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The protective effect of GSP on UVB irradiation in zebrafish growth and development

Author: ZuoYongJian
Tutor: GuoBingRan；ShiYongXiang
School: Qufu Normal University
Course: Zoology
Keywords: ultraviolet B zebrafish damage single-cell gel electrophoresis immunohistochemistry
CLC: X174
Type: Master's thesis
Year: 2009
Downloads: 73
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Abstract

Objects:Environmental factors are the most important factors influenting the organisms on the earth and the changes of environments have direct effects on the growth,propagation,inheritance and species continuity.The ultraviolet(UV) is a very important factor in environmental factors,especially the ultraviolet-B(UVB).In the past few years,human’s activities cause more and more UVB in environment and that has a serious effect on the organisms.The zebrafish was used as model animals to research the harmfull effects of UVB and antioxidation effects of grape seed proanthocyanidin(GSP),in order to provide helpful meterials for environmental protection.Methods:1) Zebrafish embryos were grouped according to different irradiation intensities and irradiation time of UVB,while non-irradiated control group was set up.We observed the morphological changes of the zebrafish embryos in different groups,and counted survival rates,mortality rates,abnormal rates and other related data as evaluation of target.2) The DNA damages of Zebrafish embryos which were irradiated by UVB were detected by single cell gel electrophoresis technique.We researched on DNA damages caused by UVB radiation in embryonic cells,the tolerances to UVB irradiation on different embryonic stages,and the changes of DNA damage on different exposure conditions.3) We injected GSE of a certain concentration into the egg yolk of gastrula stage zebrafish embryos by micro-injection method,using distilled water as a control.The embryos were incubated under the same conditions.The results were observed and counted.4) We added GSE of a certain concentration to the food and the surrounding living environment of zebrafish.The expression of Caspase-3 protein in the retina of adult fish was tested by immunohistochemical methods.The normal feeding fishes were set up in as a control.Caspase-3 protein expression could indirectly reflect the GSP’s antioxidant role against UVB.Results:1) UVB-irradiation has a great impact on morphology of Zebrafish embryonic development,and a clear dose-response relationship was not found in a short period.With the increase of exposure time and the strength of UVB irradiation,the mortality of embryos and the abnormal rate of larval increased,showing a significant positive correlation,and the rates eventually reached 100%.UVB exposure could lead to many kinds abnormal zebrafishes, such as curve of the tail,expanding of pericardial cavity,spinal twist and so on.With the intensity and duration of UVB irradiation increase,the rates of malformation and severe malformation had an obvious increase,and all of these embryos eventually became malformation or even severe malformation.2) UVB irradiation could also cause obvious DNA damages in the embryos of zebrafish, and the injury depended on the irradiation time and irradiation intensity directly,but a clear cumulative effect has not been found.We compared larval fishes on the gastrula stage,body section stage and 2-days stage on the same irradiation conditions.We found the embryo’s abilities to anti-UVB radiation continuously strengthened.3) GSE was injected into the yolk of zebrafishes’ gastrula stage embryos.We found that the abnormal and mortality rate of the injected embryos was significantly lower than that of the controls.Some juvenile fishes were feeded in the water with a certain concentration of GSE.GSE caused the level of activity lower and the mortality rate increased.If the concentration of GSE was high enough,it cause all juvenile fishes dead.4) Immunohistochemical staining showed that Caspase-3 protein mainly expression in the outer layer of the retina.Expression of Caspase-3 protein was increased in the retina of adult fishes irradiated by UVB.However,in the group which was fed GSE and irradiated by UVB,expression of Caspase-3 protein is lower than that of the foregoing group.Conclusion:1) UVB irradiation has significant teratogenic and lethal effects on the early stage of zebrafish embryos.2) UVB irradiation has injury effect on the genetic material of zebrafish embryos with an obvious dose-response relationship.3) GSP can effectively enhance the survival rate of zebrafish embryos which were exposed to UVB.4) GSP has significant protective effect and can reduce the damage degree of zebrafish retina exposed to radiation.

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