Keywords

10.1 Tendon: Anatomy, Injury, and Healing

10.1.1 Function and Anatomy

Tendons are dense, fibrous connective tissues that link muscle to bone and are critical to the overall function of the musculoskeletal system. While the primary function of tendons is to efficiently transmit tensile forces from muscle to bone, they are also subjected to compression and shear forces. Mature tendon is characterized by its hierarchical structure, and its mechanical function is dependent upon the biochemical composition and organization of its extracellular matrix (ECM) (Voleti et al. 2012; Screen et al. 2015; Snedeker and Foolen 2017). Tendon ECM is primarily composed of fibrillar collagens, proteoglycans, glycosaminoglycans, glycoproteins, and elastin (Sharma and Maffulli 2006) (Fig. 10.1). Collagen is the primary constituent of the tendon ECM, accounting for 60 to 85% of the dry weight of the tissue (Screen et al. 2015). Type I collagen is the dominant structural component, constituting nearly 95% of the total collagen content (Screen et al. 2015), while collagen types III, V, XI, XII, and XIV are present in much smaller proportions (Benjamin et al. 2008; Screen et al. 2015). The typical, highly organized collagen matrix can become disrupted in chronic injuries such as tendinopathy, while the presence of collagen III, which typically amounts to 3–5% of the total collagen in a healthy tendon, may be elevated in tendinopathic tissue (Snedeker and Foolen 2017). Non-collagenous ECM components include: proteoglycans (whose hydrophilic nature allows for rapid diffusion of water-soluble molecules and cell migration), glycoproteins such as fibronectin (which contributes to tendon repair and regeneration processes), and elastic proteins such as Tenascin-C (which is associated with collagen fiber alignment and orientation, and is upregulated by mechanical strain) (Mackie and Ramsey 1996; Mehr et al. 2000) (Fig. 10.2).

Fig. 10.1
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Tendon hierarchical structure (reproduced with permission from Marqueti et al. (2019))

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Fig. 10.2
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Interactions of proteoglycans and other matrix macromolecules within the extracellular matrix of the tendon proper (reproduced with permission from Parkinson et al. (2011))

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Tendon cell populations are heterogeneous and contribute to ECM dynamics and homeostasis (Costa-Almeida et al. 2019; Zhang et al. 2019). Tenoytes, also referred to as resident cells, are fibroblast-like cells that are mainly responsible for ECM turnover, collagen production and assembly. They are arranged in longitudinal rows proximal to the collagen fibrils (Benjamin et al. 2008). Additionally, tendon stem/progenitor cells (TPSCs) replenish tendon cells by undergoing self-renewal and differentiation (Bi et al. 2007; Zhang and Wang 2010). Other cell populations include endothelial and synovial cells of the tendon sheaths as well as chondrocytes which are present within tendon-bone insertion sites and in tendon regions subjected to compressive forces during physiologic loading (Benjamin and Ralphs 1998; Kannus 2000; Zelzer et al. 2014). Tenocytes are mechanosensitive, and upon experiencing mechanical load, they stretch along the collagen fibrils longitudinally, signaling collagen production. By modulating their alignment and signaling in accordance with their mechanical environment they alter ECM composition and structure (Humphrey et al. 2014; Muller et al. 2015; Popov et al. 2015). While little is known regarding the optimal loading conditions that may positively influence tendon healing, understanding how biophysical stimulation influences healing is critical to developing therapeutic treatments to restore pre-injury function.

10.1.2 Tendon Injuries and Healing

As tendons are frequently subjected to continuous or intermittent high magnitude forces, these tissues are prone to both acute and chronic injuries. Such injuries are debilitating, and are associated with ineffective healing, long-term pain, and loss of function (Nourissat et al. 2015). The type, severity, and prevalence of tendon injuries are dependent on multiple factors such as age, sex, activity levels, and genetic disposition (Sharma and Maffulli 2006; Thomopoulos et al. 2015). While tendon injuries are predominant in the elderly and athletic populations, they are becoming increasingly prevalent in the general population, due to increasing life expectancy, manual labor, and the popularity of strenuous loading activities such as exercise. Worldwide, more than half of the sport-related injuries involve tendons, and tendon damage is the most common orthopedic soft tissue injury (Walden et al. 2017). Apart from being highly prone to injury, tendons generally have a poor intrinsic capacity for healing, although the latter is dependent on the anatomic location and local environment (Thomopoulos et al. 2015). Intrasynovial tendon injuries do not exhibit spontaneous healing, while extrasynovial tendon injuries often result in fibrous tissue formation owing to a robust, scar-mediated healing response post-injury (Shen et al. 2021). Following its intrinsic repair response, tendon exhibits material properties which are inferior to those of native, uninjured tissue (Muller et al. 2015; Nourissat et al. 2015). Surgical and conservative medical interventions have limited, short-term efficacy and thus, there is significant motivation for the development of alternative approaches to improve tendon healing. Tendon injuries can be broadly categorized as acute or chronic, the former being the result of a “macro-trauma” such as a sudden mechanical overload, leading to a partial or complete rupture of the tendon. Chronic injuries typically present with an absence of inflammatory cells due to multiple stressors, including metabolic, biomechanical, genetic, and hypoxic factors; the latter may induce cellular responses that lead to disruption of matrix organization, loss of tissue material properties, and disrupted cell-matrix mechanotransduction (Nourissat et al. 2015; Sayegh et al. 2015). The etiology of chronic tendon disorders is multifaceted, and the subsequent degenerative pathway is triggered by dysregulated cell-cell and cell-matrix communication.

