency analysis. This method is capable of measuring the heart rate and heartbeat regularity simultaneously via the analysis of caudal blood flow in zebrafish embryos. With the advantages of rapid sample preparation procedures, automatic image analysis and data analysis, this method can potentially be applied to cardiotoxicity screening assay.In spite of the morphological differences of the heart, the early function is similar among different model animals studied, including zebrafish [1-3]. The zebrafish embryo has been suggested as an ideal model to uncover the molecular mechanism of cardiac development and to identify genes related to congenital cardiac defects in human [1,4,5]. In addition, although there are differences between zebrafish embryos and human in responses to human cardiotoxic drugs, such as the dissociation between the atrium and ventricle [6,7] in zebrafish embryos which is not normally observed in adult humans, recent studies demonstrated that zebrafish still has some similar physiological responses to those drugs. Bradycardia, one type of cardiac arrhythmia, was induced in zebrafish embryos after treated by well-known QT-prolonging drugs, such as terfenadine [6,8
Influence of carbon nanotube length on toxicity to zebrafish embryos
Cheng J, Cheng SH
International Journal of Nanomedicine , 2012, DOI: http://dx.doi.org/10.2147/IJN.S30459
Abstract: fluence of carbon nanotube length on toxicity to zebrafish embryos Original Research (2384) Total Article Views Authors: Cheng J, Cheng SH Published Date July 2012 Volume 2012:7 Pages 3731 - 3739 DOI: http://dx.doi.org/10.2147/IJN.S30459 Received: 01 February 2012 Accepted: 27 March 2012 Published: 20 July 2012 Jinping Cheng,1,2 Shuk Han Cheng1 1Department of Biology and Chemistry, City University of Hong Kong, Hong Kong; 2State Key Laboratory of Estuarine and Coastal Research, East China Normal University, Shanghai, China Abstract: There is currently a large difference of opinion in nanotoxicology studies of nanomaterials. There is concern about why some studies have indicated that there is strong toxicity, while others have not. In this study, the length of carbon nanotubes greatly affected their toxicity in zebrafish embryos. Multiwalled carbon nanotubes (MWCNTs) were sonicated in a nitric acid solution for 24 hours and 48 hours. The modified MWCNTs were tested in early developing zebrafish embryo. MWCNTs prepared with the longer sonication time resulted in severe developmental toxicity; however, the shorter sonication time did not induce any obvious toxicity in the tested developing zebrafish embryos. The cellular and molecular changes of the affected zebrafish embryos were studied and the observed phenotypes scored. This study suggests that length plays an important role in the in vivo toxicity of functionalized CNTs. This study will help in furthering the understanding on current differences in toxicity studies of nanomaterials.
Cell Adhesion in Zebrafish Embryos Is Modulated by March8  [PDF]
Mi Ha Kim, Martha L. Rebbert, Hyunju Ro, Minho Won, Igor B. Dawid
PLOS ONE , 2014, DOI: 10.1371/journal.pone.0094873
Abstract: March8 is a member of a family of transmembrane E3 ubiquitin ligases that have been studied mostly for their role in the immune system. We find that March8 is expressed in the zebrafish egg and early embryo, suggesting a role in development. Both knock-down and overexpression of March8 leads to abnormal development. The phenotype of zebrafish embryos and Xenopus animal explants overexpressing March8 implicates impairment of cell adhesion as a cause of the effect. In zebrafish embryos and in cultured cells, overexpression of March8 leads to a reduction in the surface levels of E-cadherin, a major cell-cell adhesion molecule. Experiments in cell culture further show that E-cadherin can be ubiquitinated by March8. On the basis of these observations we suggest that March8 functions in the embryo to modulate the strength of cell adhesion by regulating the localization of E-cadherin.
Toxic effects of TBBPA on in vivo and in vitro developments in the zebrafish (Danio rerio) embryos
四溴双酚A对斑马鱼胚胎体内外发育的毒性效应

