Skeletal muscle (SkM) is the body's largest organ, accommodating 40–50% of body mass. It is vital in controlling musculature, locomotion, body heat regulation, physical strength, and metabolism (Yadav et al., 2021). Skeletal muscle consists of distinct/different bundles of muscle fibers subtypes, collectively called fascicles, which form the whole muscle surrounded by sarcolemma, enclosed with cellular protein, myofibers, and organelles (Fig. 1). Skeletal muscles appear striated due to the organized arrangement of their contractile proteins called myofilaments, which consist of thin actin and thick myosin filaments. Interaction among them enables skeletal muscle contraction and relaxation, which regulates the limb's movements and maintains posture and individual activities (McCuller et al., 2023).

Myofibers of skeletal muscles comprise slow type-I twitch fibers (oxidative) and fast type-II twitch fibers (glycolytic) and are responsible for muscle endurance, strength, and fatigue. Skeletal muscles are primarily responsible for energy expenditure through ATP production, metabolic adaptations, glucose metabolism, homeostasis, etc. (Plotkin et al., 2021). Therefore, skeletal muscle regulation and maintenance are critical for protection against various metabolic disorders and physiological strains. Hence, skeletal muscle atrophy is a multi-regulatory disorder involving increased catabolism in the skeletal muscles. Muscle atrophy due to denervation, cachexia, disuse, altered metabolism, and reduced physical activity contribute to increased protein breakdown and decreased protein synthesis, leading to muscle wasting (Fig. 2). Therefore, the balance between synthesis and degradation is necessary to maintain the quality of life of individuals, mortality, and morbidity (Yadav et al., 2022).

Muscle innervation is critical for maintaining muscle tone and physiological functioning. In response to stimulation, nerve fibers release acetylcholine (ACh), which causes depolarization and polarization of SkM and promotes several intertwined intracellular signals and biological processes. Sciatic nerve injury abrupts neuronal signals to the muscle, characterized by progressive loss of motor neurons and ACh at neuromuscular junctions (Fig. 3). An interruption of the neuronal signal at the NMJ increases acetylcholine receptor (AChR) expression at the extra junctional membrane to extend myofibers depolarization, thereby increasing the permeability of the sarcolemma by opening calcium-gated channels and connexin/pannexin hemi channels (Cisterna et al., 2020). It led to increased Ca2+ influx in sarcoplasm and opening of mPTP and depolarized mitochondria, which not only stimulate ROS production but also initiate the fiber type switching in myofibers through inhibition of Sestrin2 (SESN2) (Duregotti et al., 2015; Kallenborn-Gerhardt et al., 2013). Sciatic damage was estimated to contribute to 90% inhibition of muscle mass. Moreover, soleus muscles retained only 8% contractibility in 14-week denervated rats. At the same time, the tibial anterior experienced 3–5% contraction in long-term denervation, leading to anonymous changes in muscle pool and T-tubulins and the heterogeneity of mitochondria (Carlson, 2014). Increasing evidence suggests that abnormal Ca2+ flux into sarcoplasm to mitochondria is a crucial factor in increased chronic oxidative stress and proinflammatory cytokine/chemokines for the activation of multifactorial sequelae of proteolysis following neuronal damage (Sanganalmath et al., 2023). Oxidative stress strongly inhibits the PI3K/AKT signaling pathway that activates the FOXO and NF-κB signaling pathways. Calcium influx also activates cAMP, cGMP, calcineurin signaling, mitophagy, and apoptosis (Yadav et al., 2022). Denervation-induced muscle loss, however, remains elusive regarding expression and linkage between different signaling pathways. Thus, the present review aims to understand the mechanism of impaired Ca2+ signaling and the relationship between the independent and intertwined signaling pathways after sciatic nerve damage.

The nervous system controls skeletal muscle primarily by two different mechanisms: the neuromotor type, which transmits motor nerve impulses, and the neurotrophic type, which encourages the growth and maintenance of SkM via a specialized structure known as neuromuscular junctions NMJ (Fig. 4). The neuromuscular junction is a well-organized and complex structure differentiated into three parts: the nerve terminal (presynaptic zone), synaptic cleft, and motor end plate (postsynaptic zone). The nerve terminal region comprises synaptosomal-associated protein 25 (SNAP25) and syntaxins (SNARE) family protein and calcium voltage-gated channels (Davis et al., 2022). In SkM, the motor neuron forms a branching mesh at the plasma membrane, which contains numerous mitochondria, mature 2α1β1δε acetylcholine receptors, and acetylcholine-filled synaptic vesicles. The walls of synaptic vesicles have synaptobrevin and synaptotagmin, which regulate the action potential through their ability to bind with SNARE proteins for effective exocytosis and Ca2+ sensing (Sauvola and Littleton, 2021). Various vital molecules and signaling pathways regulate the development and stability of the NMJ, including agrin, neuregulin, muscle-specific kinase (MuSK), and their downstream effectors. These molecules mediate the clustering of AChRs on the postsynaptic muscle membrane, the organization of the presynaptic nerve terminal, and the alignment of pre- and postsynaptic components (Zong and Jin, 2013). Apart from AChRs, a small number of dopamine and serotonin receptors are also found to be present at sarcolemma for the coordination of muscle movements (Reichart et al., 2011). However, their independent role in the skeletal muscle is still elusive.

Nicotinic AChRs (nAChR) are more abundantly present in SkM than other neurotransmitters. Once the motor neuron transmits the signal, it triggers the release of ACh through exocytosis in the synaptic cleft to interact with the nAChR. On binding, AchR increases the permeability of voltage-gated Na+ and Ca2+ channels in the postsynaptic membrane and promotes Na+ and Ca2+ influx into the sarcoplasm for depolarization and contraction of muscles (Liu and Chakkalakal, 2018). A protein rapsyn makes clusters with acetylcholine receptors at NMJs and binds with CDK2 and MuSK-agrin-LRP4, which further stimulates propagation. Additionally, this propagation is further supported by mAChR through protein kinase C (PKC)-dependent signaling (Wang et al., 2020; Zong and Jin, 2013). The isoforms of PKC viz., PKCε and PKCβI are essential for releasing AchR at NMJ. Phosphorylated PKCε(ser313) and PKCβI(Ser306) in association with its substrates Munc 18-1, an accessory SNARE protein, and SNAP-25 form PKC complex to activate nAchR. On activation, PKC phosphorylates Ser 187 subunits of SNAP-25 to initiate calcium-dependent exocytosis and vesicle fusion at the presynaptic terminal. MARCKS regulates the PKC phosphorylation, which helps reorganize the actin in the cytoskeleton toward extracellular responses (Cilleros-Mañé et al., 2021).

