Pathological changes and therapeutic strategies for sarcopenia

Abstract

• Sarcopenia is becoming a major public health concern for older adults. The incidence rate in people over 70 years of age is 30–50%. Patients with sarcopenia not only have difficulty moving and are prone to falls and fractures, but in severe cases, they may also experience heart and lung failure and even death. Early diagnosis and prevention in high-risk populations can effectively prevent the deterioration of muscle atrophy.

• In this review, we describe the physiological mechanism of muscle contraction and reveal common pathological changes in sarcopenic patients, including oxidative stress, inflammation, insulin resistance, hypoxia, and disturbance of the gut microbiota. These pathological changes synergistically inhibit the mass and strength of skeletal muscles.

• We also discuss nonpharmacological therapeutic methods for sarcopenia, such as nutrient supplementation and exercise, especially resistance training. On the basis of a thorough analysis of the pathogenesis of sarcopenia in high-risk populations, we believe that tissue synthesis and energy supply are the foundation for maintaining the normal physiological functions of muscles.

• Mitochondria are potential targets for the optimization of intervention methods. Targeted delivery of functional mitochondria to skeletal muscle cells contributes to improving biological oxidation, redox balance, and tissue remodeling. Additionally, stem cell transplantation with the stimulation of growth factors may also be an available method for the further treatment of sarcopenia.

Introduction

Sarcopenia is a progressive systemic skeletal muscle disease characterized by insufficient muscle mass and function and is an age-related developmental process (1). According to statistics, nearly 30% of community residents over the age of 50 are at risk of developing sarcopenia (2). The main clinical manifestations of sarcopenia are weight loss, decreased muscle strength, slower walking speed, decreased grip strength, and reduced mobility. Low muscle mass, strength, and physical performance are the common definitions of sarcopenia (3). In accordance with standards revised by the European Working Group on Sarcopenia in Older People (EWGSOP), handgrip strength is used to determine muscle strength, dual-energy X-ray absorptiometry (DXA) is used to measure muscle quality, and the short physical performance battery test is used to evaluate physical performance (4). The specific standards are as follows: grip strength ≤28 kg for males and ≤18 kg for females; appendicular skeletal muscle mass (ASM) detected by DXA/height² (kg/m²) ≤7.0 for males and ≤5.4 for females; and walking speed <1.0 m/s, five-time sit-to-stand test ≥ 12 s and short physical performance battery ≤ 9 points.

Sarcopenia is commonly considered to be induced by age-associated muscle loss. Other disease-mediated inflammation is also involved in the development of sarcopenia (5). Sedentary behavior and physical inactivity reduce muscle strength (6). Insufficient energy or protein intake caused by anorexia, malabsorption, and the consumption of unhealthy foods decreases muscle synthesis and increases muscle loss (7). The onset of sarcopenia is relatively insidious, but its complications threaten patients’ health and quality of life. Sarcopenia increases the risk of falls and fractures, leading to motor dysfunction and the loss of independent living ability (8). It is related to heart disease, respiratory system disease, cognitive impairment, and osteoporosis (9). With the intensification of aging worldwide, the number of older adults suffering from sarcopenia will increase annually, which will become a potential public health issue. A detailed understanding of the pathogenesis of sarcopenia can help develop effective intervention measures. Early prevention of sarcopenia in high-risk populations will reduce the occurrence of complications. In this review, we aim to analyze the process of skeletal muscle contraction, clarify the pathogenesis of sarcopenia, and propose promising treatment strategies.

Proliferation and differentiation of myocytes

Myocytes are derived from mesenchymal stem cells (MSCs) through multistage differentiation. MSCs with specific pluripotency that develop into skeletal muscle cells are called muscle progenitor cells. Wnts and Sonic hedgehog proteins mediate the biogenesis of the dermomyotome, which is positive for Pax3 and Pax7 (10, 11). The dermatome performs myogenic differentiation via the expression of myogenic factor 5 (Myf5) and class I myosin (MyoD) (12). MyoG is another myogenic regulatory factor that is essential for the regulation of myogenic differentiation and function (13). The differentiation of myoblasts into myocytes is regulated by the Notch, Wnt, and TGF-β signaling pathways (14). Additionally, damaged myocytes are replaced by newly formed muscle fibers originating from the proliferation and differentiation of silent muscle satellite cells (MuSCs) (15). Muscle hyperplasia is the process of fusion between myoblasts originating from activated MuSCs and preexisting myofibers. Damaged myofibers secrete activators and induce the proliferation and differentiation of MuSCs into myoblasts. Newly generated myoblasts promote the rational growth of muscle tissue. The MAPK pathway is involved in the growth, repair, and regeneration of muscle tissue and is activated by growth factors (16).

Excitation and contraction of skeletal muscle

The excitation and contraction of skeletal muscle cells depend on the innervation of the somatic nerves. The nerve impulses emitted by motor neurons are transmitted through neuromuscular junctions to skeletal muscles, causing excitation and contraction. Acetylcholine (ACh) mediates the directional conversion process of ‘electrical signal–chemical signal–electrical signal’ at neuromuscular junctions (17). Neural impulses are transmitted along nerve fibers to the axon terminals, causing depolarization of the prejunctional membrane. Depolarization opens up Ca²⁺ channels on the prejunctional membrane for Ca²⁺ influx. An increase in the Ca²⁺ concentration promotes the movement of synaptic vesicles containing ACh toward prejunctional membranes and their fusion. The ACh released into the junctional cleft binds to the receptors and triggers the opening of Na⁺ channels for Na⁺ influx on postjunctional membranes. The transmembrane movement of Na⁺ triggers the action potential of skeletal muscle cells and causes cell contraction (18) (Fig. 1).

Figure 1.

Excitation and contraction of skeletal muscle cells. The excitation and contraction of skeletal muscle cells depend on the innervation of the somatic nerves. Neuromuscular junctions are the sites of signal transmission. Ca²⁺/ACh/Na⁺ mediated the signal conversion of the ‘electrical signal–chemical signal–electrical signal’ at the neuromuscular junctions. The myotube systems constitute the structural foundation, and Ca²⁺ is the coupling factor for the process of ECC. RyR1 regulates the release of Ca²⁺ in terminal cisternae via conformational changes in L-type Ca²⁺ channels to induce myocyte contraction. Cross-bridges on myosin bind with actin to pull thin filaments into thick filaments, inducing myofilament sliding. Tropomyosin and troponin participate in the regulation of the actin–myosin complex.

Excitation–contraction coupling (ECC) links the excitation caused by action potentials on the muscle membrane and the contraction process of muscle cells caused by myofilament sliding (19). The myotube system is the structural foundation, and Ca²⁺ is the coupling factor for the process of ECC in skeletal muscle (20). Transverse tubules and longitudinal tubules are independent myotube systems. The transverse tubules, as the part of the muscle membrane that extends into the cell, transmit action potentials to the myofibrils surrounding the deep part of the muscle cell. The longitudinal tubules wrap longitudinally around the cytoplasm of myocytes and form terminal cisternae near the transverse tubules for Ca²⁺ storage. The terminal cisternae on both sides of the transverse tubule together form a triad structure, which plays an important role in ECC. It has been confirmed that almost all of the increased Ca²⁺ in the plasma during the ECC process comes from the release of internal calcium. The Ca²⁺ channel on terminal cisternae, also called ryanodine receptor 1 (RyR1), is sensitive to conformational changes in L-type Ca²⁺ channels on transverse tubules. L-type Ca²⁺ channels on transverse tubules are voltage-sensitive molecules that are activated by depolarization when myocytes are excited (21). After activation, they induce the opening of RyR1 on the terminal cisternae through conformational effects, releasing a large amount of Ca²⁺ into the cytoplasm. A rapidly increased Ca²⁺ concentration in the cytoplasm leads to muscle contraction. The calcium pump recovers Ca²⁺ from the cytoplasm into the longitudinal tubules to relax muscle (Fig. 1).

Skeletal muscle cell contraction is the process of myofilament sliding induced by the binding of Ca²⁺ and troponin. Many parallelly arranged myofibrils run through myocytes. Myofibrils present a structure of alternating light and dark along the long axis, forming light bands and dark bands. The light bands are composed of only thin filaments, whereas the dark bands mainly contain thick filaments. Thick filaments are composed of numerous myosin molecules. There is a cross-bridge structure on myosin with ATPase activity, which can bind and breakdown ATP to release energy. The cross-bridges are activated after binding with actin, pulling thin filaments into thick filaments (22). Thin filaments are composed of actin, tropomyosin, and troponin. There are cross-bridge binding sites on actin, which can induce myofilament sliding and muscle contraction. Tropomyosin can cover sites on actin, preventing the binding of cross-bridges to actin and relaxing muscle (23). Troponin is a regulatory protein that binds to tropomyosin. The binding of Ca²⁺ to troponin triggers conformational changes and weakens the binding ability to actin (24). The exposure of binding sites on actin converts more potential energy stored in the cross-bridges into kinetic energy to induce myofilament sliding (Fig. 1).

Pathological changes in the development of sarcopenia

Sarcopenia is a progressive and systemic skeletal muscle disease accompanied by systemic pathological changes. Pathological states induced by aging, such as oxidative stress, inflammation, and hypoxia, have been proven to be associated with the occurrence of sarcopenia (25). Insulin resistance and imbalance of the gut microbiota are also involved in the development of sarcopenia (26, 27).