A primary etiologic factor in tendinopathies is repeated mechanical loading which exceeds the tendon’s ability to heal (Steinmann et al. 2020; Millar et al. 2021). Although a biomechanical loading event individually may be of a magnitude within physiological limits, cumulative microtrauma from repetitive loading often leads to localized collagen fiber damage (Herod and Veres 2018; Leek et al. 2020; Steinmann et al. 2020). When subjected to these stresses, a tendon may experience either inflammation of its sheath or degeneration of its body, or a combination of both (Sharma and Maffulli 2006). Recent evidence suggests the role of an inflammatory response in mediating tendon pathophysiology (Millar et al. 2017; Sunwoo et al. 2020; Millar et al. 2021). Specifically, various immune cell types (mast cells, macrophages, T cells) and inflammatory cytokines (interleukin (IL)-6, IL-5, IL-17, IL-18, IL-33, tumor necrosis factor alpha (TNF-α)) have been identified to play a critical role in the initiation and progression (early stages) of chronic tendon injuries (Millar et al. 2010; Garcia-Melchor et al. 2021). Interactions between resident and infiltrating immune cells and resident tenocytes are important in directing the inflammatory response phase of tendon healing, via secretion of cytokines and chemokines that regulate extracellular matrix remodeling. Pro-inflammatory cytokines (including members of the IL- and TNF-families) have been implicated in tendinopathy and are associated with immune cell recruitment, increased collagen type III and reduced collagen type I production, and reduced tendon biomechanical strength (Lin et al. 2006; Legerlotz et al. 2012; Dakin et al. 2014; Millar et al. 2017). Very recently, Garcia-Melchor et al. (2021) reported that tenocytes upregulate the genes involved in inflammation and T cell recruitment in vitro. T cell–tenocyte interactions, in turn, resulted in the upregulation of inflammatory cytokine expression and an increased expression of collagen III. It has been proposed that this autoregulated feedback loop plays a key role in chronicity and long-term complications of tendinopathy.

Largely due to the challenges presented in studying human tendinopathy, including the difficulty in identifying the onset of the disease as well as in procuring injured tissues at different stages of the post-injury response (Dirks and Warden 2011; Hast et al. 2014), our understanding of the precise mechanisms of tendon injury and healing remains incomplete. Tendon healing has primarily been studied using animal models of acute tendon injury (e.g., transection) or other experimentally induced tendon damage/injury models (Sharma and Maffulli 2005; Docheva et al. 2015). Tendon healing consists of sequential and overlapping phases (Docheva et al. 2015; Nourissat et al. 2015) including inflammation, cell proliferation, migration, and remodeling (Voleti et al. 2012; Docheva et al. 2015); however, it is common for incomplete healing to result in fibrovascular scar tissue which does not recapitulate native composition and material properties (Nourissat et al. 2015).