Additionally, depolarization also activates L-type calcium channels (Cav1.1), which control ryanodine receptors (RyR) on the sarcoplasmic reticulum (SR) and release a significant amount of Ca2+ into the cytosol (Andersson and Marks, 2010). This Ca2+ binds to troponin C and forms a cross-bridge between the myosin and actin filaments, which shorten the sarcomere using ATP and cause force generation. During the polarization, Ca2+ from the sarcoplasm is pumped back to SR-by-SR calcium-ATPase (SERCA) pumps following muscle relaxation. This depolarization and polarization are also defined as excitation-contraction (E-C) coupling (Gehlert et al., 2015).

Nerve transaction starts depolarization in fast muscle within 2 h, which increases Na+ and decreases K+ in myoplasm and changes action potential by a decreased Na+ current inactivation of Na+ channel. Terminated muscle impulse also increases action potential duration and muscle contraction time, so prolonged depolarization keeps open Ca2+ release channels. The consequent increase in myoplasmic Ca2+ levels also increase the Ca2+ release from SR and reduces the Ca2+ removal rate, increasing the Ca2+ sensitivity of myofibers and disturbing the Ca2+ buffer capacity (Hong and Chang, 1997). Overall, due to ion leakage through sarcolemma, increased impulse generation threshold after nerve transection cannot reach the point that many fibers, especially soleus muscles, cannot generate spikes.

The prolonged decline in the resting membrane potential enhances the expression of nAChR at the end plate of muscle and degenerate NMJs. NMJs degeneration causes the opening of Ca2+ channels, connexins hemichannels, and the release of myokines, neurotrophin-3/4 (NT3/4), brain-derived neurotrophic factor (BDNF), and ciliary neurotrophic factor (CNTF), to degenerate myofibers and axons (Zhao et al., 2004). Axon degeneration increases the sensitivity of muscles towards succinylcholine, which increases the hyperkalemic responses at NMJ to increase depolarization further (Martyn et al., 2006). Sciatic resection increases the expression of non-depolarizing neuromuscular blocking agents (NDMRs) at the synaptic cleft of NMJs to inhibit the binding of nicotinic AchR and generate the action potential (Kim et al., 2017). Interestingly, under normal conditions, the stability of NMJs is critically regulated by E3 ligase MuRF-1 at NMJs in a lysosomal-dependent manner. Denervation also inhibits the expression of rapsyn and PKA and increases the co-localization of MuRF-1-Bif-1 endocytically to destabilize the AchRs at NMJs, resulting in NMJ degeneration and remodeling (Rudolf et al., 2013).

In addition, nerve axotomy increases the expression of miR-206, which is found in the synaptic area of myofibers, and inhibits histone deacetylase 4 (HDAC4), and fibroblast growth factor-binding protein 1 (FGFBP1) at the NMJ to activate the degenerative signaling pathways by connexin43, Fstl1, and Pax 3/7. Upregulated connexin43/45 has been shown to enhance the ionic unbalance that instigated the production of ROS and decrease the quality of the mitochondria for increased muscle fibrillation (Chen, 2020). Recently, it has been believed that autocrine activation of AChRs is related to fibrillation in muscle fibers (Pond et al., 2014). On the contrary, electrical stimulation after sciatic injury triggers nerve sprouting and axonal regeneration at NMJ via increased production of nerve growth factor, which results in the opening of Na+-gated channels at NMJ for membrane excitation and inhibits Ca2+-gated channels (Samiee and Zarrindast, 2017).

Connexins and pannexins are the hemichannels or gap junctions that control the ionic and small molecule's mobility at NMJs. Skeletal muscles express connexins hemichannels before the late differentiation phase and remain quiescent until the muscle encounters any injury. While pannexins consistently express themselves to maintain the membrane potential by releasing ATP, NO, prostaglandin, etc. Any metabolic alteration like deficient Na+/K+ ATPase, denervation, aging, and decline resting membrane potential strongly upregulate the connexin/pannexins channels in sarcolemma to adaptive response and intracellular communication (Cea et al., 2012).

Connexins and pannexins hemichannels opening after sciatic transection triggers the outflow of ATP molecules, which activates the purinergic receptor at NMJ to release inflammatory myokines, and reduces the axonal regeneration for neuronal signaling. Decreased resting membrane potential across sarcolemma was also reported to upregulate the expression of Cnx39, Cnx43, 45, panexin1, TRPV2, and P2X7R (Cea et al., 2013). Cnx 43 hemichannel releases ATP into the extracellular space, activating purinergic receptors and directing the assembly of inflammasome NOD-like receptor protein 3 (NLRP3)-procaspase 1 complex to activate apoptosis. Additionally, the opening of these channels directly impacts mitochondria quality and generates ROS for the release of TNF-α, IL-1β, vascular endothelial growth factor, and NF-κB transcript in the extracellular space, including synaptic cleft (Gombault et al., 2012).

Continuous opening of Cnx43 and Cnx45 caused the marked decline in (∼75%) fiber cross-section area due to activation of μ-calpain, and p-p70S6K in skeletal muscles (Cisterna et al., 2016). Increased expression of purinergic receptors and mobilization of Ca2+ triggers the calcineurin/NFAT signaling of muscle proteolysis (Kipanyula et al., 2016). Similarly, denervation increases the opening of Panx1/3 hemichannels via binding with T-tubulin and releases the sarcoplasmic Ca2+ in an exocytosis manner (Cea et al., 2014). Therefore, control at the opening of hemichannels is critical for maintaining ionic and ATP balance for proper neuronal conductivity.

