The Expression Characteristics And Relationship Of HSP70 And JNK In The Senescence Of Brachionus Calyciflorus

The Brachionus calyciflorus is a new type of potential model animal for aging research.It has characteristics of short life cycle,easy to be cultivated,widely distributed,and has obvious aging characteristics.So,it's a valuable biological organism for the study of aging.The HSP?Heat shock protein?family is thought to beactivated by stimulates and then protects cells from stress damage.And the effect of HSP70 is more prominent.On the other hand,apoptosis,as the programed cell death,has been shown by many studies that one of the main executors of apoptosis is c-Jun N-terminal kinase?c-Jun N-terminal kinase,JNK?.However,it is still unknown whether HSP can control the aging of rotifers by regulating the JNK signaling pathway.This paper aims to explore whether there is a regulatory effect between HSP70 and JNK pathway by comparing the relationship between HSP70 and JNK at different temperatures,and also at the inhibition of JNK in this case.In this study,B.calyciflorus,an internationally used freshwater rotifers,was used as the research object to carry out a series of experiments.In this study,the activity of HSP70 in B.calyciflorus was discussed,and studied its expression under different stress conditions;The relationship between HSP70 and JNK in rotifers was explored:Ecological indicators of rotifers at different temperatures,such as progenitor period,embryo development period,life cycle and survival rate,in each group of normal and JNK inhibitor SP600125;And the comparison of HSP70 and JNK at different temperatures and aging conditions.The main studies result of this paper are as follows:1.Effect of different temperatures and JNK inhibitor on the life span of B.calyciflorusThe results showed that temperature had an effect on the life span of brachionus rotifers,and with the decrease of temperature,the pre-reproductive period,the embryonic development period,the life cycle and survival rate of rotifers increased.Under normal conditions,the pre-reproductive period was significantly increased at 15,20,25?than 30?,and the longest time at 15 was 39.739±8.72 h.the embryonic development period at 15?and 20?have grown significantly then 30?,and 15?has longest time?26.167±2.44 h?.The life cycle increased significantly at the other three temperatures compared with 30?,with the most significant increase at15??241.541±27.82 h?.The lower the temperature was,the longer the survival time of brachionus rotifers.The longest survival time of rotifers was up to 20 d at 15?.Under the action of JNK inhibitor,the data showed that the all parameters of rotifers increased compared with the normal conditions.The pre-reproductive period of rotifers in the inhibition group was similar to that in the normal group,with the longest time of 45.739±8.62 h at 15?,and there was a significant difference between the two groups at 20?and 25?compared with the normal group.After the inhibition,Time of the embryonic development period increases with temperature decrease,the longest at 15??31.125±5.46 h?.Compared with the normal conditions,there were significant differences in each temperature.After the inhibition,the life cycle is longer than normal,the difference was most pronounced at 30?and 25?,the longest at 15??254.125±32.28 h?.The longest survival time of brachionus rotifers was up to 21.5 d after inhibition.These results indicate that the life span of brachionus is related to temperature and it might have something to do with JNK.2.Effect of temperature and aging on the expression of HSP70 and JNK genesThis experiment consists of two parts:?1?Expression levels of HSP70 and JNK genes in B.calyciflorus at different temperatures under normal conditions;?2?after joining JNK inhibitors to B.calyciflorus,expression of HSP70 in different temperature is compared with the JNK gene.Under normal conditions,the results showed that at different times,the expression of HSP70 under 15?,20?and 30?increased first and then decreased with the aging of rotifer,and 25?assumes the declining trend.These results indicate that high and low temperature can induce HSP70.The results of the JNK inhibitor group were similar,but all temperature increased first and then decreased.Rotifers at different temperatures in the same time,we found from day 1 to day 4,the other three temperature compared with 25?have been increased obviously,but in day 5,30?than 25?decreased significantly.The results of the JNK inhibitor group were similar to those of the normal group.JNK,on the other hand,is affected by the HSP70 gene,JNK decreased when HSP70 increased.However,after the decline of HSP70,it increased with the increasing age of rotifers.JNK in the JNK inhibitor group was maintained at a low level.3.Activation characteristics of HSP70 and JNK in B.calyciflorus under four stress factorB.calyciflorus in this study experienced four kinds of stress respectively:short time and high temperature stress?40?,1h?,oxygen stress?induced by H₂O₂?,ultraviolet?UV?stress,dietary restriction?DR?,and used fluorescence quantitative analysis.The results showed that all the four stresses had an effect on the expression of HSP70 and JNK in B.calyciflorus.Under high temperature stress,the expression of HSP70 reached the highest on the second day,and then the JNK decreased with the increase of HSP70.Under the oxygen stress,the results showed that HSP70 attenuated with time under the stimulation of H₂O₂,while JNK returned to the normal level with the attenuation of HSP70.When B.calyciflorus were restricted by diet,that is,after 24h of starvation treatment,the expression of HSP70 was shown to reach the highest level at the next day,but then began to decline,the fifth day was significantly lower than the control group.The expression of JNK was similar to that of H₂O₂.The expression levels of HSP70 and JNK in B.calyciflorus were similar to that of dietary restriction under the stress of ultraviolet.These experiments indicated that the expression of HSP70 in B.calyciflorus was stimulated by these stresses,and there may have a relationship with JNK.