In order to study mechanisms of mechanical “overload” on the development of tendinopathy, researchers have developed a variety of preclinical approaches (Thomopoulos et al. 2015; Theodossiou and Schiele 2019). Some of these methods include uphill and downhill treadmill running in rats or mice (Heinemeier et al. 2012; Pingel et al. 2013; Zhang et al. 2020) and application of controlled, in vivo fatigue loading to rat or mouse tendons (Fung et al. 2010; Andarawis-Puri et al. 2012; Sereysky et al. 2012). Furthermore, biochemical induction of tendon injury has been studied using collagenase injections in various models including rabbits, sheep, and rats (Chen et al. 2014; Lacitignola et al. 2014; Solchaga et al. 2014; Urdzikova et al. 2014). Surgical repair following tendon transection has been particularly useful in studying acute injury healing mechanisms (Yoshida et al. 2016; Moser et al. 2018). In vitro cell culture and tendon explant models (Goodman et al. 2004; Wunderli et al. 2020), as well as ex vivo rodent and equine tendon models, have been used to examine the effects of repetitive mechanical loading (e.g., cyclic strain or fatigue) on tendons (Arnoczky et al. 2007; Fung et al. 2009; Spiesz et al. 2015). Alteration of the tendon mechanical loading environment (e.g., by transection) can be effectively used to study biochemical responses (Maeda et al. 2011). An in vivo tendinopathy model was developed (Bell et al. 2013a) by injecting TGFβ-1 into adult mouse Achilles tendons. This injury model induces tendinopathic changes consistent with human histopathology (Bell et al. 2013a, b) and is amenable to therapeutic mechanical interventions (Bell et al. 2013a; Rezvani et al. 2021) simulating human treatments (Heinemeier et al. 2012; Dirks et al. 2013; Pingel et al. 2013; Reuther et al. 2013).

10.1.3 Mechanotherapy for Treatment of Chronic Tendon Disease

There exist several common, conservative, and surgical approaches for the treatment of chronic tendinopathies, such as rest and immobilization, anti-inflammatory drugs, growth factor injections (i.e., platelet-rich plasma), and surgical repair (Lim et al. 2019; Tsai et al. 2021) (Table 10.1). However, there is limited evidence of their long-term efficacy (Maffulli et al. 2010; Cardoso et al. 2019). The molecular mechanisms of disease initiation and progression, as well as the reasons for a failed healing response in lieu of restoration of tissue, are not well understood (Tsai et al. 2021). However, it is hypothesized that dysregulated and/or missing cues underlie the deficient healing response of a tendon; hence, a detailed understanding of such cues and mechanisms will greatly assist in the identification and design of novel therapeutic strategies to augment existing strategies to heal tendons. Rehabilitation protocols aim to robustly repair injured tissues in a manner that reduces their risk for reinjury (Gray and Brolinson 2001). This strategy involves the design of therapeutic modalities and rehabilitative exercises that address the type of injury (acute vs. chronic), symptoms, and tissue performance, via an in-depth understanding of tissue biomechanics and pathophysiology of injury (Gray and Brolinson 2001). Rehabilitation protocols generally utilize “mechanotherapy” to induce adaptation of the musculoskeletal tissues to mechanical forces and/or strain by directing cellular and molecular responses to achieve healing and/or regeneration. Identifying optimal mechanical loading regimes defined by transcriptional, molecular, and cellular responses is crucial in designing strategies for tendon repair and healing. For example, eccentric exercise (lengthening of the muscle and tendon while under load) is commonly used as a therapeutic modality to manage tendinopathy. These exercises have been shown to improve tendon structure and mechanical properties with corresponding improvements in clinical outcomes (Mafi et al. 2001; Fahlstrom et al. 2003; Yu et al. 2013) and have emerged as the most efficacious therapy across numerous tendinopathies (Kingma et al. 2007; Murphy et al. 2018; Irby et al. 2020; Vander Doelen and Jelley 2020). Given that the goal of tendinopathy treatments is to restore normal tendon function, controlling the mechanical cues directed to the injured tendon can potentially promote healing via mechanotransduction mechanisms (Özer Kaya 2020). Regenerative rehabilitation approaches are, thus, central to translational tendon healing research (Gottardi and Stoddart 2018; Rando and Ambrosio 2018; Willett et al. 2020).

Table 10.1 Advantages and limitations of existing treatments for chronic tendon injuries
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10.2 Therapeutic Ultrasound and Tendon Healing