Oxidative stress

Serum markers of oxidative stress are nonlinearly associated with sarcopenia (28). Redox parameters, such as reduced (GSH) and oxidized (GSSG) blood glutathione and plasma malondialdehyde (MDA) and 4-hydroxy-2,3-nonenal (HNE), are measured in sarcopenic patients. The results revealed that both the GSSG/GSH ratio and the MDA/HNE-protein adducts increased in sarcopenic blood, which indicates that redox imbalance is involved in the pathogenesis of sarcopenia (29). The oxidative balance score, which represents the extent of exposure to oxidative stress-related factors related to diet and lifestyle, is also negatively associated with decreased skeletal muscle mass and handgrip strength (30). Oxidative stress in muscle tissue is caused by the accumulation of reactive oxygen species (ROS). The sources of ROS are mainly mitochondria, NADPH oxidase (NOX), and xanthine oxidase (XO) (31).

Oxidative stress-related mitochondrial dysfunction and mitophagy are involved in the development of skeletal muscle atrophy (31). Mitochondria are potential targets of exercise training and nutritional intervention for the treatment of sarcopenia (32). Mitochondrial DNA (mtDNA) deletion and electron transport chain truncation reduce the energy production of oxidative phosphorylation (OXPHOS) for muscle contraction (33). Increased glucagon-like peptide 1 (GLP-1) levels inhibit the membrane translocation of glucose transporter 4 (GLUT4) to block glucose intake and ATP production in mitochondria for muscle contraction (34). Histone β-hydroxybutyrylation can increase gene transcription associated with mitochondrial pathways to increase OXPHOS and ATP metabolism for energy supplementation (35). The antioxidant enzymes in mitochondria include superoxide oxidoreductase (SOD), catalase (CAT), and glutathione peroxidase (GPx). The stable activity and function of antioxidant enzymes contribute to preventing the cytotoxic effects in myoblasts (36). SOD knockout leads to the loss of innervated neuromuscular junctions and inhibits exercise capacity by increasing ROS accumulation and decreasing oxygen consumption, which is reversed by CAT overexpression (37). SOD is essential for regulating intracellular calcium transients for calcium handling at neuromuscular junctions (38). SOD deficiency decreases the activity of Ca²⁺-ATPase in the sarco/endoplasmic reticulum, which can be reversed by GPx4 overexpression to maintain the homeostasis of ECC (39). GSSG/GSH-mediated redox metabolic processes are involved in the regulation of PGF2α and PGE2 expression to influence muscle growth and regeneration (40). A deficiency in glutathione peroxidase increases the accumulation of lipid hydroperoxides, causing muscle weakness (41). PGC-1α is an important factor in the modulation of antioxidant signaling pathways in mitochondria. PGC-1α deficiency inhibits the expression of Nrf2 and induces the production of ROS to decrease grip force and hanging time in mice (42). AMPK is involved in the regulation of the PGC-1α/Nrf2 axis to restore mitochondrial function (43). The activation of PGC-1α increases mitochondrial biogenesis and increases antioxidant activity to promote the expression of insulin-like growth factor 1 (IGF1), MyoD, myoglobin, and muscle-specific RING finger (MURF) (44). Mitophagy is also associated with the development of sarcopenia by regulating the selective removal of damaged mitochondria (45). PGC-1α deficiency impairs mitophagy to increase mitochondrial defects and enhances H₂O₂-mediated oxidative damage to limit the physiological functions of muscle tissue (46). The activation of mitophagy contributes to maintaining mitochondrial homeostasis and increasing mitochondrial antioxidant capacity to improve myocyte activity (47). Mitophagy also relieves oxidative stress to improve the regenerative ability of satellite cells (48) (Fig. 2).

Figure 2.

Pathological changes in oxidative stress in sarcopenia. Mitochondria-, NOX-, and XO-mediated ROS accumulation is the main cause of oxidative stress damage in the development of sarcopenia. Energy metabolism regulated by the electron transport chain and mtDNA in mitochondria provides ATP to maintain muscle strength. GLP-1/GLUT4 is involved in the transduction of glucose from the cell membrane to the mitochondria. Antioxidant enzymes in mitochondria, such as SOD, CAT, and GPx, influence the expression of genes related to muscle growth. SOD is involved in the modulation of Ca²⁺-ATPase activity in the endoplasmic reticulum to regulate the secretion of Ca²⁺ in ECC. NOX and XO decrease the MMP and impair the antioxidant system in mitochondria. They can also induce the secretion of interleukin factors by activating the NF-κB pathway to aggravate oxidative stress in skeletal muscle cells.

NOX- and XO-mediated ROS production participate in defects in antioxidant defense and peroxidative damage in skeletal muscle (49). Elevated NOX and XO activity decreases the mitochondrial membrane potential (MMP) and induces ROS accumulation to aggravate the apoptosis of myoblasts (50). A clinical study indicated that NOX-mediated H₂O₂ production is the main source of oxidative damage through the inhibition of the antioxidant effect of GPx in the development of sarcopenia (51, 52). NOX-mediated ROS generation shifts the mitochondrial fission–fusion balance toward fission and induces mitochondrial damage in skeletal myoblasts (53). NOX dysregulation increases the activity of antioxidant enzymes to increase IGF1 levels to promote muscle hyperplasia (54). Higher levels of NOX activate the interleukin (IL)-17-mediated NF-κB pathway to induce muscle atrophy (55). Plasma XO is independently associated with sarcopenia and increases significantly in sarcopenic patients (56). Increased XO activity accelerates the aging phenotype in skeletal muscle by opposing the antioxidant enzyme system and increasing the protein levels of inflammatory factors (57). Exposure to XO with xanthine activates the transcription of IL-6 via an NF-κB-dependent pathway to increase ROS production in skeletal myocytes (58). XO is also associated with iron overload in myocytes to attenuate autophagic flux and increase ROS production (59). Targeted inhibition of XO contributes to improving muscle functions in older populations by decreasing H₂O₂ levels, inhibiting lipid peroxidation, and reducing caspase-3 activity (60, 61) (Fig. 2).

Deficiencies in antioxidants, such as vitamin E, are relevant to aging-induced metabolic disturbances in purines, amino acids, and phospholipids, which induce oxidative damage to skeletal muscle (62). Antioxidant exposure plays a protective role in improving muscle strength and preventing muscle atrophy (63). Additionally, both peripheral motor neuron impairment and muscle innervation loss can induce redox imbalance in muscle tissue (64). Protein arginine methyltransferase 1 in motor neurons participates in the repair of the MMP and reverses ROS-induced cell apoptosis to improve neuromuscular dysfunction (65).

Inflammation

Inflammation is an important inducer of the development of sarcopenia (66). The systemic immune–inflammation index is positively correlated with sarcopenia (5). The number ratio of immune cells is an important predictor of the incidence rate and mortality of sarcopenia and its correlation with high-risk diseases (67).

Macrophage dysfunction is a common pathological change in aging-induced skeletal diseases, including osteoarthritis, osteoporosis, and sarcopenia (68). It has been reported that macrophages constitute the majority of immunocytes in the microenvironment of skeletal muscle, and transcriptome sequencing of skeletal muscle tissue indicates that macrophage-rich inflammation is significantly enriched in sarcopenia (69). A single-cell transcriptomic atlas of skeletal muscle across the lifespan also revealed that alterations in macrophages drive changes in collagen synthesis in the extracellular matrix to induce muscle fibrosis and weakness (70). Abnormal activation of macrophages impairs regeneration and promotes fibrosis of muscle fibers by increasing the expression of Trem2 and Spp1 (71). The M1/M2 macrophage ratio is an important factor in immune homeostasis regulation in sarcopenia (72). The transition from M1 to M2 polarization attenuates the inflammatory response and promotes the proliferation and differentiation of MuSCs (73). Macrophage colony-stimulating factor (CSF) is also significantly associated with a high risk of appendicular lean mass (ALM), a sarcopenia-related trait (74). Additionally, macrophages cooperate with MuSCs and fibroadipogenic progenitors involved in the repair of impaired skeletal muscle tissue (75). IL-25 can improve chronic inflammation-induced sarcopenia by promoting the secretion of Sonic hedgehog from M2 macrophages and activating the Sonic hedgehog/Akt/mTOR signaling pathway to increase the proliferation and migration of MuSCs (76) (Fig. 3).

Figure 3.

Inflammation-, insulin resistance-, and hypoxia-mediated development of sarcopenia. Inflammation, insulin resistance, and hypoxia are important pathological changes in sarcopenia. Macrophages and T lymphocytes are the main immunocytes involved in the regulation of muscle tissue. Macrophages induce muscle fibrosis by increasing collagen synthesis. The M1 and M2 polarization of macrophages influences the proliferation, differentiation, and migration of MuSCs. CD4⁺ T-cells play a negative role in myocyte maturation by building an inflammatory microenvironment. CD8⁺ and regulatory T-cells contribute to the synthesis of myotube systems. Insulin promotes muscle contraction by increasing energy supply, promoting protein anabolism, and inhibiting muscle decomposition. Insulin resistance induces oxidative stress and inflammation by increasing fat filtration, causing muscle atrophy. Several types of hypoxia activate the negative signals mediated by HIF-1α and HIF-2α. HIF-1α promotes mitochondrial apoptosis and protein degradation in muscle tissue and induces myofiber transversion by increasing glycolysis as a substitute for aerobic oxidation. HIF-2α activates the ferroptosis process by increasing lipid peroxidation. Mild or moderate hypoxia promotes the metabolism, differentiation, and proliferation of myoblasts for muscle contraction.

