Time-dependent Expression Of MT1A MRNA And MT2A MRNA In The Contused Skeletal Muscle Of Rats

Objective:To investigate the expression of MTIA mRNA and MT2A mRNA changes compared with Ribosomal Protein L13 induced by contusion in rat skeletal muscle using Real-time PCR, aimed to provide some help for forensic estimation of wound age.Methods:A total of 60 Sprague-Dawley rats, weighing between 200 and 250 g, were used in this study. All animals were approved by the Animal Center of Shanxi Medical University. The rats were divided into 3 parts:control group (n=6),experimental groups (contain 0.5h,1h,6h,12h,18h,24h,30h,36h after contusion, n=6) and curative group (n=6)..After the rats were anesthetized with ethylether and the right posterior limb was shaved, a depilatory agent was applied to remove residual hair. Subsequently, the rats were fixed, and a 250g counterpoise was raised and allowed to fall freely 150cm through a clear Lucite guide tube onto the right posterior limb of rats. The samples were extracted respectively and stored in liquid nitrogen. Total RNA were isolated from skeletal muscle of three groups, and reverse transcription polymerase chain reaction was conducted to synthesize the 1-st strand cDNA. With the use of sequence-specific primers, the expression level of MT1A mRNA and MT2A mRNA was studied by SYBR Green I Real-time PCR.Results:(1)Our results normalized by Ribosomal Protein L13(RPL13)showed that the expression levels of MTIA mRNA up-regulated in the 0.5h and the peak appeared in the 18h after contusion compared with the control group. The second peak of the expression levels of MTIA mRNA appeared in the 30h after contusion.There were significant changes in the expression levels of MTIA mRNA in the other experimental groups (P<0.05), with the exception of 0.5h,compared with the control group.And there were significant changes in the expression levels of MT1A mRNA at adjacent experimental groups (P<0.05).(2) The expression levels of MT2A mRNA up-regulated in the 0.5h and the peak appeared in the 18h after contusion compared with the control group. The second peak appeared in the 30h after contusion. There were significant changes in the expression levels of MT2A mRNA in the other experimental groups (P<0.05), with the exception of 0.5h, compared with the control group. And there were significant changes in the expression levels of MT2A mRNA at adjacent experimental groups (P<0.05).(3) Although the expression levels of MTIA mRNA and MT2A mRNA after contusion were different, but both expressed the same trend. Conclusions:(1) The time-dependent expression of MT1A mRNA and MT2A mRNA after contusion within 36hs was potentially indicative for estimation of early wound age.(2) The similar expression trend of MT1A mRNA and MT2A mRNA after contusion showed that they play the same roles in the process of anti-injury response.(3) The high expression levels of MT1A mRNA and MT2A mRNA in the 21d after contusion indicated that they worked not only at the early inflammation but also at the Period of tissue repair.