Normally, 50–100 nm free Ca2+ in the cytosol and 0.5–1.0 mM in SR is crucial for maintaining muscle homeostasis and contraction (Schwaller, 2010). In response to denervation, the NMJ stops ACh release and maintains the myofibers in a depolarized state. Depolarization can accumulate Ca2+ in the cytosol through various molecular mechanisms, changing the Ca2+ levels in different organelles and impacting the Ca2+ signaling pathways. First, it increases the permeability of the sarcolemma, expression of voltage-gated calcium channel (VGCC), and nAChR that prolongs myofibril depolarization and promotes gradient base increase of Ca2+ in the cytosol. Second, Ca2+ influx through sarcolemma increases due to a defective activity of the K+/Na+ ATPase pump and reduced membrane potential (Vm) (Ferraro et al., 2012). The opening of connexin43, P2X7Rs, Panx1 hemichannels, etc., reduces the activity of the ATPase pump, which efficiently pumps out the intracellular ATP and infuses the Ca2+ intracellularly (Kameritsch and Pogoda, 2020). Third, RyRs respond to several kinds of cytosolic signals and integrate their multitude to cope with steady and dynamic state of calcium fluctuations, phosphorylation (β-adrenergic stimulation), metabolic states, and nitrosylation to release appropriate amounts of Ca2+ (Kobayashi et al., 2021). However, sarcolemma depolarization promotes dihydropyridine receptor (DHPR or Cav1.1) activation, the regulator of RyR in the SR membrane, which boosts up Ca2+ release into the cytosol. Fourth, store-operated calcium entry (SOCE) is another mechanism responsible for replenishing the muscle SR Ca2+ by modulating the expression of proteins STIM1 and Orai1 (Agrawal et al., 2018). According to reports, decreased Orai1 gene expression is linked with specific forms of muscle weakness and can improve neuromuscular transmission following denervation (Mosqueira et al., 2021). Hence, the continuous influx of Ca2+ prolongs AchR exposure and depolarizes myofibers that cause myofiber fibrillation.

The increased concentration of cytosolic Ca2+ affects several other cell organelles and enzymes, including mitochondria, essential for ATP synthesis. The Ca2+ electrochemical gradient in mitochondria primarily regulates Ca2+ uptake and acts as a buffering agent in the mitochondrial matrix and cytosol (Rossi et al., 2019). According to a recent study, mitochondrial protein parvalbumin also helps to buffer the mitochondrial Ca2+ pool after denervation on the 3rd and 7th day. Mitochondria maintain their matrix Ca2+ levels using PTP and mitochondrial calcium uniporter (MCU) complex. The PTP opens during the depolarized state of the myofibers and remains open during the prolonged depolarized state after denervation. However, the MCU's rate of taking up Ca2+ is inversely proportional to extra-mitochondrial Ca2+ (Butera et al., 2021). Hence, prolonged depolarization and increased concentration of cytosolic Ca2+ overloads the mitochondria matrix with Ca2+ through the MCU complex. The MCU dimeric complex at the inner mitochondrial membrane comprises MICU1 and MICU2 and the central subunit essential MCU regulator (EMRE). MICU1 and MICU2 increase the membrane threshold to allow Ca2+ uptake through the MCU complex (Wu et al., 2020). The transient receptor channel 3 (TRPC3), uncoupling proteins 2/3 (UCP2/3), and LETM1 also support MCU for Ca2+ uptake in the mitochondrial matrix (Matuz-Mares et al., 2022).

Apart from E-C coupling, which keeps muscles in the strained form in the case of excess Ca2+, accumulation of Ca2+ in mitochondrial through MCU and prolonged opening of PTP results in uncoupling of an electrochemical gradient, which prevents ATP synthesis and ATP hydrolysis, therefore, altered Ca2+ homeostasis involves in cell death paradigms (Wu et al., 2020). Uncoupling of the electrochemical gradient in mitochondria leads to ATP depletion and ROS generation, which subsequently affects several cellular processes and signaling pathways, including SR Ca2+ pumps regulation through RyRs and IP3Rs, which are sensitive to the redox state of the cell and S-nitrosylation (Madreiter-Sokolowski et al., 2020). ATPase pumps push Ca2+ into or out of the cell permanently, and ATPase is required for E-C coupling, affecting purinergic signaling pathways and apoptosis. The Ca2+- levels also affect the transcription factors (Brini and Carafoli, 2009). For example, Ca2+-dependent CaMK promotes myogenesis by inhibiting the formation of complexes between transcription factors myocyte enhancer factor 2 (MEF2) and histone deacetylases (HDACs) and phosphorylating HDAC4 and HDAC5 to promote their nuclear export (Ginnan et al., 2012).

Intercellular Ca2+ signaling takes place through gap junctions and transmitter-gated ion channels (Fig. 4). Voltage-gated channels increase periplasmic Ca2+, which triggers protein fusion, and vesicles containing transmitter molecules to fuse to the plasma membrane and dominate neuron health and communication (Brini and Carafoli, 2009; Mosqueira et al., 2021). Transient receptor potential (TRP) ion channels are weakly voltage-sensitive, nonselective channels that span only intracellular membranes. Most TRP channels are plasma membrane channels that depolarize cells and increase intracellular Na+ and Ca2+. Cytosolic Ca2+ activates phospholipase C (PLC), which potentiates TRP through G protein-coupled or tyrosine-kinase receptors (TKR) and modulates sensory systems after being activated by environmental signals (Gees et al., 2010). However, the central unanswered question in this field is how TRP channels get activated in vivo. Cytosolic Ca2+ activated PLC also activates protein kinase C (PKC) through diacylglycerol, which has been reported for anti-proliferative effects, whereas PKCδ is emerging as a significant negative regulator. Moreover, PKC phosphorylates various protein targets and participates in cellular signaling pathways. PKC family members have distinct expression profiles and roles in various processes, including cell adhesion, transformation, and volume control (Kolczynska et al., 2020).

Accumulation of Ca2+ also affects several signaling pathways essential to regulate apoptosis, cell metabolism, proliferation, mitogen-activated protein kinase (MAPK) and PI3K signaling pathways, and various other signaling pathways. To promote gene transcription and protein synthesis, Ca2+ signals to activate downstream protein kinases (MAPK) and immediate early genes (IEGs), which phosphorylate ERK. ERK cascades involved in fundamental cellular functions like cell differentiation and proliferation are tightly controlled cascades (Patergnani et al., 2020). Moreover, Ca2+ also compromises the cAMP/PKA signaling pathway. Disruption of nerve input declines the cAMP production, which reduces the phosphorylation of proteins (synapsin) involved in synaptic vesicle exocytosis and opens the Ca2+ channels (Menegon et al., 2006). Furthermore, an increase in mitochondrial calcium involves the upregulation of the PGC-1α4 signaling, a regulator of muscle hypertrophy (Butera et al., 2021).

