Therapeutic Ultrasound

Ultrasound (US) has long been used in many medical specialties as an imaging modality and it has seen limited use in a therapeutic capacity by allied health practitioners. However, US is not commonly used as a therapeutic modality among medical professionals.
The impetus for using US for treating muscular injuries was based on evidence demonstrating the efficacy of US in decreasing bone healing time following fractures and facilitating healing in non-unions. It has been postulated that US promotes healing in bone through a combination of mechanical loading, generation of pressure waves and localised heating; these then go on to effect various cellular and biochemical changes favouring the normal healing of bone. Healing of muscular injuries follows a sequence of inflammation, regeneration and repair/remodelling; healing of bone follows a similar pattern of inflammatory reaction, regeneration, consolidation and remodelling. Multiple mechanisms mediate and regulate these processes, and these may be amenable to alteration in muscle tissues, as they are in bone.Rosenburg A. Bones, Joint and Soft Tissue Tumours. In: Robbins and Cotran Pathologic Basis of Disease. Philadelphia: Elsevier Saunders; 2005. p. 1273–324 
This work will aim to review literature of evidence for the use of US in muscle injury, including in vitro and in vivo evidence as well as clinical evidence for its use. Additionally, this work will discuss potential advantages of therapeutic US in the context of general practice and musculoskeletal medicine in New Zealand. 
== In vitro evidence ==
Mouse-derived C2C12 cells are an in vitro model of myoblast cells, which are important in muscle healing as they are able to fuse with injured myofibrils to assist in healing. TNF-á and IL-1â are cytokines involved in the inflammatory process and induces expression of cyclooxygenase-2 (COX-2), a key inflammatory enzyme. In one study, C2C12 cells primed with TNF-á or IL-1â were exposed to a pulsed US signal at 3 MHz for 15 minutes, pulsed 2s on and 8s off with an average energy intensity of 30 mW/cm2. These cells had a significant decrease in expression of COX-2 mRNA at three hours after stimulation, when compared to control cells. Additionally, the cells exposed to US had an increased expression of myogenin mRNA, which is associated with differentiation of myoblasts to myocytes.
The authors concluded that in this in vitro model, US attenuates the initial inflammatory response so as to allow regeneration and repair to occur. Additionally, US promotes the differentiation of myoblasts to mature myocytes, further promoting regeneration and repair.
== In vivo animal evidence ==
In an experimental animal model of muscle damage, the left tibialis anterior muscles of 48 C57BL/6 mice were damaged by low dose myotoxic snake venom in vivo. These muscles were then treated with US 24 hours after exposure to the snake venom; they received treatment for 15 minutes at 1 MHz for 15 minutes, pulsed 2s on and 8s off with an average energy intensity of 150 mW/cm2. US treatment continued for one, three, five or seven days after injury. The right tibialis anterior muscle of each mouse was left undamaged as a control. The muscle tissue underwent histological examination for evidence of expression of paired-box transcription factor 7 (Pax7), a marker of satellite cell activation, using immunofluorescent tagging. The study demonstrated that there was significantly increased expression of Pax7 at day 5 in treated muscle compared to untreated muscle. The authors concluded that US increases satellite cell activation following injury. 
US-mediated increase in activation of satellite cells have also been demonstrated in a rat model of muscle injury. This study evaluated the effect of US following laceration of the gastrocnemius muscle in Wistar rats. The treatment group underwent surgical laceration of the muscle, followed by daily US treatment (1 MHz with an intensity of 0.57 W/cm2, pulsed at 5 ms intervals) for five minutes starting two days after the injury; the control group only underwent surgery. Desmin was used as a marker of satellite cell activation and early formation of skeletal muscle; histological evaluation of treated muscle at day 14 after injury demonstrated a statistically significant increased expression of desmin when compared to control muscles. Additionally, this study demonstrated that injured muscles treated with US had significantly denser and better-oriented collagen fibres (aligned parallel to the connective sheaths of surviving and repairing myocytes) at day 7 after injury, compared to the control group, but this effect was not observed at day 14. The authors concluded that US treatment promotes activation of myoblasts and helps orient collagen fibres more appropriately; they hypothesised that the better orientation of collagen promotes biomechanical recovery of injured muscle. 
