Ethanol disrupts the formation of hypochord and dorsal aorta during the development of embryonic zebrafish

Exposure to ethanol during human embryonic period has severe teratogenic effects on the cardiovascular system. In our study, we demonstrated that ethanol of gradient concentrations can interfere with the establishment of circulatory system in embryonic zebrafish. The effective concentration to cause 50% malformations (EC₅₀) was 182.5 mmol/L. The ethanol pulse exposure experiment displayed that dome stage during embryogenesis is the sensitive time window to ethanol. It is found that 400 mmol/L ethanol pulse exposure can induce circulatory defects in 43% treated embryos. We ruled out the possibility that ethanol can interfere with the process of hematopoiesis in zebrafish. By employing in situ hybridization with endothelial biomarker (Flk-1), we revealed that ethanol disrupts the establishment of trunk axial vasculature, but has no effect on cranial vessels. Combined with the results of semi-thin histological sections, the in situ hybridization experiments with arterial and venous biomarkers (ephrinB2, ephB4) suggested that ethanol mainly interrupts the development of dorsal aorta while has little effect on axial vein. Further study indicated the negative influence of ethanol on the development of hypochord in zebrafish. The consequent lack of vasculogenic factors including Radar and Ang- 1 partly explains the defects in formation and integrity of dorsal aorta. These results provide important clues to the study of adverse effects of ethanol on the cardiovascular development in human fetus.     

Ethanol disrupts the formation of hypochord and dorsal aorta during the development of embryonic zebrafish

It has been demonstrated that maternal drinking during pregnancy had serious adverse effects on the health of the newborns. Fetal alcohol syndrome (FAS) is the most important developmental abnormality caused by maternal alcohol abuse during pregnancy. Clinically, it is characterized by head and facial ab-normalities, cardiovascular malformation, and perma-nent nervous system damage[1,2]. A lot of experimental models have been developed to study the ethanol’s effects on embryonic development,…     

Differential effect of ionizing radiation exposure on multipotent and differentiation‐restricted bone marrow mesenchymal stem cells

Debilitating effects of bone marrow from ionizing radiation exposure has been well established for hematopoietic stem cells; however, radiation toxicity of mesenchymal stem cells (MSCs) has been controversial. The present study addressed if ionizing radiation exposure differently affected bone marrow MSCs with various differentiation commitments. Mouse bone‐marrow‐derived MSCs, D1 cells of early passages (≤5 passages; p5) maintained the complete characteristics of multipotent MSCs, whereas, after ≥45 passages (p45) the differentiation capability of D1 cells became partially restricted. Both p5 and p45 D1 cells were subjected to single dose irradiation by radioactive isotope ¹³⁷Cs. Radiation treatment impaired cell renewal and differentiation activities of p5 D1 cells; however, p45 D1 cells were less affected. Radiation treatment upregulated both pro‐ and anti‐apoptotic genes of p5 D1 cells in a dose‐dependent manner, potentially resulting in the various apoptosis thresholds. It was found that constitutive as well as radiation‐induced phosphorylation levels of histone H2AX was significantly higher in p45 D1 cells than in p5 D1 cells. The increased repair activity of DNA double‐strand breakage may play a role for p45 D1 cells to exhibit the relative radioresistance. In conclusion, the radiation toxicity predominantly affecting multipotent MSCs may occur at unexpectedly low doses, which may, in part, contribute to the catabolic pathology of bone tissue. J. Cell. Biochem. 111: 322–332, 2010. © 2010 Wiley‐Liss, Inc.     

Small molecule screening identifies targetable zebrafish pigmentation pathways

Small molecules complement genetic mutants and can be used to probe pigment cell biology by inhibiting specific proteins or pathways. Here, we present the results of a screen of active compounds for those that affect the processes of melanocyte and iridophore development in zebrafish and investigate the effects of a few of these compounds in further detail. We identified and confirmed 57 compounds that altered pigment cell patterning, number, survival, or differentiation. Additional tissue targets and toxicity of small molecules are also discussed. Given that the majority of cell types, including pigment cells, are conserved between zebrafish and other vertebrates, we present these chemicals as molecular tools to study developmental processes of pigment cells in living animals and emphasize the value of zebrafish as an in vivo system for testing the on- and off-target activities of clinically active drugs.     