Hence, it has been widely established that Ca2+ signaling is involved in various stresses, including those caused by high or low temperatures, oxygen stress, pathogenic agents, hypoxia, osmotic stress, and heavy metal ions (Brini and Carafoli, 2009; Mosqueira et al., 2021; Patergnani et al., 2020). However, the exact mechanisms and consequences of altered Ca2+ ion influx after denervation are complex and need further research to fully understand the intricated interactions among the denervation, Ca2+ channels, and Ca2+ signaling in muscle fibers.

Sciatic nerve resection elevates Ca2+ levels in the cytosol as well as in the mitochondrial matrix (Fig. 4). Cytosol accumulates a high concentration of Ca2 after prolonged depolarization due to denervation; hence, MCU complex transports Ca2+ to the mitochondrial matrix on a gradient basis from the cytosol at the initial stage. This steady-state elevation of Ca2+ level in the mitochondrial matrix exacerbates ROS generation, and prolonged polarization sensitizes the opening of mPTP, induces programmed cell death, and ultimately muscle atrophy (Patergnani et al., 2020; Wu et al., 2020).

Elevated levels of Ca2+ promote the activity of glycerol phosphate dehydrogenase, isocitrate dehydrogenase, α-ketoglutarate dehydrogenase, and OXPHOS complexes in mitochondria, which increases O2 consumption rate, free radicles and uncouple the oxidative phosphorylation (Li et al., 2020). However, it has been found that an early rise in oxidative stress, after denervation, is supported by up-regulation of cytochrome P450 and mono-oxygenase transcripts (Shen et al., 2019). Ca2+ also promotes the activity of nitric oxide synthase (NOS), responsible for increasing nitric oxide (NO) production, which competes with O2 for binding on complex IV/cytochrome c oxidase and impedes electron flow increases the local O2 which, further increase ROS/RNS production. ROS reacts with NO and produces peroxynitrites (ONOO−), for oxidation of proteins, especially the tyrosine residues of cysteine and promotes cytochrome c release and lipid peroxidation and damages several vulnerable targets (Tengan et al., 2012). Also, Increased ONOO−, oxidation of cysteine and carbonylated proteins reported to inhibit actin dependent ATPase-myosin activity due to increased unfolding of myosin subfragment-1 (S1) in muscles as confirmed by Tiago et al. (2006), 2010.

Elevated Ca2+ levels also promote the opening of mPTP, which changes the ionic strength and thus disrupts the electrostatic interaction between cytoC and cardiolipin in the intermembrane space (Fig. 5). It inhibits the activity of complex III, which increases the accumulation of the one-electron donor ubisemiquinone and, therefore, the production of ROS. Moreover, the opening of mPTP causes conformational changes in complex I and contributes to the increased production of ROS (Li et al., 2020). Moreover, denervation also increases the assembly of the Nox2 complex due to a profound increase in the interaction of gp91phox/Nox2 and p22phox after 14 days of post-denervation. Increased assembly of Nox2 is also supported by an increased content of Rac 1 and p47phox, p40phox, p67phox, and Prx 6. Active Nox2 is an enzyme that promotes the catalyzation of O2 into ROS (Scalabrin et al., 2019). A significant increase in ROS production was observed after 48 h of denervation, which may be in conjunction with increased defective and misplaced mitochondria at the A band of the sarcomere (Pietrangelo et al., 2019). It was demonstrated that denervation-induced ROS production in the soleus muscle down-regulates the expression of Pink 1, Prdx2, Sirt2, 3, 5. Declines in Sirt2, 3, and 5 levels suppress the transcript of MnSOD in the soleus muscle and are additive in oxidative stress (Qiu et al., 2018). Moreover, skeletal muscles lose their contractile function after nerve resection, causing a decline in blood flow and promoting a hypoxic state in muscles. Increased hypoxia contributes to the formation of reactive oxygen species (Jatwani and Tulsawani, 2021). Therefore, denervation supports the production of ROS/RNS in myofibers in a positive feedback manner.

Phosphoinositide-3-kinase/AKT serine-threonine kinase/mechanistic target of the rapamycin kinase (PI3K/AKT/mTOR) signaling pathways is the primary pathway for glucose metabolism as well as to regulate other signaling pathways related to skeletal muscle stress. PI3K/AKT/mTOR and oxidative stress have a reciprocal modulation of several biological processes, i.e., insulin resistance, apoptosis, autophagy, senescence, cell differentiation, etc (Shiau et al., 2022). Accumulated ROS/RNS oxidized the redox-sensitive cysteine residues of protein tyrosine phosphatase 1 B (PTP1B) and protein phosphatase 2 A (PP2A) and inactivate them, which in turn inhibits PI3K and Akt correspondingly and lead to insulin resistance in skeletal muscle (Lennicke and Cochemé, 2021). Inhibition of Akt also promotes the activation of FOXO transcription factors. ROS also activates FOXO3a directly through phosphorylation. PI3K and AMPK signaling pathways are the prominent stress signal sensors that regulate FOXO3a-mediated cellular homeostasis. Furthermore, the MAPK–FOXO3a axis is the core signaling pathway to maintain physiological balance in response to oxidative stress (Roy et al., 2010). The overproduction of ROS also activates the NF-κB transcription factor. Both FOXO and NF-κB increase the transcription of proteolytic enzymes and proinflammatory cytokines. ROS activates the NF-κB signaling pathway and upregulates and accumulates S100B required to transition myoblasts into brown adipocytes. Myoblasts and brown adipocytes share common Myf5+ progenitors and bone morphogenetic protein 7 (BMP-7; TGF family member) levels that decide cell fate (Morozzi et al., 2017).

The redox-sensitive MAPK activates several protein kinases, nuclear proteins, and transcription factors required for downstream signal transduction. Oxidative stress also activates functionally distinct MAPK pathways, i.e., p38 MAPK, c-Jun NH2-terminal kinase (JNK), and ERK 1/2, which promotes protein catabolism and apoptosis, leading to skeletal muscle atrophy (Sugden and Clerk, 1998). Gadd45a and Cdkn1a further upregulate ROS production through p38MAPK signaling. Activated p38MAPK was found to endorse the adult human stem cells toward senescence (Qiu et al., 2018). Oxidative stress has also been found to enhance the production of myostatin (MSTN), a protein that triggers the MSTN-Smad2/3 pathway, leading to reduced protein synthesis, raised levels of IL-6 through p38MAPK activation and eventually causing skeletal muscle atrophy (Liu et al., 2018).