T-cell senescence-induced systemic inflammation is associated with handgrip strength (77). Aging-related T-cell phenotypes are potential predictors of decreased muscle health (78). Stable T-cell compartments are able to maintain immune homeostasis and inhibit the expression of inflammatory biomarkers for the physiological activity of muscle (79). Measurement of T-cell gene expression in peripheral blood revealed that seven genes were significantly correlated with the pathogenesis and prognosis of sarcopenia (80). A cross-sectional study indicated that CD4⁺CD28^null T lymphocytes were negatively related to the skeletal muscle mass index (81). Th1/Th17 cells participate in the regulation of the pro-inflammatory microenvironment of skeletal muscle tissue in the development of sarcopenia (82). The number of CD8⁺ T-cells is lower in sarcopenic patients (83). A decrease in the interferon-gamma (IFN-γ) response of macrophages, which are secreted from CD8⁺ T-cells, leads to the dysfunction of MuSC differentiation for myogenesis (84). The combined immunoregulation of macrophages and T-cells is a promising therapeutic strategy for sarcopenia (85). Regulatory T-cells (Tregs) increase the expression of amphiregulin, EGFR, and ST2 to maintain the regeneration and function of muscle (86). IL-6Rα deficiency in T-cells clearly causes Treg deficits and decreases the number and function of MuSCs and fibroadipogenic progenitor cells to inhibit myocyte maturation (Fig. 3).

Moreover, inflammatory factors are important mediators of immune cell-induced inflammation, affecting muscle mass and strength. Higher neutrophil-to-lymphocyte ratios and IL-6, IL-10, and tumor necrosis factor (TNF) levels can be observed in sarcopenic patients (29). Mendelian randomization analysis revealed that circulating IL-16 is associated with ALM and grip strength. IL-1β and CXCL10 are associated with appendicular lean mass. IL-12 and IL-15 are associated with grip strength (87). Another bidirectional Mendelian randomization also revealed that IL-16, a cutaneous T-cell-attracting chemokine, macrophage inflammatory protein 1b, and platelet-derived growth factor BB are significantly involved in aging-induced sarcopenia (88). IL-1β obviously elevates the levels of inflammatory biomarkers, especially the NLRP3 inflammasome, to cause inflammatory myopathies (89). IL-6 increases the expression of MCP-1 to increase inflammation by activating STAT3 in myoblasts, resulting in necrotizing myopathy (90). TNF-α inhibits the expression of Myf5, MyoD, and MyoG to induce muscle atrophy (91). TNF-α and IL-1β activate NF-κB signaling, and IL-6 activates the glycoprotein 130/JAK2/STAT3 pathway to increase SOCS-box protein 1 expression, which leads to regenerative dysfunction in MuSCs and severe muscle atrophy (92). IL-4 is a positive cytokine for myogenesis that increases the expression of MyoD, MyoG, and myomerger to promote myoblast fusion and differentiation after it combines with the IL-4 receptor (93). IL-4 activation reportedly contributes to increasing the effectiveness of stem cell transplantation for the treatment of sarcopenia by increasing the expression of Pax7 (94) (Fig. 3). Tryptophan metabolites that degrade the kynurenine pathway have been confirmed to mediate inflammation in muscle loss (95). Cyclo(histidine–proline) can reduce fibrosis and inflammation in skeletal muscle to inhibit muscle atrophy and restore muscle contraction (96).

Insulin resistance

Insulin sensitivity is important for maintaining glucose metabolism and mitochondrial dynamics to increase muscle mass and function (97). Insulin stimulation is positive for glucose uptake and glycogen storage in muscle tissue (98). Insulin increases the plasma membrane translocation of GLUT4 and glucose intake (99). p21-Activated kinase 4 phosphorylates AMPKα2 at Ser491 to promote insulin resistance via GLUT4 inhibition, which disrupts glucose homeostasis in skeletal muscle (100). Deactivation of the AMPK pathway obviously elevates insulin resistance to limit basic glucose supplementation in myotubes (101). Insulin participates in the regulation of the metabolic switch between OXPHOS and glycolysis (102). Increasing insulin sensitivity contributes to improving mitochondrial respiration to provide energy for myocyte contraction (103). The amino acids required for muscle growth can also be driven into muscles by insulin. Insulin promotes nitrogen retention and acid transport for protein synthesis in muscle (104). Insulin sensitivity is associated with the ability of amino acids to activate the mTOR signaling pathway to increase muscle anabolism (105, 106). Moreover, insulin inhibits the promoting effect of adrenocortical hormones on protein decomposition to reduce the consumption of muscle tissue. Glucocorticoids inhibit insulin resistance to increase muscle catabolism (107). Insulin resistance pathways are involved in the regulation of muscle metabolism (108). Long-term insulin resistance is associated with a high prevalence of decreased muscle mass and strength (109). The triglyceride–glucose index, a biomarker reflecting the degree of insulin resistance, is positively correlated with the development of sarcopenia (110). Ectodysplasin A2 receptor signaling has been proven to mediate insulin resistance-mediated sarcopenia (111). The inhibition of insulin resistance can decrease the expression of MuRF1 and atrogin-1, which are involved in muscle atrophy (112). In addition, myosteatosis is associated with the loss of muscle strength (113). An increase in intermuscular adipose tissue is positively correlated with insulin resistance. Insulin resistance increases fat infiltration in muscle to reduce muscle quality (114). The secretion of inflammatory factors increases, and p38 activity increases after fat infiltration into the muscle to promote protein degradation (115). Lipid deposition without insulin limitation inhibits the β-oxidation of fatty acids and increases the production of mitochondrial ROS, leading to metabolic disturbances in skeletal muscle (116). Muscle tissues also secrete cytokines to induce low-grade inflammation in adipose tissue and local hyperlipidemia, which results in a vicious cycle involving insulin resistance, fat infiltration, chronic inflammation, and muscle atrophy (Fig. 3) (117).

Insulin resistance is a common pathological change in sarcopenia and type 2 diabetes mellitus (T2DM) (118). Sarcopenia is considered a new complication of T2DM (119). Muscle mass is also a potential factor for predicting the risk of T2DM development (120). Type I fiber reduction is a typical characteristic of muscle tissue in T2DM patients (121). T2DM accelerates the decrease in muscle strength and muscle mass induced by disturbances in glucose metabolism and energy supplementation (122). Disturbance of glucose metabolism increases the levels of NOX4 to induce Sirt1 decay, which is mediated by endoplasmic reticulum stress in skeletal muscles (123). Oxidative damage induced by disordered mitochondrial dynamics and the NADPH oxidase system and elevated inflammatory status induced by the secretion of IL-6 and TNF-α are also involved in the occurrence of sarcopenia in T2DM patients (124).

Hypoxia

As age increases, hypoxic conditions accelerate the gradual degradation of the musculoskeletal system (125). Hypoxia exacerbates senescence and increases the molecular phenotype of senescence in skeletal muscle (126). A decreased diameter, low viability, and protein phosphatase 2A activity of myotubes can be observed with hypoxic exposure (127). Hypoxia-related factors are associated with the physiological activity of muscle fibers and the recruitment of MuSCs (128). Long-term hypoxemia inhibits mitochondrial OXPHOS and protein synthesis mediated by hypoxia-inducible factor (HIF)-1α and HIF-2α (129). HIF-1α participates in the regulation of MMP9 and MyoD expression during myogenesis (130). Hypoxic signals mediated by HIF-1α inhibit myogenic differentiation by decreasing PGC-1β and pAMPKα1 expression and promote myoblast apoptosis by inducing mitochondrial apoptosis (131). Activation of HIF-1α induces the transversion of type I muscle fibers with biological oxidation to type II muscle fibers with glycolysis for energy supplementation (132). HIF-1α increases the expression of connective tissue growth factor to induce skeletal muscle fibrosis (133). HIF-1α is also involved in NF-κB-mediated IL-6 secretion to inhibit protein metabolism in muscle (134). HIF-2α is involved in the regulation of myosin heavy chain type I and myosin heavy chain type II, which negatively affect the synthesis of slow-twitch muscle fibers (135). Chronic hypoxia stabilizes HIF-2α to increase local angiotensin-converting enzyme levels, which induces regenerative deficits and inhibits the proliferation of muscle stem cells (136). HIF-2α aggravates intracellular lipid peroxidation and induces ferroptosis in myoblasts (137) (Fig. 3).

However, long-term anemia leads to insufficient muscle tissue support, which is significantly associated with the occurrence of sarcopenia (138). Hypoxia conditioning with mild or moderate hypoxia has been confirmed not only to prevent skeletal muscle dysfunction by increasing PGC-1α and transcription factor A levels to increase mitochondrial biogenesis and metabolic enzyme activity but also to prevent severe hypoxic and ischemic damage (139). Mild hypoxia increases the expression of MyoG to promote muscle differentiation and hypertrophy by activating metabolic pathways (140). The transplantation of stem cells under hypoxic conditions is beneficial for muscle repair (141). The activation of hypoxic signaling increases the expression of vascular endothelial growth factor to induce muscle regeneration (142).