10.2.1 Physical Principles, Characteristics, and Modalities

Therapeutic Ultrasound (TUS) utilizes acoustic pressures and/or intensities higher than those of diagnostic ultrasound to elicit biological responses in tissues. The ultrasound beam is directed into a specific area or a region within the tissue of interest to avoid damage to surrounding tissues. Existing TUS methods (e.g., in physiotherapy to provide deep heating to tissues such as tendons, ligaments, and skeletal muscles (Watson 2008) deliver low-intensity energy through the targeted tissue via the propagation of sound waves applied by an external source. The most common musculoskeletal application for TUS to date remains for pain and physiotherapy in conditions such as osteoarthritis-related knee pain, chronic back pain, lateral epicondylitis, and myofascial pain (Dedes et al. 2020; Gulati and Ottestad 2020; Petterson et al. 2020). Within the categorization of TUS, multiple ultrasound modalities have been developed and examined with various delivery modes, acoustic pressures, and duty cycles, to elicit different biological mechanisms in tissues (Fig. 10.3). Several examples include low-intensity pulsed ultrasound (LIPUS) (Warden et al. 2008; Hsu and Holmes 2016; Tanaka et al. 2020), low-intensity continuous ultrasound (cLIUS) (Lucchetti et al. 2020; Mittelstein et al. 2020), and pulsed focused ultrasound (pFUS) for soft tissue healing (Burks et al. 2011; Poliachik et al. 2014), nanoparticle delivery (O’Neill et al. 2009; Tharkar et al. 2019), physiotherapy for pain relief by providing deep heating to soft tissues (typically combined with physical therapy) (Brown et al. 2015; Papadopoulos and Mani 2020) and high-intensity focused ultrasound (HIFU) for thermal (tHIFU) ablation (Dubinsky et al. 2008; Vidal-Jove et al. 2015; Mauri et al. 2018) and non-thermal (histotripsy) tissue ablation (Vlaisavljevich et al. 2013; Bader et al. 2019; Xu et al. 2021).

Fig. 10.3
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Schematic of waveforms (different amplitudes) illustrating different modalities of TUS (Modified with permission from Liu et al. (2020))

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A variety of labels have been utilized in published literature to describe different types of therapeutic ultrasound, with some overlap between commonly used terms. In general, these groups are broadly categorized based on the ultrasound intensity (high vs. low) and exposure mode (continuous vs. pulsed) (Liu et al. 2020). For clarity, in this chapter, we use the acronym HIFU to describe High Intensity Focused Ultrasound exposures that induce thermal and/or non-thermal irreversible changes within a short time frame (typically, it requires microseconds for direct effects during the pulse or histotripsy cavitation, milliseconds to elicit physiological responses such as in neuromodulation, to seconds or minutes for thermal changes). FUS refers to focused ultrasound methods (see Sect. 10.3 below), while TUS is used to describe low-intensity ultrasound exposures used currently for musculoskeletal physiotherapy and pain relief applications, as noted above. Different forms of TUS can be further categorized by understanding its basic physical parameters. A brief description of ultrasound parameters critical to the generated bioeffects and safety is given in Table 10.2.

Table 10.2 Ultrasound parameters
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Thermal bioeffects of ultrasound result from absorption of the applied ultrasonic energy. The amount of absorption and the accompanying heat generated depend upon ultrasound acoustic parameters as well as tissue properties. The primary determinants of thermal effects in tissues include tissue absorption coefficients and ultrasound exposure conditions, such as duration, intensity, beam width, and frequency. Importantly, there exists a direct relationship between the absorption capacity of a tissue and its protein content. Highly collagenous tissues such as tendon and ligament are known to absorb ultrasound energy more efficiently (Watson 2008). Other determinants of tissue temperature changes include ultrasound pulse repetition frequency and pulse duration, along with tissue characteristics such as density, acoustic impedance, and thermal conductivity (Dalecki 2004; Shankar and Pagel 2011).

Research suggests that an increase of temperature between 1 °C and 4 °C from baseline can provide therapeutic effects in tendons; however, higher elevations could potentially result in harmful effects such as thermal denaturation of collagen (Vlaisavljevich et al. 2015). An in vivo study investigated the ability of ultrasound to heat human patellar tendon and found that ultrasound frequency, intensity, duration of treatment, and size of the treatment area influenced heat production in tendon (Chan et al. 1998). The rate of temperature rise was found to be higher in the tendon compared to the adjoining muscle. Using a 3-MHz continuous ultrasound treatment lasting 4 minutes, temperature rises in the range of 8 °C to 10 °C were achieved using templates measuring two times the effective irradiation area of the transducer head. Such controlled thermal effects via TUS application are desirable to achieve pain relief, increased blood flow, decreased joint stiffness, and hyperdynamic tissue metabolism (Watson 2008; Papadopoulos and Mani 2020). Typically, physiotherapists utilize thermal effects of TUS to treat injuries such as tendonitis, joint pain, low back and neck pain, muscle strains, plantar fasciitis, ligament sprains, and arthritis pain (Brown et al. 2015; Papadopoulos and Mani 2020).

