| Cadmium,one of the major heavy metal pollutants in the environment,has high toxicity and severe bioaccumulation.It can transmit and accumulate in the organisms and cause cell oxidative damage,cell apoptosis and tissue necrosis and so on,thus affecting the normal growth and development of organisms and the fitness of the offspring.When the organism is subject to heavy metal stress,the body itself will have a mechanism to respond to reduce the damage,and excrete harmful substances,so as to maintain the body environment homeostasis.Heat shock protein is an important medium for organisms to adapt to environmental changes,not only covers the biological significance of high-temperature stress,but also responses to heavy metal pollution,thereby producing the corresponding physiological changes to accommodate the environment.The present paper chooses Red-eared slider(Trachemys scripta elegans),which belongs to the Phylum vertebrata,reptilia,Testudines,Emydidae,Trachemys,as experimental material to assess the physiological and behavioral effects of cadmium to turtles embryos and 96 h-LD50 and the relative expression of hsc70 and hsp90 in six tissues of turtles larvae exposed the different cadmium dose and time.Firstly,this study was carried out four groups,the cadmium dose was set to 0 mg/kg,25 mg/kg,50 mg/kg and 100 mg/kg vermiculite respectively,the turtles eggs were incubated under four groups.The aim was to test the effects of cadmium with different dose on morphological characteristics and motility of offspring.The results showed that the incubation rates in four groups were significantly different.ANCOVA with eggs weight revealed that there were significant differences in carapace length and plastron length and nonsignificant differences in body weight and carapace width and plastron width of larvae in different groups.When the larvae were fed up to 80 days,the morphological characteristics and the motor performance were measured.The results showed that body weight had not significant increase,but carapace length,carapace width,plastron length and plastron width of larvae had significant increase.And when 80 days,body weight,carapace length,carapace width,plastron length and plastron width of larvae had not significant difference,But the maximal distance and sprint speed of the larvae were significantly different in four groups,and the greater the cadmium dose was,the smaller the maximal distance and sprint speed were.In summary,the results revealed different cadmium stress affect the morphology and the movement of turtles offspring.Secondly,in order to study the sensitivity of cadmium to turtles and the median lethal dose after 96 hours of exposure.The experiment was used to inject different cadmium dose and then calculated the death rates after 96 hours by Linear regression and Karben method.The results showed that the 96 h-LD50 of acute exposure was 13.69 mg/kg body mass calculated by Linear regression but was 13.44 mg/kg body mass calculated by Karben method,and they have not significant difference.This results provided the basis for dose setting for the next study phase.To provide a digital basis for assessing the toxicity of cadmium to Trachemys scripta elegans larvae and subsequent molecular experiments.Finally,Quantitative Real-time PCR was performed to quantity the relative transcript levels of heat shock cognate gene 70(hsc70)and heat shock protein 90(hsp90)in the liver,kidney,muscle,lung,brain and heart in the turtles larvae exposed different cadmium dose.The dose of cadmium were set to 0 mg/kg,3.49 mg/kg,7.24 mg/kg and 10.42 mg/kg body mass,respectively,according the second phase.The results showed that after 96 hours,the relative expression level of hsc70 mRNA in the liver,kidney and brain of larvae was significantly different,and the level of hsc70 was significantly higher in the group of 7.24 mg/kg than those other groups.Compared with the control group,the expression of hsc70 mRNA increased firstly and then decreased.The expression of hsc70 mRNA in other tissues,such as muscle,lung and heart was not significantly affected by different cadmium dose.The effect of different cadmium dose on the expression of hsp90 mRNA in six tissues was only significantly affected in liver.We investigated that the relative expression levels of hsc70 and hsp90 mRNA in different tissues under the same cadmium level.The results showed the transcript levels of hsc70 and hsp90 were different between the different tissues.The expression levels of hsc70 in the kidney and brain were higher than that other four tissues.The expression level of hsp90 in the brain was higher than that other five tissues when the cadmium level of 0 mg/kg and 3.49 mg/kg body mass.However,the level of hsc70 and hsp90 was not affected when 7.24 mg/kg and 10.42 mg/kg body mass.All in all,the liver,kidney and brain are more sensitive to cadmium than other tissues,and with the increase of exposure dose,the toxic effect of cadmium is gradually weakened.This may be the reason why the Trachemys scripta elegans is an invasive species and they have own strong defense function.Meanwhile,we investigated its time-related effects on mRNA transcript levels of hsc70 and hsp90 in six tissues of turtles exposed one cadmium level which is 7.24 mg/kg body mass.The results showed,in the liver,the relative expression of hsc70 mRNA increased as time extends,and reached the highest level at the 96th hour;the level of hsp90 mRNA was not significantly affected.In the kidney,the level of hsc70 mRNA increased as time extends;the level of hsp90 mRNA was higher at 24th hour than that other time group.In muscle,the level of hsc70 mRNA was increased with the time and reached the highest at 96th hour;the level of hsp90 mRNA was increased as time extends and was significantly higher at 72nd hour and 96th hour than other time.In the lung,the level of hsc70 mRNA increased significantly as time extends and reached a significant peak at 96th hour;the level of hsp90 mRNA was significantly increased at 24th hour and was higher at 24th hour and 72nd hour than other time groups.In the heart,the level of hsc70 mRNA was significantly increased and reach to a significant peak at 96th hour,the level of hsp90 mRNA at the 24th hour was significantly higher than the other time.All in all,cadmium does produce toxicity to Trachemys scripta elegans embryos and affect morphological and behavioral performance of offpring;acute cadmium exposure could induce shortened turtles life and cadmium could induce the relative expression of hsc70 and hsp90 mRNA of the larvae.In summary,cadmium could induce the transcript levels of hsc70 and hsp90 mRNA of the turtles,and the changes were dose-tissue-time-dependent induction.That is to say,the toxicity of cadmium on the turtles depends on the combination of cadmium dose,exposure time and tissues and other comprehensive factors.In addition,the up-regulation or down-regulation of hsc70 and hsp90 mRNA indicates that Hsps plays an important protective role in Trachemys scripta elegans response to cadmium stress. |
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