DU Qingping,PENG Run,LIU Wuxiang,JIA Xiaoshan,WEI Dongyang,
杜青平
,彭润,刘伍香,贾晓珊,魏东洋

环境科学学报 , 2012,
Abstract: Tetrabromobisphenol A (TBBPA) is a widely used brominated flame retardant that is present widely in the environment. The developmental toxicity effects of TBBPA on the zebrafish (Danio rerio) embryos were studied both in vitro and in vivo. The results showed that exposure to water-borne TBBPA would cause teratogenic effects and death of zebrafish embryos. The teratogenic defects included pericardial edema, hemoglutinations and axial malformation. The lethality rate significantly increased when the embryos were treated with higher concentrations of TBBPA. Furthermore, the death of zebrafish embryos mainly happened in 24 h post-fertilization (hpf). The zebrafish embryos exposed to 6.4 mg·L-1 TBBPA were all dead during 48 hpf. The activities of spontaneous movement of zebrafish embryos exposed to different TBBPA concentrations higher than 0.4 mg·L-1 decreased in 20s. The main toxic effect to the embryos in 48 hpf was pericardial edema. No difference in the heart rates of embryos was observed between the control group and treated groups. The main toxic effect for the embryos in 72 h was pericardial edema and curved trunk. The hatching rates and survival rates were significantly decreased. These results suggested that TBBPA in the water were toxic directly to zebrafish embryos' early development in a dose-response manner. The lethality of offspring embryos was increased at the endpoints of 24 h and 72 h in a time-response manner when the adult zebrafish were exposed to 1.5 mg·L-1 TBBPA for 3~7 days. The lethality rate showed significant difference in 72 hpf between TBBPA treated groups and the control group in vivo. The hatching rates of offspring embryos decreased when adult zebrafish were treated with TBBPA, however, no significant difference was observed. The teratogenesis rates of offspring embryos significantly increased, and showed marked difference between the treated groups and the control groups. The results indicated that TBBPA residue in water was potentially hazardous to the development of fishes.
Effects of bisphenol A toxicity on zebrafish embryos
双酚A对斑马鱼胚胎的毒性作用

DUAN Zheng-Hua,ZHU Lin,
DUAN
,Zheng-Hua,and,ZHU,Lin

水生生物学报 , 2006,
Abstract: In order to evaluate the toxicity of the bisphenol A (BPA) on the growth of zebrafish embryos, fertilized eggs were exposed to the concentration of 2.00, 4.00, 6.00, 8.00, 10.00, 15.00, 18.00, 22.00 and 25.00mg/L BPA for72 h at 26± 1 ℃. The results revealed that the sublethal toxicological endpoints induced by BPA were: delayed hatch > blood balk > cyst > altered axial curvature and tail malformation. The median embryo lethal concentration (LC50) after 24 h was 16.36 mg/L. We concluded that the BPA toxicity on zebrafish embryos were caused before 8h exposure and it was not the result of long-term accumulation. Therefore, BPA maybe cause altered gene expression at the early stage of zebrafish embryos. In the further studies, we will use the technology of genetic chips to look for the toxic mechanism of BPA.
The paracrine effect of exogenous growth hormone alleviates dysmorphogenesis caused by tbx5 deficiency in zebrafish (Danio rerio) embryos
Tzu-Chun Tsai, Jenn-Kan Lu, Sie-Lin Choo, Shu-Yu Yeh, Ren-Bing Tang, Hsin-Yu Lee, Jen-Her Lu
Journal of Biomedical Science , 2012, DOI: 10.1186/1423-0127-19-63
Abstract: Using an oligonucleotide-based microarray analysis to study the expression of special genes in tbx5 morphants, we demonstrated that GH and some GH-related genes were markedly downregulated. Zebrafish embryos microinjected with tbx5-morpholino (MO) antisense RNA and mismatched antisense RNA in the 1-cell stage served as controls, while zebrafish embryos co-injected with exogenous growth hormone (GH) concomitant with tbx5-MO comprised the treatment group.The attenuating effects of GH in tbx5-MO knockdown embryos were quantified and observed at 24, 30, 48, 72, and 96?h post-fertilization. Though the understanding of mechanisms involving GH in the tbx5 functioning complex is limited, exogenous GH supplied to tbx5 knockdown zebrafish embryos is able to enhance the expression of downstream mediators in the GH and insulin-like growth factor (IGF)-1 pathway, including igf1, ghra, and ghrb, and signal transductors (erk1, akt2), and eventually to correct dysmorphogenesis in various organs including the heart and pectoral fins. Supplementary GH also reduced apoptosis as determined by a TUNEL assay and decreased the expression of apoptosis-related genes and proteins (bcl2 and bad) according to semiquantitative reverse-transcription polymerase chain reaction and immunohistochemical analysis, respectively, as well as improving cell cycle-related genes (p27 and cdk2) and cardiomyogenetic genes (amhc, vmhc, and cmlc2).Based on our results, tbx5 knockdown causes a pseudo GH deficiency in zebrafish during early embryonic stages, and supplementation of exogenous GH can partially restore dysmorphogenesis, apoptosis, cell growth inhibition, and abnormal cardiomyogenesis in tbx5 knockdown zebrafish in a paracrine manner.
Effects of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos

Jun'an Cui,Zhiyong Zhang,Wei Bai,Ligang Zhang,Xiao He,Yuhui M,Yan Liu,Zhifang Chai,

环境科学学报(英文版) , 2012,
Abstract: In recent years, with the wide applications and mineral exploitation of rare earth elements, their potential environmental and health effects have caused increasing public concern. Effect of rare earth elements La and Yb on the morphological and functional development of zebrafish embryos were studied. The embryos were exposed to La3+ or Yb3+ at 0, 0.01, 0.1, 0.3, 0.5 and 1.0 mmol/L, respectively. Early life stage parameters such as egg and embryo mortality, gastrula development, tail detachment, eyes, somite formation, circulatory system, pigmentation, malformations, hatching rate, length of larvae and mortality were investigated. The results showed La3+ and Yb3+ delayed zebrafish embryo and larval development, decreased survival and hatching rates, and caused tail malformation in a concentration-dependent way. Moreover, heavy rare-earth ytterbium led to more severe acute toxicity of zebrafish embryo than light rare-earth lanthanum.
Low Temperature Mitigates Cardia Bifida in Zebrafish Embryos  [PDF]
Che-Yi Lin, Cheng-Chen Huang, Wen-Der Wang, Chung-Der Hsiao, Ching-Feng Cheng, Yi-Ting Wu, Yu-Fen Lu, Sheng-Ping L. Hwang
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0069788
Abstract: The coordinated migration of bilateral cardiomyocytes and the formation of the cardiac cone are essential for heart tube formation. We investigated gene regulatory mechanisms involved in myocardial migration, and regulation of the timing of cardiac cone formation in zebrafish embryos. Through screening of zebrafish treated with ethylnitrosourea, we isolated a mutant with a hypomorphic allele of mil (s1pr2)/edg5, called s1pr2as10 (as10). Mutant embryos with this allele expressed less mil/edg5 mRNA and exhibited cardia bifida prior to 28 hours post-fertilization. Although the bilateral hearts of the mutants gradually fused together, the resulting formation of two atria and one tightly-packed ventricle failed to support normal blood circulation. Interestingly, cardia bifida of s1pr2as10 embryos could be rescued and normal circulation could be restored by incubating the embryos at low temperature (22.5°C). Rescue was also observed in gata5 and bon cardia bifida morphants raised at 22.5°C. The use of DNA microarrays, digital gene expression analyses, loss-of-function, as well as mRNA and protein rescue experiments, revealed that low temperature mitigates cardia bifida by regulating the expression of genes encoding components of the extracellular matrix (fibronectin 1, tenascin-c, tenascin-w). Furthermore, the addition of N-acetyl cysteine (NAC), a reactive oxygen species (ROS) scavenger, significantly decreased the effect of low temperature on mitigating cardia bifida in s1pr2as10 embryos. Our study reveals that temperature coordinates the development of the heart tube and somitogenesis, and that extracellular matrix genes (fibronectin 1, tenascin-c and tenascin-w) are involved.
Toxic Effects of Silica Nanoparticles on Zebrafish Embryos and Larvae  [PDF]
Junchao Duan, Yongbo Yu, Huiqin Shi, Linwei Tian, Caixia Guo, Peili Huang, Xianqing Zhou, Shuangqing Peng, Zhiwei Sun
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0074606
Abstract: Silica nanoparticles (SiNPs) have been widely used in biomedical and biotechnological applications. Environmental exposure to nanomaterials is inevitable as they become part of our daily life. Therefore, it is necessary to investigate the possible toxic effects of SiNPs exposure. In this study, zebrafish embryos were treated with SiNPs (25, 50, 100, 200 μg/mL) during 4–96 hours post fertilization (hpf). Mortality, hatching rate, malformation and whole-embryo cellular death were detected. We also measured the larval behavior to analyze whether SiNPs had adverse effects on larvae locomotor activity. The results showed that as the exposure dosages increasing, the hatching rate of zebrafish embryos was decreased while the mortality and cell death were increased. Exposure to SiNPs caused embryonic malformations, including pericardial edema, yolk sac edema, tail and head malformation. The larval behavior testing showed that the total swimming distance was decreased in a dose-dependent manner. The lower dose (25 and 50 μg/mL SiNPs) produced substantial hyperactivity while the higher doses (100 and 200 μg/mL SiNPs) elicited remarkably hypoactivity in dark periods. In summary, our data indicated that SiNPs caused embryonic developmental toxicity, resulted in persistent effects on larval behavior.
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