A combination of non-thermal effects such as acoustic streaming, acoustic cavitation, and radiation force displacement (Dalecki 2004; Izadifar et al. 2017) can also be produced by TUS or FUS application. Acoustic cavitation is the stable oscillation (inertial cavitation) or collapse (non-inertial cavitation) of a gas bubble in the presence of an acoustic field (Holland and Apfel 1990; Bader et al. 2019). Acoustic radiation force is that which results from momentum transfer from the sound field to the tissue of interest (Nightingale 2011; Urban 2018; Wang 2018). This effect itself is a consequence of radiation torque and acoustic streaming. The acoustic streaming phenomenon occurs when acoustic field propagation induces an increased rate of fluid flow (Dalecki 2004).

TUS is an appealing method to safely transfer mechanical energy to tendon tissue and elicit thermal and mechanical stimulation pathways in a prescribed manner. Although the therapeutic efficacy of TUS has been demonstrated in multiple studies, there is a substantial knowledge gap in understanding relationships between ultrasound dose (acoustic parameters) and the bioeffects elicited in these tissues. Optimal identification of TUS parameters and their correlation with molecular and tissue-level responses will largely benefit future research in ultrasound-induced tissue regeneration and rehabilitation.

10.2.2 TUS in Tendon Healing

The most widely studied form of TUS for musculoskeletal tissue repair and regeneration is low-intensity pulsed ultrasound (LIPUS), with applications in osteoporosis, fracture healing, mesenchymal stem cell recruitment and homing, and tendon–bone junction healing (Warden et al. 2008; Khanna et al. 2009; Zhang et al. 2017; Tanaka et al. 2020). Targeted application of TUS during different phases of tissue repair can produce a synergistic effect on healing (Saber and Saber 2017). During the inflammatory phase, TUS can stimulate mast cells, platelets, and macrophages, activating inflammatory mediators (Maxwell 1992; Leung et al. 2004). Efficiency of the proliferation phase of healing is also enhanced by TUS, by increasing collagen production and scar tissue formation (Zhou et al. 2004; Watson 2008). Lastly, TUS has also been shown to enhance the remodeling of scar tissue by improving collagen fiber orientation and increasing tensile strength (Nussbaum 1998; Maan et al. 2014).

The therapeutic potential of TUS, specifically low-intensity ultrasound (LIUS), in stimulating healing of acute tendon injuries has been investigated predominantly in animal studies, which enable the concurrent evaluation of tissue biomechanics and physiological responses to LIUS application (Table 10.3). Biomechanical metrics such as ultimate load, tensile strength and energy absorption, and structural metrics such as collagen organization and aggregation have been commonly characterized after applying treatments to assess the efficacy of TUS on healing (Ng et al. 2003; Yeung et al. 2006; Jeremias Junior et al. 2011). A 2016 review by Best et al. summarized the effects of LIUS on tendon, tendon–bone junction, muscle and ligament injuries (Best et al. 2016). The authors concluded that LIUS improves tendon strength and accelerates collagen formation after acute injury in preclinical models. Tensile strength and collagen expression (types I and III) were found to be greater in LIUS treated tendons compared to untreated controls (Jackson et al. 1991; da Cunha et al. 2001; Fu et al. 2008, 2010; Jeremias Junior et al. 2011; Kosaka et al. 2011). Regarding treatment time and duration (Fu et al. 2008, 2010), ultrasound treatment in the earlier (relative to later) stages of healing appears to improve tensile strength and matrix synthesis. While the available literature generally indicates that LIUS enhances biomechanical and structural properties of injured tendons, ultrasound parameters (e.g., intensity, stimulation frequency, and mode) and animal models (species, tendon of interest, and injury type) have varied across studies (Tsai et al. 2011). In consideration of these inconsistencies, direct comparison of results is likely not warranted and researchers should exercise caution in deducing the cellular and molecular mechanisms attributable to treatments. Hence, there is not only an urgent need for standardization of TUS parameters and experimental conditions (treatment time, injury models) in future investigations, but also clinically relevant animal models whose tendon injury characteristics mirror those observed in human injuries.