Imbalance of the gut microbiota

Gut microbiota disturbance is an important mediator of senescence-related sarcopenia (143). Lower gut microbiota diversity is observed in older adults with sarcopenia (144). The pathogenesis of the gut microbiota in sarcopenia has been verified by microbiota transplantation from older adults to healthy youth (145). Disruption of the intestinal mucosal barrier accelerates the deterioration of sarcopenia (146). The transplantation of microbiota from young to aged individuals also has therapeutic effects on improving muscle phenotypes and functions (147). The data from multisample Mendelian randomization indicate that the relative abundance of more than 20 gut microbiota is related to walking pace, appendicular lean mass, and grip strength (148). Among them, Blautia, Lachnospiraceae_unclassified, and Subdoligranulum are potential diagnostic markers of sarcopenia (149). Sex-specific differences indicate that Haemophilus parainfluenzae and Roseburia faecis are more abundant in males with a high skeletal muscle mass index, but Bifidobacterium longum is mostly correlated with female sarcopenia (150, 151). Increasing the abundance of Clostridium sensu stricto 1 is helpful for activating the anabolic processes of muscle by regulating the cannabinoid receptor system (152). An improvement in the Allobaculum genus contributes to relieving muscle loss by increasing the production of branched-chain amino acids to increase the transcription of myosin heavy chain family genes (153). Enhancing Ruminococcaceae_UCG_013, Lactobacillus murinus, Algibacter, Bacillus, Gordonibacter, Porphyromonas, and Prevotella_6 improves the activity of antioxidant enzymes and the levels of short-chain fatty acids to increase the appendicular skeletal muscle mass index (154). The genera Subdoligranulum, Alistipes, and Faecalibacterium prausnitzii, associated with the production of short-chain fatty acids, also decrease the expression of atrogin-1 and MuRF1 to inhibit skeletal muscle degradation by promoting FoxO3a-, Akt-, and mTOR-mediated anabolic phosphorylation (155). Prevotella copri promotes the production of branched-chain amino acids, which is correlated with the prevention of sarcopenia (156). By regulating bile acid composition, Bacteroides fragilis, Blautia marseille, Sutterella spp., and Veillonella parvula are positively associated with the development of sarcopenia (157). Parabacteroides distasonis and Duncaniella dubosii increase the levels of circulating aminoadipic acid degraded from lysine to disrupt mitochondrial function by inhibiting mitophagy (158). A decreased composition of Parabacteroides, Akkermansia, and Enterobacteriaceae and their metabolites prevents the aggravation of sarcopenia by influencing nucleotide metabolism, β-alanine metabolism, histidine metabolism, ABC transporters, and the calcium signaling pathway (159) (Table 1). Additionally, more than 170 metabolites derived from the gut microbiota clearly vary in sarcopenic patients (149). Myogenic biomarkers are significantly associated with the gut microbiota for butyrate production (160). The composition of bile acid and its metabolites are also important in the regulation of the gut–muscle axis (157). Phenolic metabolites from the gut microbiome are essential for muscle mitochondria to perform the normal process of energy metabolism (161). Tryptophan metabolites, including kynurenine, 5-hydroxytryptamine, and indole, promote inflammatory responses in muscle tissue (162).

Table 1.

Gut microbiota in the regulation of muscle pathophysiology.

Microbiota	Effects on muscle tissue	Mechanism	Reference
Clostridium sensu stricto 1	Activates the anabolic processes of muscle	Regulating cannabinoid receptor system	(152)
Allobaculum genus	Increases the transcription of myosin heavy chain family genes	Increasing the production of branched-chain amino acids	(153)
Ruminococcaceae_UCG_013, Lactobacillus murinus, Algibacter, Bacillus, Gordonibacter, Porphyromonas, and Prevotella_6	Elevate appendicular skeletal muscle mass index	Improving the activity of antioxidant enzymes and the levels of short-chain fatty acid	(154)
Subdoligranulum, Alistipes, and Faecalibacterium prausnitzii	Decrease the expression of atrogin-1 and MuRF1 to inhibit skeletal muscle degradation	Promoting FoxO3a-, Akt-, and mTOR-mediated anabolic phosphorylation	(155)
Prevotella copri	Prevents sarcopenia	Promoting the production of branched chain amino acid	(156)
Bacteroides fragilis, Blautia marseille, Sutterella spp., and Veillonella parvula	Induce the development of sarcopenia	Regulating bile acid composition	(157)
Parabacteroides distasonis and Duncaniella dubosii	Disrupt mitochondrial function of myoblasts by inhibiting mitophagy	Increasing the levels of circulating aminoadipic acid degraded from lysine	(158)
Parabacteroides, Akkermansia, and Enterobacteriaceae	Aggravate sarcopenia	Influencing the nucleotide metabolism, β-alanine, histidine metabolism, ABC transporters, and the calcium signaling pathway	(159)

High-risk populations for sarcopenia

As demonstrated above, the contraction of skeletal muscles depends on the regulation of the autonomic nervous system, the transmission of neuromuscular junctions, and the mass and strength of myocytes. Motor neuron damage and neuromuscular junction degeneration induce muscle atrophy in sarcopenia (163, 164). In addition, long-term insufficient protein intake or malnutrition significantly reduces muscle strength and quality (165). Populations with these high-risk factors are more susceptible to sarcopenia and require early prevention.

Patients with chronic wasting diseases

People who are in a long-term state of malnutrition are at the highest risk of sarcopenia. Chronic wasting diseases, such as respiratory diseases and heart and renal failure, are the causes of inadequate nutritional supplies for patients.

Respiratory diseases, such as chronic obstructive pulmonary disease (COPD), asthma, and pneumonia, increase the risk of nutrition-related diseases in patients (166, 167, 168). Sarcopenia is regarded as a complication of COPD and is influenced by age, body mass index (BMI), and smoking (169). Nocturnal hypoxemia in COPD patients decreases the pectoralis muscle index, and low oxygen saturation increases the incidence rate of sarcopenia (170). A cohort study revealed that a muscle loss phenotype was common in COPD patients and highly correlated with COPD-related mortality. The results of immunofluorescence microscopy and RNA sequencing of skeletal muscles also revealed that genes differentially expressed in both type I and type IIa myofibers were enriched in abnormal myofibers, which suggested altered transcriptional regulation of muscle tissue in COPD (171). ROS accumulation is a main driver of muscle atrophy (172). Serum lipoprotein-associated phospholipase A2 is negatively associated with the mass and function of skeletal muscles and is a potential biomarker for predicting the risk of sarcopenia in COPD patients (173). Vitamin D deficiency and inflammation are the pathogenic factors of sarcopenia in COPD patients (174). Vitamin D supplementation contributes to improving vascular functions in muscle and increasing muscle strength (175). Greater systemic immune–inflammation and sarcopenia are important factors for evaluating the severity of COPD (176). The dietary inflammatory index (DII) is positively associated with sarcopenia incidence, and appendicular skeletal muscle mass has a nonlinear association with all-cause mortality in COPD patients (177). Asthma is another respiratory disease that clearly increases the risk of sarcopenia (178). Nearly 20% of asthma patients are likely to suffer from sarcopenia (179). Systemic inflammation is the mediator of asthma-induced muscle loss. The DII is also a potential factor for predicting the incidence and severity of sarcopenia in asthma patients (180). Low skeletal muscle mass is related to obesity and increases airway obstruction in asthma (181). Sarcopenia participates in the development of asthma by influencing lung function and comorbidities and is an available therapeutic target for asthma (182). Additionally, the occurrence and outcome of pneumonia are associated with the loss of skeletal muscle (183). During the epidemic of COVID-19, the pectoralis and erector spinae muscles were used to assess the clinical course (184). Acute skeletal muscle loss is an important clinical symptom in COVID-19 patients with poor outcomes (185). Muscle wasting is also a common sequela of COVID-19 (186) (Table 2).

Table 2.

Development of sarcopenia in patients with chronic wasting diseases.

Chronic wasting diseases/effects on muscle tissue	Mechanism	Reference
COPD
Decreases pectoralis muscle index and increases the incidence rate of sarcopenia	Nocturnal hypoxemia	(170)
Induces abnormal myofiber proportion	Altering transcriptional regulation of type I and type IIa myofibers	(171)
Derives muscle atrophy	Increasing ROS accumulation	(172)
Decreases the mass and function of skeletal muscles	Upregulating serum lipoprotein-associated phospholipase A2	(173)
Inhibits vascular functions in muscle	Causing vitamin D deficiency	(174, 175)
Decreases appendicular skeletal muscle mass	Inducing higher systemic immune-inflammation	(174, 176)
Asthma
Increases the incidence and severity of sarcopenia	Systemic inflammation	(180)
COVID-19
Induces acute skeletal muscle loss and muscle wasting	-	(185, 186)
Induces oxidative damage of myocytes and causes muscle atrophy	Increasing circulating TNF-α to destroy mitochondrial function	(193)
Heart failure
Decreases muscle quality	Increasing interleukin factors to induce hemodynamic abnormalities	(194)
Leads to muscle weakness and weight loss	Cachexia	(195)
Destroys muscle composition and function	Cachexia	(196)
Hypertension
Leads to the poor prognosis, increases mortality of sarcopenia and lowers sarcopenic index	-	(200)
Elevates the threshold of the nutrient sensing mTOR pathway in muscle tissue	Inhibiting the production of anabolic hormones and promote the secretion of growth factors	(205)
Accelerates muscle atrophy and reduces muscle strength	Activating protein degradation	(206)
Decreases the skeletal muscle mass	Low glomerular filtration rate	(207)
Inhibits muscle mass and physical function	Metabolic acidosis	(208)
Chronic kidney disease
Induces sarcopenia development	Endothelial dysfunction-mediated peripheral vascular lesion	(209)
Leads to uremic myopathy	Inducing the imbalance between the protein synthesis and catabolism	(210)
Blocks the process of myocyte fusion and induces frequent musculoskeletal injuries	Uremia	(211)
Induces the weakness of signal transduction and mitochondrial dysfunction in myocytes	Insulin resistance	(212)
Influences sarcopenia development	Regulating uremic toxins and inflammatory cytokines mediated by gut microbiota	(213)