Increased ROS and Ca2+ overload impaired the tubular network of mitochondria through upregulation of miR-142a-5p in denervated gastrocnemius muscle. Upregulated miR-142a-5p triggers the fragmentation of neuronal mitochondria via binding to the 3′UTR region of MFN1 mRNA. The binding of miR-142a-5p and MFN1 depolarizes the membrane potential of mitochondria to induce mitophagy responses (Yang et al., 2020). Elevated superoxides interact with the axonal transport of synaptic vesicles and inhibit the formation of new trans-golgi vesicles at the NMJ, resulting in a reduction in the release of acetylcholine at the NMJ, which disrupts proteostasis signaling by motor neurons (Jang and Van Remmen, 2011). Additionally, another study reported that Hsp 70 increases neurotransmitter release and motor neuron regeneration by interacting with the DnaJ-like chaperone cysteine string protein to prevent the development of ER stress and NMJ integrity (Gorenberg and Chandra, 2017).

In contrast, few studies claim that oxygen species alone cannot induce muscle atrophy but that NF-κB levels are elevated on day 3 of denervation, a primary inhibitor of the antioxidant system in skeletal muscle (Takayama et al., 2023). Despite everything, ROS of mitochondrial origin regulates muscle mass through fiber branching (Barbieri and Sestili, 2012). Therefore, the kinetics of ROS in sciatic injury associated with different molecular signaling pathways need to be explored. It will facilitate the discovery of a consistent mechanism of ROS release and associated activation of proteolytic systems in denervation-induced muscle loss.

High Ca2+ levels in cytosol activate several proteolytic enzymes, including caspases and calpains, which disintegrate muscle fibers (Patergnani et al., 2020; Schwaller, 2010). Muscle repair depends on satellite cell (SC), which differentiates into myotubes after acute or chronic injury (Mierzejewski et al., 2020). Mild levels of ROS are required to activate p38MAPK to corroborate IL-6/JAK/STAT and FGF/p38αβ MAPK signaling pathways to initiate SC differentiation (Belizário et al., 2016). In a state of equilibrium, the dormant SCs are arrested in the initial phases of myogenesis due to the expression of transcription factor Pax7 (Rigillo et al., 2021). A cascade of cytokines and growth factors are released during muscle injury, originating from the damaged myocytes (Tu and Li, 2023). The bioactive molecules increase the expression of Myf5 or MyoD, which are vital factors responsible for the SC proliferation and induction of cellular differentiation. Activated satellite cells, known as myoblasts, commence the proliferative stage which facilitates the development of myogenic progenitors. During this particular stage, there is a decrease in the expression of Pax7, while there is an increase in the expression of myogenin and Myf5. This subsequently leads to increased expression of surface proteins such as β1-integrin, caveolin-3, and VCAM to facilitate the process of fusion. Simultaneously, the heterodimerization of myogenic regulatory factors (MRFs) with enhancers, specifically MEF2, drives the expression of muscle-specific genes such as actins, myosins, and troponins (Yin et al., 2013). These genes are crucial for the proper morphology and function of skeletal muscle. Furthermore, activated SCs can exit the cell cycle without the induction of differentiation. They can self-renew to restore the population of quiescent SCs by delaying the drop in the expression of Myf5 and maintaining Pax7 protein, while losing MyoD (Yanay et al., 2020). This mechanism ensures a constant SC cell number, with the potential for myogenic differentiation, even after multiple rounds of activation. Consequently, this process ensures the life-long regenerative capacity of the muscle.

However, excessive ROS reversed the process and directed the pathways to inflammation. Sustained elevated ROS decreases glutathione (GSH), glutathione peroxidase (GPx), and nuclear factor erythroid 2–related factor 2 (Nrf2) levels, abrogates Nrf2 signaling that impairs skeletal muscle recovery (Fig. 6), and leads to muscle SC senescence. High ROS levels can result in loss of myoblast function and increased cell death. ROS/RNS depletes the GSH pool of myofibers that cause NF-κB activation, resulting in decreased MyoD expression. In addition, NF-κB also mediates the YY1 activation, an inhibitor of myogenic transcription. Hence, decreased MyoD expression and increased activation of YY1 discourage myoblasts from differentiation. Moreover, ROS declines p21 promoter activity, increasing myoblasts' apoptosis rate. It was also found that the activity of the p21 promoter in myoblasts decreased in response to high ROS and that apoptosis occurred in myoblasts to a greater extent than in myotubes. However, increased ROS levels are sensed by antioxidant enzymes, and levels elevate along with peroxisome proliferator-activated receptor coactivator-1 protein (PGC-1) promoter activity to cope with oxidative stress (Lian et al., 2022). Still, the high level of ROS/RNS modifies and inactivates these antioxidant enzymes and PGC-1α.

Skeletal muscles undergo different stages of regeneration, degeneration, and fibrosis after sciatic nerve damage to maintain the state of equilibrium. Oxidative stress is known to induce inflammation through the activation of several pathways (Liu et al., 2018). With increased oxidative stress, mitochondria deviate from normal ATP synthesis, Ca2+ signaling, and release mitochondrial-derived damage-associated molecules (MDPs) that interact with the environment, increasing the inflammatory response required for myofibril regeneration (Barbieri and Sestili, 2012; Scalabrin et al., 2019). However, with denervation, large-scale signaling associated with mitochondrial dysfunction can continue synthesizing cytokines and chemokines, resulting in chronic inflammation that damages muscle outcomes. Moreover, inflammation and oxidative stress reciprocate each other.

ROS can induce inflammation by activating the transcription factor NF-κB through TNF-α. The NF-κB pathway is a typical proinflammatory signaling pathway (Fig. 6), and it can promote the expression of inflammatory factors, including cytokines, chemokines, and adhesion molecules. Also, ROS damages the mitoDNA and leads to DNA base modification that induces inflammation by activating NF-κB (Morgan and Liu, 2010). It has been shown that base excision repair by 8-oxo guanine-DNA glyoxalase-1 culminates in the NF-κB pathway, hence inflammatory cytokines (Pan et al., 2016). Moreover, ROS-induced arachidonic acid fragmentation and oxidation generate 8-isoprostane, which has been shown to increase inflammatory chemokine IL-8 expression by activating MAP kinases (Scholz et al., 2003).

