| Research objective:In order to find the roles of the muscle-specific miRNAs have played during theprocess of skeletal muscle injury and regeneration induced by the exercise andprovide an original evidence for the enrichment of the theory about muscleregeneration and get a new visual angle on the injury induced by exercise or thediagnogsis and therapy applying the gene technologies. In this study we applied theestablished the animal’s model of skeletal muscle injury and regeneration induced byexercise in rats from our lab; and then the dynamic expression changes ofmuscle-specific miRNAs(miR-1and miR-206) has been examined successfullyduring the process of the skeletal muscle injury and regeneration in rats; theexpression from which the transcriptional level and translational level of their targetgenes had been detected as well.Research methods:There were72healthy male Sprague-Dawley (SD) rats in8-weeks-old, weight264.44±11.94g, were randomly divided into control group, immediate,6hours,12hours,1day,2days,3days,1week and2weeks group after exercise,and8in eachgroup. Except for the control group, the rats in remaining8groups employed atreadmill training.The gradient, speed, time of the treadmill training is-16o,16m/min,90min respectively. The gastrocnemius muscle of both side legs were removed fromthe nesthetized rats by0h、6hours、12hours、1day、2days、3days、1week and2weeks after exercise respectively. We observed samples of the left legs through HEstaining, and the other ones were examined the miRNAs and their target genes byRT-PCR and Western blot.Research results:1HE staining of traverse section of gastrocnemius muscleWe could find the polygonal myofibers arranged tightly each other with generalmorphological shapes; The cells membrane were integrated while the nucleus besidesthe sarcolemma; From the group of0h, several swelling cells could be found in themuscle with augmented volumes and enlarged gaps between the neighboring cells; inthe following groups around6h、12h、1d and2d，consecutive aggravated swellingcells could be found by the decreasing the gaps compared with the control group；Bythe3d group after the exercise，the most serious swelling infiltrated cells could befound，there could be found increasing number of inflammatory cells between thegaps of neighboring cells，the red blood cells could be found as well；In the group of1w after the exercise, the number of inflitrated inflammtory cells was decreased, In the group of2w, the process of regeneration got accomplished with the shapes andstructures of the cells getting recovery.2. The expression changes of the muscle-spcific miRNAs during the injury andregeneration of skeletal muscle in rats.Compared with the control group,there were no significant changes on theexpression of muscle-specific miR-1in rats,the expression of miR-206wereupregulated after the exercise；statistical analysis shows that only the result in thegroup of1w after the exercise have significant changes when compared with thecontrol group（p<0.05）.3. The expression level of downstream target genes of the miRNAs during the processof muscle regeneration.Compared with the control group, We find that the mRNA expression level ofCx43and HDAC4was decreased in the group of0h after the exercise；The mRNAlevel in the following groups after the treadmill running were upregulated；comparedwith the control group, mRNA level of HDAC4in the group of1w has the significantdifference （p<0.05），mRNA level of Cx43in all of the groups after exercise have nosignificant difference （p>0.05).Compared with the control group, We find that the protein expression level ofCx43and HDAC4were declined in the group of0h after the exercise;we also findthere was another downregulation of the protein level in the group of3d after thetreadmill running；compared with the control group，protein level of Cx43in thegroup of0h and3d have the significant difference（p<0.05），protein level of HDAC4in all of the group after the exercise have no significant difference （p>0.05).Conclusion:1. In this work, based on the model of skeletal muscle injury and regeneration inducedby exercise in rats that had been estabilished formerly in our lab, we find that therewas little changes of muscle-specific miR-1during the process of injury andregeneration induced by the exercise,but miR-206was upregulated after the exercise，and we found the difference of Cx43in the group of0h and3d after the treadmillrunning that indicated miR-206and its target gene Cx43appeared to be involved inthe process during the injury and regeneration in the gastrocnemius muscle throughpost-transcriptional level. |
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