Sarcopenia is a common comorbidity in patients with heart failure (HF) (187). Nearly 20–50% of HF patients are at risk of developing sarcopenia (188). The rate of skeletal muscle wasting is as high as 50% with HF (189). The presence of sarcopenia indicates a worse prognosis for heart failure patients (190). The amount of water extracted from the intracellular water of skeletal muscle is a potential index for evaluating muscle quality in patients with heart failure (191). The ratio of appendicular lean mass to body mass index (BMI) can significantly predict muscle function (192). Systemic inflammation and low caloric intake might be the reasons for muscle wasting in HF. The level of TNF-α, which is increased in HF patients, is the main inflammatory factor that destroys mitochondrial function to induce oxidative damage to myocytes and cause muscle atrophy (193). Interleukin factors also induce hemodynamic abnormalities to decrease muscle quality (194). Cachexia occurs in one-third of older patients with heart failure and leads to muscle weakness and weight loss (195). Cachexia increases the possibility of anorexia to reduce the substance basis and energy supply for muscle composition and function in these patients (196). Microbiota regulation is a potential target for the prevention and treatment of sarcopenia in HF patients. The sarcopenic index (SI) obtained from the ratio of serum creatinine to cystatin C is associated with the taxa and composition of the digestive tract microbiota (197). The levels of metabolites, including isobutyric acid, isovaleric acid, and valeric acid, are lower (198). Sarcopenia is associated with the risk and survival of individuals with other cardiovascular diseases (199). A low SI leads to a poor prognosis and increases mortality in hypertensive patients (200). The incidence of sarcopenia is 43% in coronary artery disease patients and 30% in cardiac arrhythmia patients (201). The psoas muscle thickness/height ratio is also an applicable tool for predicting the mortality of acute type A aortic dissection (202) (Table 2).

Almost half of patients with chronic kidney disease (CKD) exhibit significant muscle atrophy (203, 204). CKD can inhibit the production of anabolic hormones and promote the secretion of growth factors, which elevates the threshold of the nutrient-sensing mTOR pathway in muscle tissue (205). The activation of protein degradation also accelerates muscle atrophy and reduces muscle strength (206). A cross-sectional study indicated that a low glomerular filtration rate clearly decreased skeletal muscle mass (207). Metabolic acidosis is a negative factor for muscle mass and physical function (208). Additionally, endothelial dysfunction-mediated peripheral vascular lesions play an important role in the development of sarcopenia in patients with CKD (209). An imbalance between protein synthesis and catabolism in skeletal muscle could lead to uremic myopathy (210). The occurrence of uremia blocks the process of myocyte fusion and induces frequent musculoskeletal injuries (211). Insulin resistance induces a decrease in signal transduction and mitochondrial dysfunction in myocytes (212). The gut microbiota is also involved in the regulation of uremic toxins and inflammatory cytokines, influencing the development of sarcopenia (213) (Table 2). Hemodialysis is the ultimate treatment for renal failure and plays a negative role in muscle strength and bone health (214). Chronic inflammation and insulin resistance are common pathological changes in patients undergoing hemodialysis (215). A lower serum 25(OH)D level is a risk factor for decreased skeletal muscle function (216). Elevated blood manganese is also associated with the occurrence of sarcopenia in patients on maintenance hemodialysis (217). Sarcopenia is also a potential factor for predicting mortality in kidney transplant recipients (218). Low muscle mass and strength decrease the survival rate of these patients.

Sarcopenic obesity (SO)

Both overweight in young people and malnutrition in older adults are considered high-risk factors for sarcopenia (219). The prevalence rate of sarcopenia in obese individuals is greater than 10% (220). A long-term high-fat diet (HFD) can induce muscle atrophy (221). Serum adipocyte fatty acid-binding protein is positively associated with the incidence of sarcopenia (222). Myogenic inhibition was observed with decreased expression of Myf5, MyoD, and MyoG in a high-fat state (223). There is mutual regulation between myogenesis and adipogenesis. MyoD-positive myoblasts secrete antiadipogenic factors to inhibit the formation of white adipocytes (224). Fat infiltration is a pathogenic factor that induces dysfunctional myogenic differentiation (225). Excessive accumulation of white adipose tissue is a high-risk factor for sarcopenia (226). Peroxisome proliferator-activated receptor (PPAR) and CCAAT enhancer-binding protein (C/EBP) are potential transcription factors for inducing the transdifferentiation of myoblasts into adipocytes. S100B is considered a transducer that mediates the effects of ROS to promote the myoblast–brown adipocyte transition (227). Insulin resistance combined with abnormalities in glucose metabolism is an important pathogenesis of sarcopenia induced by obesity (228). Insulin resistance negatively affects aerobic oxidation in mitochondria to inhibit muscle metabolism via the Akt/NOR-1/mTORC1 pathway (229). Obesity also aggravates mitochondrial dysfunction and the energy burden to induce oxidative damage, which inhibits the proliferation of satellite cells and the differentiation of myocytes (230). Mitochondrial dysfunction causes the hypertrophic conversion of adipocytes (231). DNAJA3 is a mitochondrial cochaperone protein involved in the regulation of mitochondrial respiration and fatty acid metabolism. DNAJA3 deficiency increases body fat mass and inhibits mitochondrial respiratory complex activity to aggravate fat accumulation and muscle loss (232). Chemokines can promote the migration of subcutaneous adipocytes to skeletal muscle and increase the content of intramuscular fat (233). Retinoic acid receptor-related orphan receptor-α (RORα) is involved in regulating the number and antioxidant activity of mitochondria to inhibit fat infiltration in muscle by suppressing oxidative MyHC2a fibers (234). SO refers to the presence of overweight and low muscle mass (235). A lower physical activity level and sedentary behavior are causes of SO (236). Inflammation and oxidative stress are the main pathological changes in SO (237). The systemic immune–inflammation index is positively correlated with the risk of SO occurrence (238). Endoplasmic reticulum stress decreases insulin sensitivity, aggravates inflammation in adipocyte tissue, and induces muscle contractile dysfunction and atrophy (239). The gut dysbiosis-mediated occurrence of inflammation is also a main trigger for SO (240). A high-fat diet induces systemic inflammation and insulin resistance and increases the production of trimethylamine N-oxide to activate the ROS-AKT/mTOR signaling pathway to reduce muscle quality and strength by increasing deleterious bacteria in the gut (241). The GSSG/GSH ratio and MDA levels in the blood are strongly associated with the development of SO (242). ROS are the effectors of insulin resistance induced by obesity, causing muscle atrophy (243). ROS disturb mitochondrial functions and arrest the cell cycle in myoblasts (244) (Fig. 4). Physical activity together with nutritional modulation is regarded as the most effective method to manage SO (245).

Figure 4.

Sarcopenia in obesity and osteoporosis. Both obese individuals and osteoporotic patients are at high risk of sarcopenia. There is mutual regulation between myogenesis and adipogenesis. MyoD-positive myoblasts secrete antiadipogenic factors to inhibit the formation of white adipocytes. PPAR and C/EBP induce the transdifferentiation of myoblasts into adipocytes. S100B activates ROS for the myoblast–brown adipocyte transition. Inflammation and oxidative stress are the main pathological changes in SO. Insulin resistance decreases the efficiency of the TCA cycle. DNAJA3 deficiency inhibits the activity of respiratory complexes. RORα deactivation reduces antioxidant activity. All of the above mitochondrial dysfunctions aggravate fat infiltration in muscle tissues. A HFD also induces endoplasmic reticulum stress to cause muscle atrophy in obese individuals through insulin resistance and inflammation. Wnt signaling and growth factors mediate the crosstalk between bone and muscle tissues. Muscles secrete irisin to promote osteogenesis and inhibit osteoclastogenesis. Osteocalcin combines with GPRC6A to promote myoblast proliferation, but sclerostin inhibits myoblast differentiation by deactivating Wnt3a, both of which are produced in bone. Redox imbalance and the immune response are common pathological changes associated with osteoporosis and sarcopenia. The accumulation of ROS caused by mitochondrial dysfunction is negatively related to muscle mass and bone mass. Macrophages and T lymphocytes highly express Spp1, resulting in muscle atrophy and bone resorption. Adipogenesis induced by oxidative stress and inflammation plays an important role in the development of sarcopenia and osteoporosis.

Osteoporosis

Sarcopenia and osteoporosis are highly correlated skeletal diseases and are collectively referred to as ‘dyskinetic syndrome’. Osteoporosis and sarcopenia usually coexist in individuals with long-term physical inactivity (246). Approximately one-fifth of patients with sarcopenia also suffer from osteoporosis (247). Moreover, a follow-up study of subjects over 60 years of age indicated that osteoporosis increased the incidence of sarcopenia (248). Impaired muscle domains are usually observed in patients with osteoporosis (249). High-risk groups of osteoporosis patients, such as postmenopausal women and diabetic patients, also have a potential risk of sarcopenia. Total body skeletal muscle mass is considered an important parameter for assessing the risk of postmenopausal osteoporosis (250). Several types of plasma microRNAs are also correlated with muscle wasting in postmenopausal women (251). Signal transducer and activator of transcription 3 (STAT3) is a potential common hub gene that mediates osteoclast differentiation and myoblast proliferation in postmenopausal osteoporosis and sarcopenia patients (252). STAT3 activation prioritizes the adipogenic differentiation of MSCs over osteogenesis and myogenesis (253). Musculoskeletal damage has an early onset with central fat distribution in patients with diabetes (254). Hyperglycemia-induced pyroptosis exacerbates adverse remodeling of muscle tissue after mass loss (255). The secretion of irisin in muscle also enhances the pyroptosis response in bone tissue induced by diabetes (256).