Table 10.3 Results of studies investigating therapeutic ultrasound (TUS) treatment of tendon injuries
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Clinical studies have examined the influence of TUS on lateral epicondylitis (D’Vaz et al. 2006; Dedes et al. 2020), patellar tendinopathy (Warden et al. 2008; de Jesus et al. 2019) and Achilles tendinopathy (Hsu and Holmes 2016). A 2015 study on the short-term effectiveness of LIPUS on human Achilles tendinopathy revealed that all participants who had undergone traditional, nonsurgical treatment modalities prior to using LIPUS, showed good to excellent improvements in pain relief and function post-treatment (Hsu and Holmes 2016). In three studies that examined the influence of TUS on epicondylitis and patellar tendinopathy, a significant decrease in tendon pain was observed after daily treatments with continuous or pulsed LIUS treatment for 6 weeks (Best et al. 2016). However, two randomized controlled trials were used to assess the utility of LIPUS to treat chronic tendinopathies and reported that LIPUS provided no additional benefit to physical therapy for chronic tendinopathies (D’Vaz et al. 2006; Warden et al. 2008). Thus, while acute injury animal models demonstrate moderate effectiveness of LIUS in tendon healing, human studies specifically investigating chronic tendon injuries do not demonstrate promise as a noninvasive treatment option. Human studies investigating TUS effects on acute tendon injuries would provide further evidence to guide clinical practice, particularly if treatment paradigms across such studies were standardized, as noted above.

Extracorporeal Shockwave Therapy (ESWT) utilizes rapid, short-duration pressure waves (“shockwaves”) intended to elicit physicochemical and cellular reparative responses and has shown promise in treating a variety of musculoskeletal conditions (Simplicio et al. 2020). ESWT can be regarded as high-intensity therapeutic ultrasound, since shockwaves can be generated under both continuous and pulsed modes via transducers that induce nonlinear propagation effects resulting in shock formation. Alternatively, shockwaves can also be generated by non-ultrasonic sources such as electrohydraulic and electromagnetic systems (Simplicio et al. 2020). To date, the US FDA has only approved the use of ESWT for plantar fasciitis and lateral epicondylitis (Wang 2012). Studies have shown that ESWT acts as a mechanical stimulus and promotes healing via mechanotransduction (Moya et al. 2018; Simplicio et al. 2020) and may also alleviate pain (Hausdorf 2008). In humans, ESWT may aid tendon healing by providing mechanical stimulation to aid inflammatory and catabolic processes that are associated with damaged matrix constituents (Waugh et al. 2015). A recent study compared the effectiveness of ESWT and LIUS on pain, return to functionality, and quality of life in patients with Achilles tendinopathy (Dedes et al. 2020). Although both interventions resulted in significant improvements in pain and functionality immediately and 4 weeks post-treatment, the effects of ESWT were more pronounced compared to LIUS. Another recent, randomized controlled trial evaluated the effectiveness of point-focused (small treatment volume focused on the point of maximum pain) and line-focused (larger treatment volume with equally distributed energy density but smaller maximum pressure compared to point-focused) ESWT on patients with confirmed Achilles tendinopathy and concluded that both modalities showed superior outcomes in terms of pain relief compared to placebo treatment (Gatz et al. 2021). Previous in vitro studies have shown the positive effects of ESWT on tenocyte viability and proliferation, collagen fiber synthesis and organization, expression of TGF-β1 and IGF-1, and decreased expression of MMPs and pro-inflammatory interleukins (ILs) (Banes et al. 1999; Chen et al. 2004; Notarnicola and Moretti 2012). While ESWT is currently deemed as a safe and effective “mechanotherapy” to treat many musculoskeletal pathologies including chronic tendon injuries, unfortunately, there exists very little guidance with regard to parameter selection to ensure repeatability and effectiveness for degenerative tendinopathy (d’Agostino et al. 2017; Fan et al. 2020).

10.3 Focused Ultrasound: A Novel Therapeutic for Tendon Healing?

Recently, Focused Ultrasound (FUS) methods have shown great promise for noninvasive tissue ablation, neuromodulation, and drug delivery (Daoudi et al. 2017; Miller and O’Callaghan 2017; Chua and Faigel 2019). FUS treatments are currently approved for numerous applications including treatment of painful bone metastases, essential tremor, uterine fibroids, and prostate cancer (Duc and Keserci 2019). FUS is also gaining considerable interest as a musculoskeletal treatment option, with clinical research in applications such as osteoarthritis, bone and desmoid tumors, epicondylitis, rotator cuff injury, and plantar fasciitis (Liberman et al. 2009; Weeks et al. 2012; Masashi Izumi et al. 2013; Chan et al. 2017). Another emerging application of FUS is pain relief. Although the precise mechanisms of FUS-induced analgesia are not clear, localized denervation of tissue targets and neuromodulatory effects have been presumed (Brown et al. 2015).

