Muscle multiorgan crosstalk with MG53 as a myokine for tissue repair and regeneration

Abstract

Through stress and injury to tissues, the cell membrane is damaged and can lead to cell death and a cascade of inflammatory events. Soluble factors that mitigate and repair membrane injury are important to normal homeostasis and are a potential therapeutic intervention for regenerative medicine. A myokine is a type of naturally occurring factors that come from muscle and have impact on remote organs. MG53, a tripartite motif-containing family protein, is such a myokine which has protective effects on lungs, kidneys, liver, heart, eye, and brain. Three mechanisms of action for the beneficial regenerative medicine potential of MG53 have been identified and consist of 1) repair of acute injury to the cellular membrane, 2) anti-inflammatory effects associated with chronic injuries, and 3) rejuvenation of stem cells for tissue regeneration. As such, MG53 has the potential to be a novel and effective regeneration medicine therapeutic.

Introduction

The cell membrane is a dynamic barrier that serves as the interface between the intracellular and extracellular environments. The complex interactions of trafficking intracellular to extracellular contents and vice versa is influenced by autocrine, paracrine, and direct contact stimuli. Injury to the cell membrane has critical implications on cell function, regulation, and overall survival. Inherent in cells is a membrane repair mechanism to counteract any perturbations caused by stress, whether that be mechanical, chemical, ischemia reperfusion (IR), pharmacologic, or oxidative stress. Timely repair of the cell membrane is critical to cell survival and regeneration from injury.

In the field of regenerative medicine, soluble factors secreted from muscles have been found to have important impacts on aging and regeneration, often associated with exercise [1,2]. These soluble factors are now known as myokines. Myokines can have an impact on immunologic function [3], aging, and organ protection. Mitsugumin-53 (MG53) is one such endogenously secreted myokine which has beneficial effects on repair and regeneration of remote vital organs [4–17].

MG53 secretion from skeletal muscle

MG53 is a tripartite motif-containing (TRIM) family protein and plays a key role in repairing cell membrane damage and tissue regeneration. Mechanistically, MG53 facilitates the trafficking of vesicles to the cell membrane to begin the repair process [4,18,19]. First identified as a novel protein in excitation-contraction coupling and muscle physiology [20] regulated by zinc binding to RING and B-box motifs [21], MG53 was quickly identified to be a key cell membrane regeneration factor [4,5,19,22]. Not entirely unique, there have been more than 80 identified TRIM proteins in humans and all share the RING and B-Box motifs [23]. The cellular functions of these TRIM proteins encompass a wide array of critical processes [23–25]. There is a close association among MG53 within species and it is highly conserved. The human MG53 shares homology particularly with Pongo abelii (99.6%), Gorilla (99.4%) and Hylobates moloch (99.2%) [26], implying that this degree of conservation in diverse animals denotes a common ancestry.

While MG53 is primarily found and expressed in striated muscles [4,19,20], it is also found in other cells throughout the body, such as the lung and kidney [7,8] and has circulating effects throughout the body [27,28]. As circulating MG53 could play a role in tissue regeneration, targeted effects of extracellular MG53 could be translated into a treatment of injured tissues in people. To facilitate its regenerative medicine potential, recombinant human MG53 (rhMG53) has been developed as a novel cell membrane repair therapeutic [7–11,13–17,27,29–40].

Several stimulations could contribute to release of MG53 from skeletal muscle. First, noninjurious treadmill running can trigger elevation of circulating MG53 in murine model [29], suggesting the beneficial role of exercise might involve promotion of regenerative factors such as MG53. Second, several studies have shown that insulin treatment could induce release of MG53 from skeletal muscle [27,41], thus potential therapeutics could involve stimulation of the metabolic process of MG53 secretion. In addition, given the elevation of circulating MG53 after IR stress in striated muscles [10,30], noninvasive manipulations of repetitive short-term IR treatment to mimic ischemic preconditioning has emerged as an attractive means to enhance MG53 secretion to preserve remote organ function associated with surgery or chemotherapy [42].