Bone–muscle interactions are based on the neuronal reflex, which is mediated by an osteocytic mechanoreceptive network that innervates muscle excitement and contraction (257). Wolf’s law reveals that the growth, absorption, and reconstruction of bone are related to the stress state of the bone (258). The contraction and relaxation of muscles are important factors in regulating bone formation and absorption (259). The Wnt/β-catenin signaling pathway is involved in muscle–bone crosstalk (260). Muscle-derived transcription factors and myokines activate Wnt3a, Wnt4b, and Wnt10 to increase the expression of β-catenin for osteogenic differentiation. Wnt3a activated by fluid flow stress in bone promotes the expression of Pax7, MyoD, and Myf5 for myogenic differentiation. Appropriate exercise and stress can promote faster bone growth and healing. Paracrine and endocrine pathways also mediate the interrelated effects between bone and muscle tissue (261). Growth factors participate in the regulation of bone–muscle crosstalk (262). Skeletal muscle secretes fibroblast growth factor (FGF) 2 to promote osteogenesis by activating the BMP2 and Wnt/β-catenin signaling pathways and IGF1 to modulate bone homeostasis by activating BMP9/Smad-induced bone formation and RANKL-mediated bone resorption (263, 264). Osteoblasts secrete IGF1 to promote myogenesis by activating the PI3K/AKT pathway but secrete FGF23 to inhibit the transduction of insulin/IGF1 signals (265, 266). Irisin is an important hormone that is secreted from muscles and is involved in bone metabolism (262). Irisin increases insulin sensitivity to facilitate the conversion of white adipose tissue to brown adipose tissue, increases the expression of osteopontin for osteogenic differentiation via the BMP2/Smad and Wnt/β-catenin signaling pathways, and directly inhibits osteoclast differentiation. The regulatory proteins secreted by bone tissue have a bidirectional effect on muscles. Osteocalcin combines with GPRC6A receptors on myocytes to promote myocyte proliferation through the PI3K/Akt and MAPK/Erk1/Erk2 signaling pathways (267). Sclerostin binds to low-density lipoprotein receptor‐related proteins to inhibit Wnt3a-mediated myogenic differentiation by decreasing the expression of MyoD and MyoG (268) (Fig. 4).

Combined analysis of the proteome and transcriptome revealed differentially expressed genes (DEGs) in bone and muscle tissue in patients with osteosarcopenia. These DRGs are enriched in the regulation of redox balance and the immune response. Oxidative stress is a common pathological change in osteoporosis and sarcopenia (269). The impairment of connexin 43 hemichannels in osteocytes destroys mitochondrial homeostasis and causes ROS accumulation in muscles, which reduces muscle mass and strength together with increased collagen synthesis through the activation of TGFβ/Smad2/Smad3 signaling (270). Inflammation increases RANKL-mediated bone resorption by osteoclasts, which is a high-risk factor for sarcopenia (271). Single-cell RNA sequencing of bone and muscle tissue revealed that a common subset of lipid-associated macrophages exists and that Spp1 is highly expressed to induce muscle atrophy and bone loss (272). Th1 differentiation in muscle and Th17 transformation in the bones of CD4⁺ T-cells stimulate the expression of Spp1. OXPHOS defects in muscle mitochondria influence T-cell homing to the bone marrow via the CXCL12–CXCR4 signaling axis to induce a local inflammatory response and promote adipogenesis, which is negative for bone formation (273). The measurement of biomarkers of microarchitecture in bone and muscle tissue also revealed that basophil and TNFα levels are elevated in patients with osteosarcopenia (274). Moreover, NF-κB signaling pathway-mediated osteoclast differentiation is a potential pathogenic molecular mechanism of osteosarcopenia (275). Fast type II myosin heavy chain isoform and myofiber metabolic shifts lead to unbalanced bone resorption and abnormal protein breakdown in the loss of bone and muscle mass (276) (Fig. 4).

Treatment

The popularization of knowledge and the optimization of treatment strategies for sarcopenia can help improve patients’ willingness and cooperation in treatment (277). Previous studies have revealed that low intakes of energy, protein, vitamin D, and ω-3 fatty acids are associated with sarcopenia risk, and moderate exercise, particularly resistance exercise for muscle mass and strength and aerobic exercise for physical performance, contributes to improving sarcopenia risk (278). Thoroughly analyzing the specific regulatory mechanisms of different intervention measures on muscle tissue contributes to developing specific treatment plans and improving treatment efficiency for sarcopenic patients.

Exercise

Exercise significantly affects the maintenance of muscle function and delays aging. Exercise activates MuSCs through the interaction of transcription factors and Pax7 to increase the expression of Myf5 (279). The secretion of specific exercise-induced cytokines, such as myostatin, irisin, and IL-6, promotes the myogenic differentiation of MuSCs (280). Exercise also increases the stiffness of the extracellular matrix in muscle tissue, which is a positive signal that activates kinesin-1-mediated myogenic differentiation by increasing glucose intake and utilization of GLUT4 (281). Targeted exercise contributes to repairing injured myofibers and maintaining their normal morphology (282).

Exercise can improve oxidative stress and inflammation while protecting telomeric DNA and activating repair pathways to treat sarcopenia (283). Redox signaling plays an important role in muscle remodeling induced by exercise (284). A clinical study indicated that 6 months of physical activity effectively improved serum oxidative markers and muscle mass in sarcopenic patients (285). Mitochondria are important organelles that mediate redox balance in muscle tissues via exercise. Exercise modulates the mitochondrial unfolded protein response and mitophagy through ROS and AMPK signals for mitochondrial quality control (286). Exercise increases mitochondrial biogenesis and improves mitochondrial functions, including maintaining Ca²⁺ homeostasis, enhancing antioxidant activity, and regulating mitophagy (287). The expression of sirtuin family proteins also increases after physical exercise to eliminate excessive ROS and maintain mitochondrial homeostasis for myogenic differentiation of MuSCs (288). Aerobic exercise has a positive effect on mitochondrial functions to promote protein synthesis in muscle (289). Inflammation is another target of exercise in the improvement of sarcopenia (290). A randomized controlled trial indicated that exercise could decrease the serum levels of TNF-α, IL-1β, and IL-6 in sarcopenic patients (291). Regular physical training helps preserve muscle mass and strength by inhibiting muscle inflammation (292). Exercise inhibits the secretion of PGE2-degrading enzymes from macrophages to prevent muscle atrophy (293). T-cell-specific inflammatory gene expression could also be changed after combined exercise for muscle strength performance (80) (Fig. 5). Additionally, exercise has a positive influence on the composition and distribution of the gut microbiota to improve oxidative stress and inflammation, which contributes to preventing muscle atrophy and improving muscle strength (294). Changes in intestinal metabolite abundance and production caused by exercise are involved in regulating the mass and quality of skeletal muscle (295). Different levels and types of exercise are beneficial for muscle stability. A randomized crossover trial revealed that high-intensity interval training is a potential method to prevent and treat sarcopenia (296). Moderate-intensity exercise activates PGC-1ɑ to increase mitochondrial biogenesis and regulate the intracellular Ca²⁺ distribution and ATP/ADP ratio for muscle contraction (297). Blood flow restriction aerobic exercise contributes to improving the distribution and content of fat and muscle by regulating lipid profiles and glucose metabolism (298). Voluntary wheel running improves the activity of citrate synthase, β-hydroxyacyl-CoA-dehydrogenase, and SOD to increase ATP production and antioxidant capacity in mitochondria, which is positive for relieving muscle weakness (299).

Figure 5.

Therapeutic mechanism of exercise and resistance training. Physical activities not only directly stimulate myogenesis but also improve pathological changes in sarcopenia. Myogenic factors are secreted to promote the differentiation of Pax7+ MuSCs into skeletal muscle cells. GLUT4 mediates glucose intake and utilization to activate kinesin-1 for myogenic differentiation. Pax7 and FoxO3a maintain the balance of muscle synthesis and decomposition. Resistance exercise modulates the redox balance and inflammatory response in skeletal muscle. Mitochondria are the regulatory targets involved in antioxidant effects. Mitochondrial respiration, Ca²⁺ transport, and mitophagy are involved in the oxidation–reduction process. Macrophages and T lymphocytes play important roles in maintaining muscle health in an inflammatory state. Macrophages secrete MMP14 to promote ECM and myotube formation. T lymphocytes inhibit fibroadipogenic infiltration to protect muscle growth.

Resistance training (RT)

RT is widely recognized as an effective method for improving muscle mass, increasing muscle strength, and promoting muscle metabolism (300). RT increases the ratio of muscle tissue to skeletal muscle lipid content with increasing muscle density and constant lipid content (301). Progressive resistance exercise training contributes to improving activities of daily living in aged individuals (302). It has been reported that RT can increase the muscle fiber cross-sectional area and the number of Pax7+ MuSCs (303). RT also restored the levels of MyoD, MyoG, and IGF1 (304). Cannabinoid receptors are involved in the positive effect of RT on muscle homeostasis by regulating FoxO3a-mediated catabolism and Pax7-mediated regeneration (305) (Fig. 5).