Gut metabolite Urolithin A mitigates ionizing radiation-induced intestinal damage

Ionizing radiation (IR)-induced intestinal damage is the major and common injury of patients receiving radiotherapy. Urolithin A (UroA) is a metabolite of the intestinal flora of ellagitannin, a compound found in fruits and nuts such as pomegranates, strawberries and walnuts. UroA shows the immunomodulatory and anti-inflammatory capacity in various metabolic diseases. To evaluate the radioprotective effects, UroA(0.4, 2 and 10 mg/kg) were intraperitoneally injected to C57BL/6 male mice 48, 24, 1 h prior to and 24 h after 9.0Gy TBI. The results showed that UroA markedly upregulated the survival of irradiated mice, especially at concentration of 2 mg/kg. UroA improved the intestine morphology architecture and the regeneration ability of enterocytes in irradiated mice. Then, UroA significantly decreased the apoptosis of enterocytes induced by radiation. Additionally, 16S rRNA sequencing analysis showed the effect of UroA is associated with the recovery of the IR-induced intestinal microbacteria profile changes in mice. Therefore, our results determinated UroA could be developed as a potential candidate for radiomitigators in radiotherapy and accidental nuclear exposure. And the beneficial functions of UroA might be associated with the inhibition of p53-mediated apoptosis and remodelling of the gut microbes.     

In vivo imaging of embryonic vascular development using transgenic zebrafish

In this study we describe a model system that allows continuous in vivo observation of the vertebrate embryonic vasculature. We find that the zebrafish fli1 promoter is able to drive expression of enhanced green fluorescent protein (EGFP) in all blood vessels throughout embryogenesis. We demonstrate the utility of vascular-specific transgenic zebrafish in conjunction with time-lapse multiphoton laser scanning microscopy by directly observing angiogenesis within the brain of developing embryos. Our images reveal that blood vessels undergoing active angiogenic growth display extensive filopodial activity and pathfinding behavior similar to that of neuronal growth cones. We further show, using the zebrafish mindbomb mutant as an example, that the expression of EGFP within developing blood vessels permits detailed analysis of vascular defects associated with genetic mutations. Thus, these transgenic lines allow detailed analysis of both wild type and mutant embryonic vasculature and, together with the ability to perform large scale forward-genetic screens in zebrafish, will facilitate identification of new mutants affecting vascular development.     

Mrg15 null and heterozygous mouse embryonic fibroblasts exhibit DNA-repair defects post exposure to gamma ionizing radiation

MORF4-related gene on chromosome 15 (MRG15) is a core component of the NuA4/Tip60 histone acetyltransferase complex that modifies chromatin structure. We here demonstrate that Mrg15 null and heterozygous mouse embryonic fibroblasts exhibit an impaired DNA-damage response post gamma irradiation, when compared to wild-type cells. Defects in DNA-repair and cell growth, and delayed recruitment of repair proteins to sites of damage were observed. Formation of phosphorylated H2AX and 53BP1 foci was delayed in Mrg15 mutant versus wild-type cells following irradiation. These data implicate a novel role for MRG15 in DNA-damage repair in mammalian cells.     

Low temperature enhances petunia flower pigmentation and induces chalcone synthase gene expression

Flower coloration is controlled by internal and external factors, including temperature. The aim of the present work was to examine the effect of temperature on anthocyanin synthesis and chalcone synthase gene ( chs ) expression in petunia flowers. A moderate-low temperature enhanced both anthocyanin accumulation and chs expression in the corollas. However, the effect on chs expression was not always correlated with that on anthocyanin content, suggesting a post-translational effect. The effect was local and required the exposure of corollas, but not the whole plant, to the ambient temperature. The response of chs to moderate-low temperatures did not coincide with its expression during flower development. Moderate-low temperatures only slightly affected gibberellic acid (GA₃)-induced chs expression in the light, but activated chs expression under non-inducing conditions, i.e. in the absence of GA₃ in the dark. The results of this study suggest that moderate-low temperatures do not simply enhance the developmental regulation of anthocyanin biosynthetic gene expression; they act as a specific and separate signal.     