Furthermore, ROS oxidizes the cysteine (Cys) and its disulfide cystine (CySS) in various proteins, which trigger monocyte adhesion and activate NF-κB (Checa and Aran, 2020). As a result of ROS generation, tibia anterior (TA) muscles express TNF-α, IL-6, and IL-1β within 14 days of denervation. Elevated IL-6 levels activate its downstream JAK2/STAT3 signaling pathway in TA muscles, which increases expression of the suppressor of cytokine signaling (SOCS3) to inhibit IGF-1/Akt pathway by degrading IRS-1 that promotes insulin resistance as well as inflammation Huang et al., 2020. This process was reversed by salidroside and aspirin via down-regulation of IL-6 through sirtuin 1 (SIRT1)/PGC-α signaling and phosphorylation of Foxo3A and attenuated the fiber switching (Wan et al., 2020; Xue et al., 2019). Specifically, aspirin has also been reported to inhibit the IL-8, C-reactive proteins (CRP), HSP60, and TNF-α in circulation but remains unexplored in denervated-induced muscle atrophy (Wan et al., 2020). TNF-α is also reported to promote the expression of myostatin using the NF-κB pathway and, thereby, the production of IL-6 via p38MAPK that causes growth inhibition of skeletal muscle (Ji et al., 2022). Therefore, activation of NF-κB is crucial to mediate the entire process of muscle atrophy.

ROS-oxidized mitochondrial DNA plays a vital role in activating NLRP3 inflammasome, an oligomeric molecular complex that prompts the maturation of IL-1β, IL-18 and apoptosis. Moreover, ROS also dissociates thioredoxin-interacting protein from thioredoxin, which binds with NLRP3 and further activates the NLRP3 inflammasome (Zheng et al., 2020). Also, TNF-α was reported to induce the expression of IL-1 and IL-1β in muscles while reducing the sphingosine-1-phosphate (SIP) level in the denervated soleus muscles of rats. SIP maintains the integrity of myotubes in TNF-α accompanied L6 myotubes, hence tending to inhibit proteolysis. IL-1α and IL-1β direct the catabolism of myofibrillar proteins in skeletal muscles by increasing the IL-6 expression in cultured myotubes through MAPK and NF-κB signaling (Bernacchioni et al., 2021; Formigli et al., 2004). It has been reported that 0.1–2 ng/ml of IL-1β increases the DNA binding activity of NF-κB and ∼25-fold of IL-6 in myotubes (Luo et al., 2003). Also, IL-1 led to anorexia to increase the circulation of IGFBP-1, resulting in the decreased bioavailability of IGF1 (Clemmons, 2018), but the facts are still unexplored in denervation muscle loss. Similarly, IL-10, an anti-inflammatory cytokine, has been reported to inhibit the toll-like receptors (TLRs) mediated inflammation (Iyer and Cheng, 2012), but its expression remains under-explored in sciatic damage. In contrast, a few studies also showed that TNF-α co-stimulates IL-6 and IL-12 to regulate muscle regeneration (Collins and Grounds, 2001). However, no such experimental evidence is available in denervation-induced inflammation.

IFN-γ, another inflammatory marker, adapts different catabolic pathways to decrease the muscle protein pool. Unlike TNF-α, interferon-gamma (IFN-γ) inhibits the Akt signaling of myogenesis instead of activation of MuRF-1 and Atrogin-1 (Shum and Polly, 2012). A study also found that IFN-γ inhibits the TNF-α via its receptor TNFR2 (Horie et al., 1999), to work as an antagonist of TNF-α but data for inhibition of TNF-α via IFN-γ is limited. Therefore, further experimental evidence is much needed to justify this inhibition. TGF-β also increases the inflammatory responses to promote the proteolysis of ECM in the SMAD3-dependent pathway (Tsai et al., 2009). However, IFN-γ attenuates the expression of TGF-β in muscles (Vu et al., 2019). Hence, more studies are needed to conclude that TGF-β triggers the inflammation in skeletal muscle and the expression of IFN-γ. Similarly, overexpression of tumor necrosis factor (TNF)-like weak inducer of apoptosis.

(TWEAK) also exacerbates skeletal muscle atrophy during sciatic damage. A study reported that 3–7 days of denervation led to a 6-fold increase in the mRNA content of TWEAK in skeletal muscles. Elevated TWEAK induces the transcript of NF-κB further to upregulate the MuRF-1 and Atrogin-1-mediated ubiquitination of myosin heavy chain (MyHC). Increased expression of TWEAK also increases the accumulation of collagen fibers in soleus muscles and reduces the ∼33% cross section area (CSA), leading to ∼7.2% slow fiber and ∼36% reduction in fast fibers. The fact is further confirmed by the increased transcript of macrophage-1 antigen (Mac-1) and cluster of differentiation 68 (CD68) via increased infiltration of macrophages in the injured muscle (Mittal et al., 2010). Moreover, NF-κB (p65) activation is further supported by tumor necrosis factor receptor associated factor 6 (TRAF6) after 14 days of sciatic damage. Upregulated TRAF6 increased the transcript of MuRF-1 and Atrogin-1 in denervated TA, soleus, and gastrocnemius muscles through JNK and p38MAPK-mediated signaling. The study also reported that TRAF6 is necessary for TWEAK-dependent activation of NF-κB (Mittal et al., 2010). Therefore, inhibition of TRAF6 might help to suppress the MuRF-1 and Atrogin-1-dependent ubiquitination of muscle proteins.

