Effects And Its Mechanism Of Low-Intensiy Pulsed Ultrasound On Fibrosis After Aged Mice Skeletal Muscle Injury

Background:The injury and repair of skeletal muscle is one of the hot and difficult problems in research on trauma rehabilitation. In early animal study, we found that Low-intensity pulsed ultrasound(LIPUS) can promote the energy of repair after skeletal muscle injury, promote skeletal muscle myoblast proliferation and differentiation, and inhibition of satellite cells differntiate to fibroblast. But further research is needed on how LIPUS influence the fibrosis after skeletal muscle injury.Objective:To investigate the inhibitory mechanism and effect of low-intensity pulsed ultrasound on injury skeletal muscle fibrosis in aged mices. Clinical application of LIPUS for the prevention of injury skeletal muscle fibrosis and give a theoretical reference.Methods:By using the method of operation, 30 male Kunming rats bilateral gastrocnemius were local injured. They were divided into the control group(CG) and the ultrasound group(UG) according to the method of random digits table. were treated by LIPUS(frequency 1MHz,10 min once,duty cycle 20%,average strength 40 m W/cm2, 1/d) from the day 3 after injury, and the control group without treatment. Respectively in the day 3,7,14,21 and 28 post-injury, the specimen bilateral gastrocnemius muscles were cut down after the heart perfusion fixation, and then do frozen section. Using the HE, Masson staining, observe the muscle fiber arranged, infiltration of inflammatory cells and collagen distribution; and by using immunohistochemical staining to observe Collagen I, and alpha-SMA expression and distribution; last using the statistical analysis to compare the difference of UG and CG.Result:1. HE staining observation: The day 3 after injury, a lot of inflammatory cell infiltration were found at the skeletal muscle injury site in both groups. The day 7 and 14 after injury, new muscle fibers are gradually increased in both groups, the multinucleated muscle fiber number of UG was more and more orderly than CG, interstitial cells of the CG more obvious, the infiltration of inflammatory cells more obvious. The day 21 and 28 after injury, the multinucleated muscle fiber of CG direction relatively disordered, collagen fibers more, part of the muscle fibers fused into sheets, border is not clear; then the muscle fibers of UG were arranged in uniform, myoblasts and myotubes also increased than before.2. Masson staining observation: The day 3 after injury, two groups showed a small amount of collagen present in the interstitial damage zone, and their MOD were(6.05±1.02)% and(5.71±0.76)%,it is no no significant difference in both groups.The day7,14 and 21 after injury, the collagen fibers of both groups to hyperplasia was obvious, and gradually concentrate and shrinking; The MOD reached the peak in the day 21 after injury, respectively(13.40±1.30)% and(21.32±2.83)%, and the UG was significantly less than the CG(P<0.01).The day 28 after injury, the collagen fibers of tow groups begin to absorb, shrinkage, remodeling, and the UG was significantly less than the CG at the same time(P<0.01).3. Collagen I expression: The 3 and 7 after injury, the MOD of collagen I were(5.37±0.54)%,(6.59±0.76)%å'Œ(5.13±0.64)%,(7.23±1.65)%,and it is no difference between the 2 groups(P>0.05). The day 3 to 21 after injury, the collagen type I of both groups expression increased gradually, then stabilizing, peaked at the day 21(15.83±2.13%å'Œ11.72±1.31%).The day 14, 21 and 28 after injury, the collagen I expression of UG were significantly less than the CG(P<0.01).4. α-SMA expression: The day 3 after injury, the 2 groups α-SMA MOD were(6.30 ± 0.57)% and(6.30 ± 0.57)%, no significant difference between both groups(P>0.05).The day 3 to 14 after injury, the α-SMA expression of both groups were gradually increased, slowly decrease at the day 21 and 28. In the UG, the difference was statistically significant in the day 14 and day 7 after injury(P<0.05), but it was not statistically significant in the day 21 and 14 after injury. In the CG, there was no difference in the day 21 and day 14 after injury,but there was a statistically significant difference compared in the day 21 and day 28(P<0.05). In the day 7 and 28 after injury, the expression of α-SMA in the UG is less than the CG(P<0.05); and in the day 14 and 21 after injury(P<0.01).Conclusion:1. Using the operation to cut the aged mouse gastrocnemius muscle caused defects injury, can produce the fibrosis animal model after skeletal muscle injury.2. Low-Intensity Pulsed Ultrasound therapy can inhibit aged mice skeletal muscle after injury fibrosis.3. The inhibitory effect of ultrasonic pulse therapy on injury skeletal muscle fibrosis maybe cause by it can decrease the expression of Collagenâ… and α-SMA.