Mechanism of MG53 function in tissue repair and regeneration

As our understanding of MG53 has grown, we have come to identify three general mechanisms of action in tissue repair and regeneration: a) repair of acute injury to cell membrane, b) anti-inflammation, and c) rejuvenation of stem cells.

These three modes are separate yet complementary to each other. On one hand, MG53’s function in repair of membrane injury could have direct implication to facilitate treatment of acute injury to multiple organs [5,7–11,13,29,31]. In addition, several studies from our laboratory and other investigators have revealed an anti-inflammatory function of MG53 associated with chronic tissue injury, sepsis, or viral infection [14–16,33,40,43,44]. Targeting the anti-inflammatory role of MG53 could have broader application in regenerative medicine. Finally, another important physiological role of MG53 has been linked to protection and rejuvenation of stem cells [14,35,40,44,45]. Preservation of stem cell lineage and function associated with aging or degenerative human diseases is another important area of MG53 research.

MG53 protection in vital organs

MG53 as a myokine has been found to have protective and regenerative effects in several vital organ tissues (Figure 1). Here we summarize the various applications of MG53 in multiorgan protection.

Figure 1.

MG53 is abundantly expressed in skeletal muscle and circulates in the blood stream. As a myokine, MG53 has benefits on protection against injury to multiple organs, including heart, lung, kidney, liver, brain, cornea, and skin. In addition to facilitating tissue repair, MG53 has anti-inflammation function to preserve organ function under chronic injury conditions. Moreover, MG53 can preserve stem cells to improve the regenerative capacity of multiple tissues.

a. MG53 and the lung

Numerous stresses can induce acute lung injury (ALI)—trauma, transfusion, ventilator-induced lung injury, and infection being the most notable. MG53 has been identified in type I and II murine, rat, and human alveolar epithelial cells [7,31,46]. Through the use of the mg53−/− mice, researchers have been able to demonstrate that a lack of endogenous MG53 leads to increased susceptibility to lung injury caused by various stresses including IR injury, ventilator-induced lung injury, and lipopolysaccharide [7,31,46]. MG53 over-expression, intravenous delivery, or inhalation of rhMG53 had protective effects in repair of alveolar epithelial cells and prevented lung injury by facilitating cell membrane repair [7,31,46].

Trauma and poly-trauma has direct deleterious effects on the lung through direct contact trauma, contusive blast effect, inhalational injury, and hemorrhage/resuscitative injury. This mechanism of trauma-related injury is relevant to military service personnel as well as civilian populations. Through live cell imaging, cell membrane repair in human bronchial epithelial cells is preserved with MG53 [17]. In addition, in a porcine model of combined chest contusive lung injury and hemorrhagic shock, exogenous rhMG53 reduced percentages of circulating neutrophils and red blood cells in bronchioalveolar lavage fluid of injured animals [17].

Recent and timely critical discoveries in ALI have found that rhMG53 may have a dual therapeutic effect in treating viral associated ALI [16]. With the ongoing global SARS-CoV-2 (COVID-19) pandemic and the annual viral pneumonia cases predominated by influenza virus, the concomitant action of rhMG53 on mitigating the infectious inflammasome and inhibiting pyroptosis and the cell membrane repair properties, has seen a significantly positive beneficial effect in animal models [15,16]. These positive effects in viral associated ALI are not by a direct antiviral effect of MG53. In mice that were exposed to a lethal dose of influenza virus and treated with exogenous rhMG53, a 92% survival rate with preservation of normal body weight was seen, as compared with universal death in the control mice [16]. In these contexts, rhMG53 may be a promising directed therapeutic to suppress an inflammatory cytokine storm and facilitate lung regeneration.

b. MG53 and acute kidney injury

Although expressed at low concentrations in the kidney, MG53 is significantly expressed in areas, particularly the proximal tubular epithelium [8]. This is important since the proximal tubules are very sensitive to injury from nephrotoxic agents, IR, and infection. The MG53 deficient mice develop pathologic renal phenotypes with interstitial cell injury and are at increased risk of IR associated renal injury. This renal IR injury in mice is able to be mitigated through exogenous rhMG53 administration [8].