Moreover, micro RNAs are also being investigated and reported to have a major role in skeletal muscle atrophy. Recently, it has been reported that miR-142a-5p is an essential regulator of denervation-induced SkM atrophy, which induces mitophagy and apoptosis through mitofusin-1 (MFN1) (Yang et al., 2020). However, it was evident that TNF-α, IL-1, and TWEAK are the major inflammatory cytokines released during sciatic damage. At the same time, other inflammatory markers, including IL-17, IL-4, and IL-10, are least explored during nerve injury (Sharma and Dabur, 2018). Also, IFN-γ plays a dual role in the expression of inflammatory cytokines (Vu et al., 2019). Prolonged and over-activated inflammation promotes downstream muscle atrophy signaling, increases proteolysis, decreases synthesis, reduces muscle regeneration, and ultimately leads to SkM atrophy and fibrosis. Overall, oxidative stress and inflammation are major factors for skeletal muscle atrophy. Inhibition of ROS and inflammation can culminate in denervation-induced skeletal muscle atrophy by inhibiting the ubiquitin-proteasome system (UPS), autophagy-lysosome pathway (ALP), and mitophagy (Yadav et al., 2022). Therefore, further investigations could help provide more insights into the release of inflammatory cytokines and chemokines in skeletal muscles.

The role of apoptosis is less clear in the skeletal muscle, being myofibers are multinucleated. Apoptotic signaling is a pivotal cellular process orchestrated in response to diverse stress stimuli, including the depletion of growth factors, DNA damage, and oxidative stress. At the mitochondrial crossroads, atrophic signals compromised membrane potential hindered oxidative phosphorylation (OXPHOS), and diminished ATP synthesis (Fig. 7). In response to oxidative stress generated, mPTP opening triggers the release of pro-apoptotic initiators, including p53 upregulated modulator of apoptosis (PUMA), Bid, etc., followed by executing effectors like Bax and Bak1, leading to the swelling and rupturing of the mitochondrial membrane. Once activated, these effectors counteract the inhibition of BCL2 family proteins (Der Chen et al., 2011). Increased PTP permeability enhances the translocation of Bax/Bak1 oligomer and subsequent opening of voltage-dependent anion channel (VDAC) and adenine nucleotide translocase (ANT1) channels of mitochondria. Moreover, increased splitting of the mitochondrial membrane excites the cytochrome c and Smac release for apoptosis-inducing factor 1 mitochondria (AIFM1), endonuclease G (Endo G), and caspase activation to release mtDNA (Redza-Dutordoir and Averill-Bates, 2016). Moreover, ligands such as tumor necrosis factor receptors (TNFR) and first apoptosis signal ligand (FasL) also trigger the activation of cysteine (caspase 8) and aspartate proteases (caspase 10) cascades in muscles. Hence, it decreases DNA replication, repair and disrupts the cytoskeleton into apoptotic bodies, causing cell death. Additionally, ER stress also stimulates apoptotic signaling via increased release of Ca2+ to activate procaspase-12 (Savitskaya and Onishchenko, 2015).

Neuromuscular degeneration reduces the count of myonuclei in muscle, leading to more programmed destruction and reduction in the transcription of the hypertrophic gene (Rudolf et al., 2016). It has been reported that 14 days of sciatic resection stimulate caspase-3 and 8 through increased DNA fragmentation and Bax expression in soleus and gastrocnemius muscles in young and old rodents, contributing to a 60% loss of muscle fiber. However, caspase-3 has been implicated in non-apoptotic functions within skeletal muscle, particularly in protein degradation. Notably, it has been linked to the breakdown of actin filaments, contributing to muscle weakness (Adhihetty et al., 2007). It has been reported that nerve injury increases the permeability of Bax protein for simultaneous interaction of cytochrome c, apoptotic protease activating factor-1 (APAF-1), and procaspases 9 to form apoptosome complex for activation of poly (ADP-ribose) polymerase (PARP), caspase 9, and caspase 3 (Plant et al., 2009). The fact is supported by increased expression of apoptosis inducing factor (AIF) after 7 days of denervation, contributing to a 40% increase in DNA fragmentations in denervated rodents. In contrast, activated cytochrome c was reported to inhibit 60% of transcription factor A, mitochondrial (TFAM) and 70% of PGC-α in skeletal muscles. Similarly, opening of mitochondrial PTP increases the 88% and 115% Bax expression in gastrocnemius muscle after 21 and 42 days of denervation, followed by downregulation of Bcl2 up to 79% and 89%, therefore leading to a 16-fold increase in Bax/Bcl-2 ratio and paralleled the FoxO signaling activation for mitochondrial remodeling (Adhihetty et al., 2007). Moreover, nerve injury inhibited the neuronal expression of TIM23 and TOM20, promoting cell death due to enhanced translocation of Bax/Bcl2 ratio and Parkin (Franco-Iborra et al., 2018). However, expression of translocase of the inner membrane of mitochondria 23 (TIM23) and translocase of outer mitochondrial membrane 20 (TOMM20) are still underscored in SkM; therefore, more studies are needed that could elucidate them as a therapeutic target for mitochondrial biogenesis.

A recent study has unveiled a significant role for heat shock proteins (Hsps), particularly hsp 72, in the intricate regulation of apoptosis within skeletal muscles. Notably, studies have highlighted that the overexpression of HSP72 exerts a potent influence on the modulation of NF-κB translocation, primarily through ameliorating IkBα expression in a Sirt1-AMPK-dependent manner (Kuppuswami and Senthilkumar, 2023). In support, certain glucagon-like peptide 1 (GLP-1) agonists have demonstrated the ability to regulate AMPK signaling, thereby contributing to an enhanced expression of HSP72 (Nguyen et al., 2020). Further, inhibition of NF-κB signaling was also achieved by reducing the activity of cellular inhibitor of apoptosis protein 1 (Ciap1) in the soleus muscle. Elevated Ciap1 was reported to ubiquitinate the TRAF6 for NF-κB, and TNF-α scaffolding, to increase expression of MuRF-1 and Atrogin-1 in SkM (Lala-Tabbert et al., 2019). Moreover, few studies reported the regulation of apoptotic signaling via Id expression in aged SkM. However, their expression in denervated SkM remains intact to date. In response to denervation, nuclei of satellite cells are reported to decrease by 20% after 20–30 weeks while found to completely diminish in 10 weeks of neonatal due to simultaneous expression of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) (Castro Rodrigues and Schmalbruch, 1995). The fact is further supported by increased expression of oxidative markers, i.e., nitro-tyrosine and malondialdehyde (MDA)/4-hydroxynonenal (4-HAE) in denervated muscles (Sharma et al., 2020). Hence, the studies indicated that denervation-induced oxidative stress is directly linked with apoptosis.