RT can relieve ROS-mediated oxidative damage to improve the voluntary contraction of skeletal muscle (306). Resistance exercise mainly regulates ROS originating from mitochondria and NOX (307). Mitochondrial respiratory inhibition induced by relatively decreased ADP sensitivity leads to redox stress (308). RT inhibits the emission of H₂O₂ from mitochondria in skeletal muscle (309). The activity of antioxidant enzymes, including GSH-Px, SOD, and CAT, in mitochondria increases with RT by inhibiting the transcription of Keap1 (310). The antioxidant signal of Nrf2 is activated after RT to increase muscle-specific histone methyltransferase (Smyd1) for sarcomere assembly and myofiber folding by inhibiting oxidative stress and endoplasmic reticulum stress (311, 312). RT also relieves ROS accumulation-induced insulin resistance (313). Carbon monoxide-loaded red blood cells can increase skeletal muscle mass and strength and restore athletic ability by activating the PGC-1α and AKT signaling pathways (314). Low-dose inhaled carbon monoxide combined with exercise induces ADP-stimulated respiration to prevent mitochondrial dysfunction and overnutrition-mediated insulin resistance in skeletal muscle (315). RT decreases NOX4 activity to reduce ROS production and enhance AKT signal transduction to alleviate insulin resistance (316). However, RT increases the expression of NOX2 to mediate the signal transduction of insulin, which normalizes body fat and enhances OXPHOS in skeletal muscle (317). Insulin sensitivity is involved in extracellular matrix remodeling in skeletal muscle after RT (318). The orphan nuclear receptors Nur77 and NOR1 mediate the positive effect of RT on the insulin response in skeletal muscle (319). RT increases the expression of major urinary protein 1 (MUP1) to induce GLUT4 translocation and increase insulin sensitivity for skeletal muscle metabolism (320) (Fig. 5).

RT is an effective method for the inhibition of inflammation in sarcopenic patients (321). RT contributes to decreasing the levels of inflammatory biomarkers, especially IL-6, to prevent the progression of sarcopenia (321). RT has been revealed to increase the MuSC number and pool of macrophages in skeletal muscle (322). RT promotes the recruitment of macrophages to facilitate substance phagocytosis, myogenic differentiation, and myotube formation (323). Leukemia inhibitory factor is upregulated with RT to stimulate MMP14 secretion from macrophages, which is involved in the formation of the extracellular matrix in skeletal muscle (324). The accumulation of muscle macrophages also elevates insulin sensitivity (325). RT activates chitinase-3-like protein 1 (CHI3L1)/protease-activated receptor 2 (PAR-2) to prevent TNF-α-induced insulin resistance to promote skeletal muscle growth and repair (326). Additionally, RT inhibits macrophage-induced inflammation and increases the MuSC number to relieve inflammatory myositis (327). RT increases the sensitivity of CD4+ and CD8+ T-cells to promote the secretion of myokines and cytokines (328). The expression of T-cell-specific inflammatory genes changes after resistance exercise, which is associated with muscle strength (80). The killer T-cell-specific marker CD8α can be activated after RT to eliminate senescent fibroadipogenic progenitors and inhibit the senescence-associated secretory phenotype in skeletal muscle (329) (Fig. 5).

In a hypoxic state, RT can induce angiogenesis in muscle tissue to increase muscle strength and endurance (330). Endurance exercise is an effective supplement to RT for reducing senescence-prone T-cell-induced inflammation and oxidative stress damage (331, 332). Both endurance exercise and resistance exercise can promote collagen deposition to prevent muscle atrophy (333). Alternating endurance and resistance training increases the expression of Pax7 and Myf5, reduces body fat, and improves skeletal muscle quality (334). The combination of endurance and resistance promotes adjustments in endoplasmic reticulum stress by inhibiting the expression of CHOP and p-eIF2α/eIF2α to increase skeletal muscle strength (335). Concurrent resistance and endurance exercise training also restored the inflammatory and fibrotic transcriptomes of myofibers to improve muscle performance (336) (Fig. 5).

Nutrients

Adequate diet and dietary supplementation with muscle-targeted food are important interventions for the treatment of sarcopenia (337). High-quality proteins are the basic nutrients for amino acid intake, while vitamin D, fatty acids, and probiotics are also important in nutritional intervention (338).

Whey protein is considered an important supplement for improving muscle strength and physical performance (339). Whey protein increases the production of IGF1 and albumin to increase muscle strength (340). Whey protein also promotes mitochondrial biogenesis and antioxidant effects to induce muscle growth and prevent myoblast apoptosis via the PI3K/Akt/PGC-1α and MAPK/ERK signaling pathways (44). Post-exercise whey protein supplementation is beneficial for adapting to endurance and resistance training (341). Whey protein also increases blood leucine levels to promote protein synthesis in skeletal muscles (342). Leucine supplementation activates the mTOR signaling complex to induce muscle formation (343). Leucine also increases the mitochondrial content to improve muscle strength (344). Additionally, other branched amino acids improve muscle mass and strength (345). Branched amino acid supplementation can improve insulin resistance to increase muscle mass and prevent SO (346). Gamma-aminobutyric acid (GABA) promotes myogenesis to prevent SO by activating the PI3K/Akt pathway (347). GABA activates Nrf2 signaling to increase the activity of antioxidant enzymes and increase the expression of phase II enzymes to prevent oxidative damage in myoblasts (348) (Table 3).

Table 3.

Nutrients for the treatment of sarcopenia.

Nutrients/effects	Mechanism	Reference
Whey protein
Enhances muscle strength	Increasing the production of IGF1 and albumin	(340)
Induces muscle growth and prevents myoblast apoptosis	Promoting mitochondrial biogenesis and antioxidant effects via the PI3K/Akt/PGC-1α and MAPK/ERK signaling pathways	(44)
Adapts to endurance and resistance training	-	(341)
Promotes protein synthesis in skeletal muscles	Increasing blood leucine levels	(342)
Leucine
Induces muscle formation	Activating mTOR signal complex	(343)
Improves muscle strength	Increasing mitochondrial content	(344)
Branched amino acids
Improves muscle mass and strength	-	(345)
Enhances muscle mass and prevent SO	Improving insulin resistance	(346)
GABA
Promotes myogenesis to prevent SO	Activating the PI3K/Akt pathway	(347)
Prevents oxidative damage of myoblasts	Activating Nrf2 signal to elevate the activity of antioxidant enzymes and increase the expression of phase II enzymes	(348)
Vitamin D
Increases the expression of MHC isoforms and myotube size	Combining with the receptor on the MyoG	(351)
Prevents the aggravation of sarcopenia and the occurrence of SO	Promoting mitochondrial biogenesis and enhances oxidative respiration	(352)
Relieves muscle atrophy	Improving mitochondrial function to inhibit oxidative damage and repairs neuromuscular junctions	(353)
Enhances muscle regeneration	Inhibiting inflammation and autoimmune by regulating intracellular metabolism and mitochondrial activity	(354)
Enhances myogenic differentiation	Activating of VDR to increase the expression of SIRT1 and SIRT3 and promote the phosphorylation of AMPK and AKT	(356)
Inhibits fat infiltration in muscle and prevents the development of sarcopenia	Transduction of vitamin D signaling	(357)
Omega-3 polyunsaturated fatty acids
Improves muscle atrophy	Maintaining muscle volume and relieving fat infiltration	(360, 361)
Improves muscle membrane composition and function	Inhibiting inflammatory response and oxidative stress, activating mTOR signaling, and reducing insulin resistance	(362)
Promotes the synthesis of muscle fibers	Relieving IL-10-induced inflammatory storm and elevating the antioxidant activity	(363)
Caprylic acid
Promotes the expression of myogenic differentiation 1 and myosin heavy chain	Activating Parkin-mediated mitophagy to regulate mitochondrial quality control and relieving oxidative stress	(364)
Probiotics
Improves sarcopenia and its complications	Repairing of intestinal pathological changes	(367)
Elevates muscle mass, quality, strength, and functions	-	(368, 369)
Regulates the intestinal microbiota to relieve the muscle damage	Increasing the expression of PGC-1α, SIRT1, and myosin heavy chain, inhibiting the expression of MuRF1, muscle atrophy F-box, and p16 via gut microbiota-mediated AKT, NF-κB, and FOXO3a signaling pathways	(370, 371)

The vitamin D level is negatively associated with the risk of sarcopenia (349). Vitamin D plays an important role in maintaining the structural integrity and function of skeletal muscle (350). Vitamin D combines with the receptor on the MyoG promoter to increase the expression of MHC isoforms and myotube size (351). Vitamin D promotes mitochondrial biogenesis and enhances oxidative respiration to prevent the aggravation of sarcopenia and the occurrence of SO (352). Vitamin D supplementation improves mitochondrial function to inhibit oxidative damage and repairs neuromuscular junctions to relieve muscle atrophy (353). Vitamin D inhibits inflammation and autoimmunity to enhance muscle regeneration by regulating intracellular metabolism and mitochondrial activity (354). The vitamin D receptor (VDR) mediates the insulin response of skeletal muscle (355). VDR deficiency leads to dysfunction of carbohydrate utilization in myocytes, decreasing energy supplementation. Activation of the VDR increases the expression of SIRT1 and SIRT3 and promotes the phosphorylation of AMPK and AKT to increase myogenic differentiation (356). The transduction of vitamin D signaling contributes to inhibiting fat infiltration in muscle and preventing the development of sarcopenia (357) (Table 3).