The multitude of diverse applications of FUS are largely dependent on the exposure conditions and the manner in which they are delivered (Fishman and Frenkel 2017). In contrast to HIFU (which is primarily used for tissue ablation), therapeutic FUS can be leveraged to achieve effective, functional tissue healing via thermal and/or mechanical stimulation pathways, without inducing irreversible tissue damage. Furthermore, FUS can achieve higher precision with a wide range of acoustic exposures that can encapsulate those utilized in LIUS and ESWT (Fig. 10.4). Additionally, the pulsing parameters can be modulated in real-time during treatment across a wide parameter space. With the advent of state-of-the-art, customizable, reliable, and safe FUS systems, researchers and clinicians are beginning to leverage the potential of image-guided, therapeutic FUS for the treatment of many debilitating conditions.

Fig. 10.4
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Comparison of an approximate range of peak acoustic pressures delivered by FUS, ESWT, and Low-Intensity TUS

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Focusing on the ultrasound beams prevents them from being applied to other regions, and minimizes the potential for thermal or mechanical damage to tissues located outside the focal zone, allowing for precise treatments of targeted tissues or tissue regions. To achieve localized biological effects, FUS transducers are designed such that focused ultrasound beams converge at a single focal point, using techniques such as geometric focusing (concave transducers that cause waves to arrive at a single focal point), electronic focusing (using phased array transducers composed of multiple piezoelectric elements), or by using acoustic lenses (mimicking a concave transducer surface) (Elhelf et al. 2018).

Ultrasound waves interact with tissues to produce thermal and non-thermal bioeffects (Sect. 10.2.1). Acoustic cavitation, which is one of the most widely studied non-thermal mechanisms, is not significant at lower intensities and is often associated with “high” acoustic pressures (Bader et al. 2019). Typically, at high-pressure amplitudes, microscopic gas bubbles form and oscillate (non-inertial cavitation) or steadily grow in size and collapse above certain pressure thresholds (inertial cavitation). Stable or non-inertial cavitation may induce reversible tissue effects such as sonoporation, whereas inertial cavitation induces large stresses and strains on the tissue, ultimately leading to irreversible tissue damage, i.e., histotripsy ablation. Cavitation can also enhance thermal effects by increasing energy absorption at the focal point. Thus, acoustic amplitudes can directly alter the threshold for inertial cavitation as they can change bubble response from non-inertial to inertial cavitation. Mechanical Index (MI) is another parameter that is commonly used to determine the likelihood of cavitation. It is defined as the maximum value of negative peak pressure divided by the root square of the acoustic center frequency. The MI is frequently used to determine the exposures below which cavitation and related bioeffects would not be observed.

In the absence of cavitation, tissue displacement due to FUS application typically occurs as a result of radiation forces. Acoustic Radiation Force (ARF) is defined as the time-averaged force exerted by acoustic waves on the tissue (Urban 2018). As a result of these forces, localized tissue displacement and deformation can be observed, due to transfer of momentum from the sound field to the tissue. Depending on variables such as probe orientation, ultrasound can induce mechanical loading of the extracellular matrix, which in turn provides a biophysical stimulus to the resident cells. ARFs are known to influence cellular proliferation and protein synthesis as evidenced by augmented wound healing and bone remodeling and healing (Curra and Crum 2003; Zhang et al. 2012; Tang et al. 2015). Figure 10.5 depicts the application of radial forces, which deform the tendon transversely while, simultaneously, shear loading of the tendon occurs longitudinally (i.e., along the tendon’s long axis). Irrespective of the mode of mechanical loading, tissue deformation can be spatially and temporally quantified using speckle-tracking methods in conjunction with high-frequency imaging (Bercoff et al. 2004; Liu and Ebbini 2010; Ebbini and ter Haar 2015). Perhaps the most prominent biomedical application utilizing ARFs is in conjunction with imaging, to determine the mechanical properties of tissue by utilizing radiation forces (Wells and Liang 2011; Doherty et al. 2013; Urban 2018). Acoustic Radiation Force Impulse (ARFI) imaging utilizes short-duration acoustic radiation forces to generate localized, quantifiable tissue deformation, thereby providing a noninvasive method to quantify tissue biomechanical properties. Recently, the utility of ARFI imaging has been recognized for multiple tissue types including tendon, thyroid, breast, kidney, and pancreas (Bojunga et al. 2012; Wang 2016; Kaya et al. 2018).

