Date of Award
Campus Access Dissertation
Matthew C Kostek
Skeletal muscle injuries are common in athletic and non-athletic populations and affect muscle function, mobility, and thereby quality of life; muscle rehabilitation constitute a challenge in physical therapy and sports medicine. Therapeutic ultrasound (TUS) is one of the most commonly used modality in muscular injuries in order to reduce edema and pain, and accelerate tissue repair/regeneration, however the mechanisms of action of TUS are not clear yet. In addition, there is little clinical evidence from to support that TUS enhances regeneration of skeletal muscle following injury. The aim of this study was to determine the effects of TUS at a set combination of parameters on muscular performance and regeneration program after exercise-induced damage, and its autonomous effects on muscle cells proliferation, differentiation and growth. Two models were used for the purpose of this study, human (healthy, college-aged individuals) and cell culture of muscle cells (C2C12). Three components of muscular performance of healing human skeletal muscle were examined in response to 7 consecutive TUS treatments. Satellite cell myogenic program in humans (females and males) in response to TUS was examined 6h after TUS treatment by gene expression of myogenic regulatory factors and immunofluorescence for satellite cell (SC) counting. The autonomous effect of three consecutive TUS treatments on muscle cells was examined during proliferation, by cell counting and densitometry, and differentiation by determination of fusion index, myotube diameter and number of myonuclei per myotube. Protein synthesis and protein degradation were examined by relative protein phosphorilation of P70S6K and protein levels of MURF-1. During muscle healing process, TUS enhanced the muscle's capacity to contract throughout the range of motion (work (J)) compared to sham treatment (p=0.02). Acutely, TUS regulated the myogenic program of satellite cells based upon the skeletal muscle's tissue status and gender. In healing muscle, TUS cancelled the effect of exercise in regards the number of SCs in males and females (no differences between TUS treated vs. untreated, p=0.9), increased the expression of cyclin-D1 (1.9±0.3-fold, p=0.008) in males and down regulated gene expression of p21(1.59±0.3-fold, p=0.005) in females. In response to TUS alone, females exhibited increase in the number of activated/proliferating satellite cells of 80% (MyoD positive cells) (p=0.01) and males exhibited an increase of 122% (p=0.001). In damaged skeletal muscle, TUS did not induce change in the number of satellite cells in either females or males for any of the variables examined (p=0.09). Correlation analysis showed that undamaged muscle Cyclin D1 has a negative correlation (r=-0.48, p=0.04) with the number of MyoD positive cells following TUS treatment; and p21 expression showed a positive correlation (r=0.72, p=0.02) with the number of MyoD positive cells. In contrast, in damaged muscle Cyclin D1 showed a positive correlation (r=0.73, p=0.03) with the number of MyoD positive cells following TUS treatment (figure. 4.5E); whereas p21 expression and the number of MyoD were not correlated (r=0.05, p=0.1) (figure. 4.5F). In muscle cells in-vitro, 3 consecutive days of TUS treatment increased total number of muscle cells (1.6±0.05-fold compared to sham and control group (p=<0.03)), increased myotube diameter (2.2±0.1-fold, p=0.01) and mean number of myonuclei per myotube (2±0.02-fold, p=0.01). In differentiating myoblasts, TUS increased relative levels of p-p70s6k (5.7±0.2-fold, p=<0.001) and decrease in relative levels of MURF-1 protein (3.4±0.1-fold, p=0.04). No change in the media temperature was induced by TUS in-vitro. We conclude that TUS positively impact muscle function after exercise-induced damage, and regulates the myogenic program of satellite cells. TUS constitute a potent mechanical stimuli for muscle cells proliferation and growth, with a concomitant induction of markers of protein synthesis and inhibition of markers of protein degradation.
DELGADO DIAZ, D. C.(2012). Functional and Molecular Effects of Therapeutic Ultrasound On Skeletal Muscle Repair.. (Doctoral dissertation). Retrieved from http://scholarcommons.sc.edu/etd/1204