rhMG53 has also been demonstrated to significantly reduce acute kidney injury (AKI) from cisplatin (without reducing the oncologic efficacy of cisplatin) [8], from radiologic contrast associated AKI [36], and from renal insufficiency associated with a 30% total body surface area burn [47]. From these many animal models of AKI ameliorated by exogenous rhMG53, one can see that MG53 is a critical component to prevent tubular epithelial cell injury from several etiologies.

c. MG53 and the liver

Liver or hepatocyte injury can occur during trauma, drug effect, resection, or IR injury associated with transplantation and surgical manipulation in nontransplant settings. In a hepatic IR model, Yao et al were able to demonstrate that exogenous rhMG53 administration reduced lactate dehydrogenase, ALT, and AST, associated with liver injury [12]. Exactly how muscle-derived MG53 in circulation contributes to hepatic protection under physiological or pathological conditions remains to be studied.

d. MG53 in myocardial protection

Myocardial injury can manifest as acute, chronic, and acute on chronic pathophysiology. Typically, an acute injury is attributed to myocardial ischemia which may sometimes progress to infarction and scar. MG53 has shown a broad ability to be protective in myocardial injury [6,42,48,49,56]. Ischemic preconditioning of myocardium is able to promote MG53 release [42]. This injury/preconditioning effect may be augmented to be protective through exogenous rhMG53. Liu et al. demonstrated that exogenous administration of rhMG53 to injured cardiomyocytes is able to stimulate the prosurvival PI3K pathway in pigs [10]. It appears that there may be a common pathway irrespective of the mode of stress that is affected during myocardial infarction, which can be targeted by MG53. This is further reinforced by work reported by Han et al. who demonstrated myocardial dysfunction protection by rhMG53 in the setting of sepsis [43].

e. MG53 in neuronal injury protection

While MG53 is undetectable in the neurons, overexpression of MG53 can enhance cell membrane repair in cultured neuronal cells [11,29,50]. Yao et al. used a mouse model with sustained elevation of MG53 in circulation, and demonstrated that increased MG53 levels could protect the brain against IR injury [11]. In addition, they found that MG53 could permeate the blood-brain barrier under brain injury condition, and rhMG53 administration promotes prosurvival RISK pathway in response to IR injury to the brain [11].

Guan et al. used a mouse model of contusion-induced traumatic brain injury (TBI) and observed neuroprotective function of rhMG53 [14]. They found that TBI caused transfer of MG53 from circulation to the injured brain, and further elevation of rhMG53 was detected in the brain after intravenous administration to mice subjected to TBI. The neuroprotective function of MG53 is linked to decreased brain edema and lesion volume through activation of PI3K/Akt and ERK survival pathway. Additional studies showed that rhMG53 could protect against lipopolysaccharide-induced neuroinflammation and neurotoxicity in mice, via mitigation of the proinflammatory TLR4/NF-κB signal cascade [33].

f. MG53 protection of the cornea

In addition to circulating in the blood stream, MG53 is also found in tears and aqueous humor in murine, canine, and human [13], suggesting a physiologic role of MG53 in cornea homeostasis and wound healing. Chandler et al. showed that the mg53−/− mice exhibited delayed re-epithelialization, pronounced fibrosis and neovascularization as compared with wild-type mice following alkaline-induced injury to the cornea [13]. Topical application of rhMG53 to the eye was effective to treat alkaline-induced corneal wounds in mice [13,27] and rats [13]. Furthermore, daily application of rhMG53 had additive benefits in corneal healing in db/db mice, a type 2 diabetic model [27], without measurable side effects.