Fatty acids are important supplements for the prevention and treatment of sarcopenia (358). The essential intake of polyunsaturated fatty acids helps alleviate exercise-induced muscle damage by increasing total antioxidant capacity (359). ω-3 polyunsaturated fatty acids maintain muscle volume and relieve fat infiltration to improve muscle atrophy (360, 361). ω-3 polyunsaturated fatty acids improve muscle membrane composition and function by inhibiting the inflammatory response and oxidative stress, activating mTOR signaling and reducing insulin resistance (362). ω-3 fatty acids relieve the IL-10-induced inflammatory storm and increase antioxidant activity to promote the synthesis of muscle fibers (363). Caprylic acid, a medium-chain fatty acid for energy supplementation, promotes the expression of myogenic differentiation 1 and myosin heavy chain by activating Parkin-mediated mitophagy to regulate mitochondrial quality control and relieve oxidative stress (364) (Table 3). However, excessive intake of fatty acids increases the production of aldehydes, which could aggravate oxidative damage to myocytes (365). Plasma detection in sarcopenic patients has revealed that high levels of 7-ketocholesterol and 7β-hydroxycholesterol are associated with oxidative stress damage and the secretion of inflammatory factors in myocytes and myotubes (366).

Probiotics are involved in the repair of intestinal pathological changes to improve sarcopenia and its complications (367). Clinical statistics suggest that probiotics are positive for muscle mass and function (368). Probiotic supplementation also elevates muscle quality and strength (369). The intake of probiotics regulates the intestinal microbiota to relieve muscle damage (370). Probiotics increase the expression of PGC-1α, SIRT1, and myosin heavy chain and inhibit the expression of MuRF1, muscle atrophy F-box, and p16 via the gut microbiota-mediated AKT, NF-κB, and FOXO3a signaling pathways (371) (Table 3). Owing to the positive effects of probiotics, fecal microbiota transplantation from young donors has been confirmed to remodel the composition and metabolites of the gut microbiota, repair the gut barrier, and improve mitochondrial physiology in muscles (372). The targeted gut microbiota improves macrophage- and T-cell-induced chronic inflammation to resist muscle loss and maintain muscle health (85). As gut-derived nutrient-stimulated hormones, incretins can relieve insulin resistance to optimize body composition by reducing fat content (373).

Discussion

Skeletal muscle is the foundation of the daily behavior and life activities of humans. Muscle atrophy is an important sign of aging and increases the risk of fractures and joint injuries. The aging of skeletal muscles generally results in an average annual decrease of approximately 8% in both quantity and quality. If young people lack exercise and have insufficient muscle reserves, their muscles will age faster than those who exercise regularly in old age. The proliferation and differentiation of myoblasts maintain muscle mass. The theory of muscle filament gliding involves the process of muscle contraction. ECC is the basis for skeletal muscles to receive signals and coordinate movements. With changes in the human diet structure and lifestyle, the incidence rate of sarcopenia is increasing annually, which seriously threatens the life, health, and daily activities of patients. We aimed to explore the pathogenesis of muscle atrophy, identify high-risk populations for sarcopenia, and optimize treatment plans.

In this review, we demonstrated the main pathological changes associated with sarcopenia, including oxidative stress, inflammation, insulin resistance, hypoxia, and imbalance of the gut microbiota. Mitochondria play important roles in the regulation of redox balance during the development of sarcopenia. The activity of antioxidant enzymes in mitochondria not only participates in the clearance of ROS in muscles but also modulates the Ca²⁺ concentration at neuromuscular junctions. Moreover, mitochondrial respiration provides energy for muscle contraction, and mitophagy improves the regenerative ability of MuSCs. Macrophages and T lymphocytes mediate inflammatory damage in muscle tissue. Macrophages constitute the majority of immunocytes in the microenvironment of skeletal muscle. The harmful polarization of macrophages changes collagen synthesis in the extracellular matrix and the secretion of inflammatory factors to destroy muscle fibers. The effects of different types of T-cells on muscle tissue differ. Defects in or overactivation of T lymphocytes inhibit the differentiation and maturation of myoblasts. Stable insulin sensitivity is crucial for skeletal muscle cells to utilize nutrients and release energy. Insulin resistance inhibits the oxidation of glucose, amino acids, and fats, resulting in growth inhibition and fat infiltration in muscle tissue. HIF-mediated hypoxic signals not only inhibit mitochondrial OXPHOS and protein synthesis to induce myogenic inhibition but also aggravate the intracellular lipid peroxidation and ferroptosis of myoblasts. Disturbance of the gut microbiota influences substance metabolism, causing abnormalities in mainly mitochondrial metabolism and inflammatory responses. Additionally, these pathological changes are not isolated. Oxidative stress and the inflammatory response destroy the balance of epigenetic modifications and the composition of the gut microbiota (374). Oxidative stress and insulin resistance are considered to have interactive effects on sarcopenic inflammation, and inflammatory bowel disease is the research focus for revealing the role of the gut microbiota in the development of sarcopenia (375). Patients with chronic wasting diseases, obesity, and osteoporosis are at high risk of sarcopenia. Oxidative damage and inflammation are common in these patients.

At present, nutrition supplementation and RT are important nonpharmacological strategies for preventing the deterioration of sarcopenia (376). Whey protein is the optimal nutrient for increasing muscle mass and strength in aged patients with sarcopenia undergoing RT (377). RT combined with testosterone, calcium, vitamin D, and protein intake decreases leg fat and tiredness and improves muscle quality (378). However, the combination of dietary supplements and exercise therapy seems difficult to achieve for some patients who have lost their motor function or have eating disorders. Pharmacotherapy is generally considered an adjunctive option under specific circumstances. Myostatin is a protein that inhibits muscle growth. Suppressing its activity can promote muscle growth. Myostatin inhibitors, such as stamulumab and landogrozumab, represent one of the most promising future directions (379). Selective androgen receptor modulators (SARMs) act selectively on androgen receptors in muscles and bones, promoting anabolic effects while avoiding the severe side effects associated with traditional steroids. Their representative drug is enobosarm. Clinical trials have demonstrated its potential to increase muscle mass and strength, although long-term safety and efficacy require further validation (380). Hormone-based therapies represent another potential option. Once weekly, intermittent administration of glucocorticoids contributes to improved muscle mass by activating PGC-1α to increase mitochondrial abundance (381). Therapeutic ultrasound enhances the signal transduction of cytokines and inhibits cell recruitment in muscle, together regulating the number of neutrophils and monocytes and the ratio of M1/M2 macrophages to repair muscle fibers and induce MuSC differentiation (382). Taurine supplementation is helpful for improving gut microbiota homeostasis to prevent SO by inhibiting inflammation and oxidative stress in muscle and adipose tissues (383). The optimization of lifestyle and psychosocial support are also essential. Quitting smoking and moderating alcohol consumption contribute to improved muscle health. Quality sleep provides a critical window for muscle repair and growth hormone secretion. Social engagement and psychological support, including encouragement to participate in group activities and exercise programs, help reduce loneliness and enhance motivation for treatment adherence. Additionally, the prevention of complications and psychological distress associated with sarcopenia is particularly crucial. Declining muscle strength and balance lead to unsteady gait, increasing the risk of falls. From slowed walking speed to reliance on assistive devices, the condition may ultimately progress to long-term bedridden states and loss of independent living capacity. Daily activities, such as climbing stairs, shopping, and bathing, become challenging, resulting in reduced social participation and elevated risks of depression and anxiety. Early screening, diagnosis, and intervention for sarcopenia are crucial. Assessment of physical function and balance ability, including the timed op and go test, short physical performance battery, one-legged stance test, and gait observation, can help predict the risk of falls. Home environment assessment is a frequently overlooked yet critical component. Ensuring adequate lighting, level flooring, installing non-slip mats and grab bars in bathrooms and toilets, and placing commonly used items within easy reach can significantly reduce the risk of accidents among older adults. Ensuring that patients are free from cataracts, glaucoma, and uncorrected vision problems and have normal vestibular function and proprioception is a prerequisite for preventing the aforementioned complications. In conclusion, the treatment strategies for sarcopenia should be multifaceted.

Our research group suggested that tissue synthesis and energy supply constitute the foundation for maintaining the normal physiological functions of muscles. Adequate supplementation of nutrients is indispensable. Under this precursor, mitochondrial function is crucial for ensuring the energy supply needed for muscle contraction. Mitochondrial antioxidant effects and mitophagy also contribute to improving pathological changes in the muscle tissue of sarcopenic patients. Targeted delivery of functional mitochondria to MuSCs is a potential method to increase local biological oxidation and accelerate muscle repair (384). Additionally, cross-species pluripotent stem cell transplantation has also been shown to have the potential to induce the development of human muscle cells (385). Intramuscular injection of the stem cell secretome decreases lipid content and increases Pax7-positive myocyte abundance (386). IGF1 mediates the positive effect of exercise on muscle repair (387). IGF1 supplementation elevates oxidative and insulin-sensitive metabolism to increase myofibril formation for muscle contraction (388). Stem cell transplantation with the stimulation of growth factors may be an available method for the further treatment of sarcopenia.

ICMJE Statement of Interest

The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.

Funding Statement

This study was supported by a project funded by the National Science Fund for Distinguished Young Scholars (32200943), Shenyang Young and Middle-Aged Innovative Talents Project (RC210171), and China Postdoctoral Science Foundation (2022M723520).

Author contribution statement

KY curated the data, designed the methodology, acquired software, and wrote the original draft of the manuscript. SP curated the data, acquired software, and wrote the original draft of the manuscript. CZ designed the methodology, acquired software, and wrote the original draft of the manuscript. LG designed the methodology, performed validation, and reviewed and edited the manuscript. DL curated the data, acquired software, performed validation, and reviewed and edited the manuscript. LT acquired funding and resources, administered the project, and reviewed and edited the manuscript. All authors read and approved the manuscript.

Data availability

The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.