Fig. 10.5
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An FUS transducer producing Acoustic Radiation Force to generate tissue displacements and/or deformation

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Currently, FUS methods are being explored to achieve effective tendon healing. One approach is to utilize “low intensity” FUS methods to promote tendon healing via radiation forces (predominantly mechanical stimulation) without producing thermal damage and cavitation-like bioeffects (Meduri et al. 2020). Higher amplitude pulses (compared to traditional physiotherapy applications) are expected to induce larger tendon matrix strains, and this mechanical effect, similar to existing mechanical loading (exercise) therapies typically used for the treatment of chronic tendon injuries (Mafi et al. 2001; Kingma et al. 2007; Irby et al. 2020), in turn, can effectively stimulate tendon repair. Researchers are also exploring histotripsy, a cavitation-based therapy that utilizes short, high-intensity pulses to mechanically homogenize tissue with negligible heating in the tissue (Smallcomb and Simon 2019). Such cavitation effects can induce targeted microdamage within the tendon and promote a healing response, thus serving as an improved, noninvasive alternative to traditional dry needling approaches (Khandare et al. 2021). Initial studies have shown that this approach preserves the mechanical properties of tendons better than dry needling, without damaging the surrounding tissue.

The wide range of FUS applications utilizing both thermal and non-thermal bioeffects indicate the need for robust, real-time image guidance systems with high sensitivity and specificity, for target visualization, beam focusing, and accuracy verification. An ideal, innovative FUS system for tendon healing applications can precisely target tendons, accurately measure the resulting thermal and mechanical bioeffects and facilitate the investigation of cell/tissue responses. Given the paucity of published data on acoustic parameters and mechanisms of action associated with focused ultrasound therapies for tendon injuries, a thorough investigation of such methods in suitable preclinical models is necessary. Controlled studies in relevant animal models will provide a rigorous basis for optimizing acoustic pulsing parameters for FUS tendon treatments, strengthen the rationale for using FUS as a noninvasive treatment method of stimulating tendon healing and will establish a modular, scalable experimental platform upon which further studies of different species (e.g., rabbit, equine, and human) can readily be undertaken. Figure 10.6 depicts a modular, custom designed system that can apply controlled mechanical, thermal, and mechanical-thermal (dual) stimulation to murine Achilles tendons, under image guidance (Meduri et al. 2020). To establish the feasibility and efficacy of applying different pulsing schemes to injured tendons, investigators may utilize reliable preclinical tendon injury models such as tenotomy for acute injuries or a previously established murine Achilles model (Bell et al. 2013a; Rezvani et al. 2021) of degenerative tendinopathy. The concurrent utilization of real-time, high-frequency ultrasound imaging (as depicted in Fig. 10.6), high field magnetic resonance imaging (MRI) methods, or laser vibrometry enables the quantification of mechanical effects.

Fig. 10.6
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Schematic of a custom-built small animal FUS system allowing interchangeable transducers and driving systems (for different FUS exposures) and a high-frequency imaging system for real-time treatment monitoring and guidance

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Noninvasive (MRI thermometry) or invasive (thermocouple) assessment of dynamic temperature changes accompanying FUS treatment of tendons is a crucial component in analyzing thermal effects of pulsing. Experimental, regional measurements of temperature may further be used to validate computational simulations of predicted heating effects from FUS fields. The effect of predominantly thermal and predominantly mechanical stimulation on the healing profiles of injured tendons can then be established using biomechanical, geometric, cellular, and histologic analyses.

10.4 Future Advancements in Ultrasound-Based Stimulation of Tendon Healing

Although therapeutic ultrasound approaches are widely used for physiotherapy applications, the mechanism of action and optimal acoustic parameters are poorly understood and have not been systematically investigated in comparative studies utilizing in vivo tendon injury models. Novel in vivo data from small animal FUS studies will serve as a foundation upon which the methodology can be readily adapted to larger species in order to explore a wider range of FUS treatments in more clinically relevant animal models. Future studies aimed at identifying optimal TUS modalities (e.g., FUS and ESWT) and the corresponding acoustic parameter sets for the treatment of specific tendon injuries (acute and chronic) will strengthen the rationale for using ultrasound modalities for noninvasive stimulation of tendon healing and regeneration.

Specifically, for the treatment of chronic tendon injuries, it is widely known that mechanical loading-based treatments such as exercise-based rehabilitation can effectively treat symptomatic tendinopathy; however, the mechanism of this healing response is not well understood. Furthermore, physiotherapy requires patient compliance, frequently causes discomfort, and may require lengthy treatment periods (e.g., up to 5 years) for symptomatic relief and restored functionality. Successful development of US-based treatments for chronic tendon injuries may provide further insights into the aforementioned healing pathways in response to mechanical loading. Finally, FUS approaches for treatment of tendon injuries alone or in combination with other therapies could represent an attractive alternative for individuals who are unable or unwilling (e.g., due to pain or injury severity) to pursue physical therapy. In turn, prompt and effective treatment of injured tendons is expected to halt the progression of long-term, degenerative changes that may lead to chronic mobility issues.