Mechanistically, endogenous MG53 is present in the human corneal epithelial cells that can preserve corneal integrity against injury. MG53 in tears and aqueous humor could enter corneal fibroblasts to inhibit their activation by TGF-β [13], thus contributing to reduced neovascularization. Whether or not MG53 plays a role in anti-inflammation associated with cornea ulceration remains to be investigated. It is also important to determine if MG53 can preserve the regenerative capacity of the eye through modulation of the limbal stem cells.

g. Dermal protection of MG53

Given the similarity between corneal epithelial and dermal epithelial, rhMG53 was also used to treat dermal wounds and found to be effective [9]. GFP-MG53 could nucleate to mechanical injury site and protect human keratinocytes. Similar to corneal wounding, mg53−/− mice are more susceptible to incisional dermal wounding than wild-type mice. Formulated rhMG53 protein in hydrogel promoted healing of the dermal wounds with reduced scar formation [9,51]. More importantly, mice with elevated levels of MG53 in circulation displayed enhanced healing capacity, as revealed by the faster rate of wound closure following punch-hole injury to their ears [40].

h. MG53’s role in preservation of stem cells associated with tissue injury

As a mechanical active tissue, skeletal muscle experiences constant injuries in daily activity. The abundant expression of MG53 in skeletal muscle ensures protection against sarcolemma injury induced by extensive exercise [29], genetic alterations [37,52,53], IR injury [30], and toxin-induced injury [29,40]. We know that MG53 could interact with dysferlin and caveolin-3 for intracellular repair vesicle trafficking [19,52]. Nonmuscle myosin IIA is a motor protein for MG53-containing vesicle trafficking [54], and PTRF serves as an anchor protein to help targeting MG53-containing vesicles to the cell membrane injury site [55].

In addition to tissue repair, MG53 also promotes regeneration of skeletal muscle following injury. Muscle satellite cells derived from the mg53−/− mice proliferate slower than those from wild-type mice, and those derived from mice with increased MG53 expression grow faster than wild-type satellite cells [40]. In vivo eccentric skeletal muscle injury experiments revealed that delayed and repetitive rhMG53 administration had extended benefits to facilitate muscle recovery, demonstrating a role of rhMG53 in skeletal muscle regeneration [40].

Multiple studies also support a role for MG53 in preservation of stem cell function associated with tissue injury. Li et al. reported that cell membrane damage is involved in the impaired survival of bone marrow stem cells by oxidized low-density lipoprotein, and rhMG53 could protect oxidative stress–induced injury to these stem cells [45]. Moreover, n-acetylcysteine, an antioxidant, could prevent oxidized low-density lipoprotein-induced reduction of MG53 and enhance MG53’s protective effect on bone marrow stem cells [35]. Guan et al. found that rhMG53 could enhance survival and therapeutic benefits of human umbilical cord-derived mesenchymal stem cells to treat traumatic brain injury [14,44].

Conclusion and perspectives

MG53 is a highly conserved myokine derived from striated muscle and has the potential to be an effective biologic in regenerative medicine. rhMG53 has robust and beneficial effects on cell membrane repair through well-defined mechanisms of action. These positive reparative effects are seen in many tissues and cell types. Repetitive administration of rhMG53 may have beneficial effects on sustained viability and anti-aging. Evolving impact of MG53 on anti-inflammation and mitigating pyroptosis have encouraging implications for the use of rhMG53 as a broad therapeutic in regenerative medicine [16].

As the benefit of MG53 in tissue repair and regenerative medicine continues to evolve and mature, we formulate these questions for future directions of investigation and intervention: a) How is MG53 released from skeletal muscle? b) Is there an “injury receptor” for MG53 on the plasma membrane that facilitates the extracellular action of MG53 into promotion of cellular survival under stress conditions? c) Does the dual action of MG53 in tissue repair and inflammation control share a similar “injury receptor”, and where do they diverge at the intracellular signaling cascade? d) Can we design therapeutic means to target both anti-inflammation and tissue repair functions of MG53 to elicit long term benefits in degenerative human disease? e) As MG53 contains the conserved E3-ligase activity, how does this function participate in the quality control and stress-response of the cells under physiologic and pathologic conditions? f) Are there genetic roots for MG53-associated repair machinery in degenerative human diseases that can be exploited; and how do developmental changes impact MG53’s role in the aging aspect of regenerative medicine ? Finally, from a surgical point of view, how might MG53 be used as a novel reagent for organ preservation during surgery or transplantation?