Sensitization of ionizing radiation-induced apoptosis by ursolic acid

Radiation therapy has been widely used for treating human cancers. However, cancer cells develop radioresistant phenotypes that decrease the efficacy of radiotherapy. Ionizing radiation (IR) induces the production of reactive oxygen species, which play an important role in apoptotic cell death. Therefore, radiation therapy combined with a sensitizer, which modulates cellular redox status, has the potential to enhance therapeutic efficacy in a variety of human cancers. Here, we investigated the radiosensitizing effects of ursolic acid (UA), a pentacyclic triterpenoid found in rosemary and holy basil. IR-induced apoptosis in cancer cell lines such as DU145, CT26 and B16F10 was significantly enhanced by UA, as reflected by DNA fragmentation, cellular redox status, mitochondrial dysfunction and modulation of apoptotic marker proteins. Additionally, UA combined with IR was also effective for inhibiting tumorigenesis in B16F10 melanoma cells implanted into mice. Taken together, these results suggest that applying UA together with IR may be an effective combination modality for treating cancer.     

A dominant mutation in tyrp1A leads to melanophore death in zebrafish

Melanin biosynthesis in vertebrates depends on the function of three enzymes of the tyrosinase family, tyrosinase (Tyr), tyrosinase‐related protein 1 (Tyrp1), and dopachrome tautomerase (Dct or Tyrp2). Tyrp1 might play an additional role in the survival and proliferation of melanocytes. Here, we describe a mutation in tyrp1A, one of the two tyrp1 paralogs in zebrafish, which causes melanophore death leading to a semi‐dominant phenotype. The mutation, an Arg‐>Cys change in the amino‐terminal part of the protein, is similar to mutations in humans and mice where they lead to blond hair (in melanesians) or dark hair with white bases, respectively. We demonstrate that the phenotype in zebrafish depends on the presence of the mutant protein and on melanin synthesis. Ultrastructural analysis shows that the melanosome morphology and pigment content are altered in the mutants. These structural changes might be the underlying cause for the observed cell death, which, surprisingly, does not result in patterning defects.     

Specific activation of ERK pathways by chitin oligosaccharides in embryonic zebrafish cell lines

Chitin oligosaccharides (COs) play a role in plant development and are presumed to affect body plan formation during vertebrate embryogenesis. The mechanisms of COs recognition and cellular processes underlying embryonic development are still not understood. We analyze the possible link with the mitogen-activated protein kinase pathway that is conserved in evolution through the plant and animal kingdom and has been implicated in diverse cellular processes, including cell growth, proliferation, differentiation, survival, and vertebrate development. We show that in vivo stimulation of embryonic zebrafish cells ZF13 and ZF29 with chitin tetrasaccharides at 10-9 M concentration transiently induced activation/phosphorylation of extracellular regulated kinases (ERKs), with a maximum after 15 min. Furthermore the biological specificity of chitin tetrasaccharides and various derivatives was examined. The replacement of one or two GlcNAc residues of the chitin backbone by glucose and fucosylation of chitin tetrasaccharides at the reducing terminus caused a complete loss of their activity. We also tested a chitin tetrasaccharide analogue in which the oxygen atoms in glycosidic linkages were replaced by sulfur atoms. This analog, which could not be enzymatically hydrolyzed, was as potent an inducer as chitin tetrasaccharide. These results suggest that the observed activation of ERKs is chitin tetrasaccharide-specific and does not require further enzymatic processing. We examined possible signaling pathways leading to ERK activation by COs by use of phosphospecific antibodies and inhibitors. We conclude that a high-affinity CO receptor system exists that links to the Raf, MEK, and ERK pathway in zebrafish cells.     

KCl:Dy phosphor for thermoluminescence dosimetry of ionizing radiation

The thermoluminescence (TL) characterizations of γ‐irradiated KCl:Dy phosphor for radiation dosimetry are reported. All phosphors were synthesized via a wet chemical route. Minimum fading of TL intensity is recorded in the prepared material. TL in samples containing different concentrations of Dy impurity was studied at different γ‐irradiation doses. Peak TL intensities varied sublinearly with γ‐ray dose in all samples, but were linear between 0.08 to 0.75 kGy for the KCl:Dy (0.1 mol%) sample. This material may be useful for dosimetry within this range of γ‐ray dose. TL peak height was found to be dependant on the concentration (0.05–0.5 mol%) of added Dy in the host. Copyright © 2012 John Wiley & Sons, Ltd.