A potential mechanism by which US modulates healing in muscle is the expression of heat shock proteins (HSPs). These are a group of proteins that act as cytoprotective mediators and are expressed in response to protein-damaging stimuli. HSPs may act as chaperone molecules by modulating protein synthesis, post-translation modifications of protein and degradation.Kumar V, Abbas AK, Fausto N. Cellular Adaptation, Cell Injury and Cell Death. In: Robbins and Cotran Pathologic Basis of Disease. Philadelphia: Elsevier Saunders; 2005. p. 3–46. One rat model explored the expression of HSPs following US treatment in a non-injury experiment in nine Sprague-Dawley rats. The focus was placed on HSP 25 and HSP 72, which are known to be cytoprotective and inhibit apoptosis in cells injured by heat, cold and chemical agents. The gastrocnemius, plantaris and soleus muscles on one side were exposed to either pulsed US (1 MHz at intensity of 2.0 W/cm2 at 50% cycle) or continuous US (1 MHz at intensity 1.0 W/cm2) for 15 minutes on four consecutive days. The contralateral muscles were used as controls. Core and muscle temperatures were monitored throughout treatment and to ensure there was no increased temperature due to US treatment.
The study demonstrated that only pulsed US significantly increased the expression of HSP 25 in all muscles. Whilst there was an increased expression of HSP 72 in plantaris muscles treated with both pulsed and continuous US, increased expression of HSP 72 was only observed in gastrocnemius and soleus of muscles treated with pulsed US. The authors concluded that US increases expression of beneficial HSPs even in the absence of injury; the study found that there was insufficient temperature increase with either form of US to induce the expression of the HSPs in a temperature-dependent manner. Other in vivo studies had speculated that altering expression of HSPs may be the mechanism by which US modulates the inflammatory process in injured muscle.
== In vivo human studies ==
One experimental human model of muscle injury examined the difference in expression of insulin-like growth factor (IGF) in muscle tissue following injury; IGF-1 has been previously demonstrated to increase satellite cell activation and promote muscle repair by increasing protein synthesis and myotube differentiation. This study had 16 participants undergo 200 bilateral quadriceps femoris contraction; muscle damage was confirmed by a rise in plasma creatine kinase and decreased force production. Forty-eight hours after the damaging event, one vastus lateralis in each participant was treated with US (continuous cycle, 1 MHz at 1.5 W/cm2 for 10 minutes). There was a control group who did not undergo the muscle contractions but did have unilateral US treatment. Bilateral muscle biopsies were then taken six hours after treatment and mRNA expression of IGF-1 was assessed by polymerase chain reaction. This study demonstrated a significantly increased expression of IGF-1 mRNA, particularly that of the IGF-1Eb splice variant, in only the treated leg of the control group; there was no statistical difference in IGF-1 expression between treated and untreated muscles in the group that underwent injurious activity. This study demonstrated that US does not have an effect on IGF-1 expression in muscle injured by repeated contractions, but there was nevertheless some effect on muscle. This led the authors to hypothesise that US may not be useful for the study population, who were healthy and active men aged 18-29, but there were effects on muscle that may be useful in other patient populations.
== Clinical studies ==
A small trial investigated the efficacy of therapeutic US in modulating the pain sensitivity of myofascial trigger points in trapezius. The investigators had two cohorts of 22 participants aged 28-65, identified as having trigger points, with pain threshold measured by constantly applying an increasing pressure to the trigger points, at a rate of 5 N/s, until participants reported pain. The treatment group then underwent continuous US treatment to the trigger points at 1 MHz, intensity of 1.0 W/cm2 for five minutes; the control group also underwent continuous US at the same frequency but at a much lower intensity of 0.1 W/cm2. The authors then found that the treatment group had a significantly greater tolerance of force applied to trigger points in the period immediately after treatment, whereas the control group did not demonstrate any significant difference. They concluded that therapeutic US increases pain threshold in those with trapezius trigger points in the short term.
== Discussion ==
Evidence demonstrates that US has an effect on an in vitro model of myoblasts even in the absence of injury, which creates biological plausibility for further investigating US as a specific therapeutic modality. In vivo studies of two mammalian models of injury have demonstrated that US promotes satellite cell activation and helps orient collagen fibres in a manner that promotes recovery of biomechanical performance. Potential molecular mechanisms have also been identified, in that US may attenuate the initial inflammatory process to promote healing. The inflammatory response is important early in the repair process because it promotes clearance of cellular debris by macrophages and increases blood flow to the area to allow for healing, among other actions. But a persistent inflammatory response can inhibit efficient regeneration of the contractile mechanism. US also increases expression of beneficial HSPs in a temperature-independent manner, which aids correcting folding of expressed protein, which may be important in reconstitution of muscular contractile mechanisms. Experimental human studies demonstrated that pulsed US does increase expression of IGF-1 in muscle, but only in a non-injurious state, suggesting there is a role for therapeutic US, but not in injuries due to extensive muscular contractions. However, there is little clinical evidence for the efficacy of US in a clinical setting. The author of this work was only able to find one small scale clinical trial demonstrating that US has an antinociceptive effect on trigger points in trapezius muscle. 
It is difficult to gauge which lesion US would be most effectively used in based on the animal models. The murine study caused injury using a snake venom, whereas the rat model used a laceration injury. The experimental human model was different again, using excessive contractions and the clinical study investigated trigger points. The aetiology of trigger points was never described in this study and therefore it can only be speculated whether their existence was due to a true muscular injury. Heterogeneity of the injury mechanisms means there is insufficient data to determine an indication for the use of US in a clinical setting, which is further exacerbated by the nature of the lesions investigated in the human studies. 
Another barrier is the marked differences in the sonographic parameters of US used in the studies. All in vivo studies, both animal and human, used 1 MHz, whereas the in vitro study used 3 MHz. The former frequency was likely used by investigators as it has been demonstrated to penetrate tissues to a depth of 5cm, whereas a frequency of 3 MHz is more effective at more superficial penetrations. The intensities used ranged from 0.03-2.0 W/cm2; the former being used in the laboratory model of myoblasts and the latter being used in a rat injury model. An effective intensity is therefore difficult to gauge.
The single clinical study included for consideration in this work used a low-intensity US (0.1 W/cm2) as it’s control. However, as in vitro studies have demonstrated, a significantly lower intensity (0.03 W/cm2) can have an effect on satellite cells. Therefore, a criticism of this clinical study is that it did not compare therapeutic US to a true sham. Additionally, this study did not blind the investigators to the intervention being performed, which raises the possibility of introducing inadvertent bias. Other issues with this study are that this did only examine the immediate effects of US and that there was no apparent radiological standardisation of the lesion (e.g., size, location within muscle) being investigated, albeit experienced clinicians were used to identify the lesion. As discussed above, the aetiology of these trigger points is not known. This study also examined a very subjective endpoint with no objective measure of change. 
Further work is needed to determine the utility of therapeutic US in treating muscle injuries. Potential barriers would include enrolling patients with comparable demographic characteristics (e.g., age, gender, activity levels and comorbidities), injury characteristics (e.g., timing, location, size) and pain tolerances into the study.  There would have to be sufficient numbers of comparable patients to allow for adequately sized treatment and control groups so as to provide statistical power. In addition to assessing objective parameters (e.g., location and size of lesion), the study will need to utilise validated tools to determine clinical improvement (e.g., pain, mobility). The study would also have to be of long enough duration to determine the effectiveness of therapeutic US.
The advantage of therapeutic US is that it is a potential bedside treatment that can enhance healing and promote return to work and pre-morbid functioning. The patient will be able to access community-based treatment which may be delivered at the patient’s local medical practise; as injuries sustained in New Zealand are covered by the Accident Compensation Corporation, the patient may also experience little or no financial barriers to accessing treatment that may hasten return work and normal functioning. 
== Conclusion ==
In vitro, in vivo animal and in vivo human studies have demonstrated a potentially beneficial effect of US on muscle tissue. However, there is little clinical evidence demonstrating its efficacy, with significant barriers to being able to determine the utility of therapeutic US in treating muscle injuries. Identifying indications, doses and durations of treatment using therapeutic US would allow for development of community-based treatment options that can help patients suffering muscle injuries return to pre-injury function. New Zealand’s accident compensation infrastructure would allow for viable treatment in the community with little or no financial barriers to accessing treatment.
== References ==
Category:Miscellaneous