The AMPK and mTORC1 signaling pathways are pivotal in cell survival, proliferation, and metabolism. AMPK monitors the level of ATP inside cells and the balance between catabolism and anabolism (Fig. 7). The high ratio of AMP/ATP and levels of ROS activates AMPK, which increases ATP production through the promotion of catabolism and a decrease in anabolism (Chun and Kim, 2021). AMPK relocates FoxO3a to the nucleus to stimulate autophagy-related gene expressions, such as LC3β-II, Gabarapl1, and Beclin 1. Additionally, AMPK phosphorylates unc-51 like autophagy activating kinase 1 (ULK1) at Ser 317 and Ser 777 to release ULK1 from the complex composed of ULK1, AMPK, mTORC1, focal adhesion kinase family interacting protein of 200 kD (FIP200), and autophagy-related gene 13 (Atg 13) to initiate autophagy for energy and nutrition (Sanchez et al., 2012). The mTOR can be divided into two groups: mTORC1 and mTORC2. The mTORC1 is sensitive to rapamycin and consists of raptor, mTOR associated protein, LST8 homolog (MLST8), proline-rich Akt substrate of 40 kDa (PRAS40), and DEP-domain containing mTOR-interacting protein (DEPTOR), while mTORC2 includes rictor and mSIN1, and shares DEPTOR and MLST8 with mTORC1. Sustained activation of mTORC1 maintains LC-3α and LC-3β levels and blocks autophagy (Takahara et al., 2020). Even though activation of FoxO3 cannot induce autophagy, which is blocked by mTORC1-mediated inhibition of ULK1 through its phosphorylation at Ser 757 to prevent the interaction between ULK1 and AMPK. Autophagy, on the contrary, can be triggered through mTORC1 activation after the separation of the raptor, even after inhibition of FoxO-dependent transcription, which transcribes autophagy genes LC3, BECLIN1, BNIP3, VPS34, ATG4, ATG8B, ATG12, ULK1, and ULK2 in skeletal muscle (Zou et al., 2022). Hence, mTORC1 is one of the upstream regulators of autophagy in skeletal muscle.

Also, AMPK phosphorylates and inhibits mTORC1, which disassembles FIP200/RB/CCI, Atg 13, Atg 101, and Ulk1/2 complex and lets Ulk1 form a complex with Atg 1, which initiates autophagy. Ulk1/2 is the only core protein that possesses the serine/threonine kinase activity and is involved in initiating the autophagy signaling pathway (Ravikumar et al., 2010). Subsequently, the nucleation stage requires activation of PI3KC3–C1 complex containing PIK3C3, Beclin-1, PIK3R4, and Atg14/Atg14 L also gets initiated after inactivation of mTORC1. Several studies have shown that Atg14 or UVRAG-PI3KC3–C2 containing complexes PI3KC3–C1 are involved in autophagolysosomal maturation. Phagophore, a double membrane structure, starts to enclose cytoplasmic material (Yu et al., 2015). Two proteins, ATG12 and microtubule-associated proteins 1A/1 B light chain 3 A (MAP1LC3A)/MAP1LC3B and LC3, are essential for the growth and expansion of phagocytic membranes. ATG7 and ATG10 help conjugate ATG12 to ATG5, which interacts with ATG16. Parallelly, LC3 is cleaved by ATG4 to produce cytoplasmic LC3-I linked with phosphatidylthanolamine (PE) in a reaction similar to ubiquitin that needs ATG7 and ATG3 to form LC3-β. LC3-β, in lipid form, is incorporated into the autophagosomal membrane and is considered an autophagosomal marker. Phagophore closure leads to the sequestration of cytoplasmic components and the formation of autophagosomes. The autophagosome then fuses with endosomes and vacuoles or lysosomes to form autophagolysosomes. These autophagolysosomes degrade the inner membrane and its contents and are sent back to the cytoplasm for reuse in cellular metabolism, providing energy from catabolism (Xia et al., 2021).

Autophagy, as a process, is not random but rather selective in nature, aiming to remove specific cargoes. Mitophagy and mitochondrial biogenesis are essential to maintain the proper functioning of mitochondrial complexes and cellular respiration. However, mitophagy plays a crucial role in maintaining the quality and turnover of mitochondria under normal circumstances. But, in the event of denervation, the disturbance of Ca2+ balance results in prolonged oxidative stress, causing initial damage to the mitochondria. The presence of Bcl-2/E1B 19 kDa-interacting protein 3-like protein (BNIP3L) is regarded as an indicator of mitochondrial malfunctioning, which in turn prompts mitophagy. Various types of stress, such as oxidative stress and hypoxia, may stimulate the transcription of BNIP3L. There are controversies that BNIP3L induces transmembrane potential loss and thus initiates mitophagy (Saito and Sadoshima, 2015). However, concerning denervation, the loss of transmembrane potential is not necessary by BNIP3L for mitophagy, as it has already been lost due to denervation. BNIP3L disrupts the complex of BCL2-beclin1 (BECN1), causing the release of BECN1, thereby promoting the formation of autophagosomes. BNIP3L also regulates the activation of autophagy, possibly depending on the quantity of BECN1 or mTORC1. The LC3-interacting region (LIR) domain of BNIP3L interacts with LC3s found in the autophagosome membrane (Wang et al., 2019). Hence, BNIP3L interacts with the Atg 8 family proteins to recruit autophagosomes to specific mitochondria.

Some studies also reported that denervation-induced autophagy causes muscle loss at a slower rate compared to other conditions like fasting. Increased expression of autophagic flux was stimulated by upregulation of runt-related transcription factor 1/2 (Runx1/2), a transcription factor highly expressed after nerve injury. Certain studies demonstrated that Runx1 deficient muscle accompanied more myofibril disorganization and expression of atrophying genes in muscles (Wang et al., 2005). Therefore, further studies are warranted to reveal the effect of Runx1 on skeletal muscles.

It is believed that the ubiquitous protein-proteasome system (UPS) degrades the contractile skeletal muscle's proteins and maintains muscle homeostasis and health. While the UPS is crucial in facilitating the breakdown of proteins during muscle atrophy, its deficiency detrimentally affects muscle homeostasis and gives rise to various pathological phenotypes (Kitajima et al., 2020). Denervation-induced prolonged depolarization leads to disrupted calcium balance, causing oxidative stress that activates and inhibits various signaling pathways associated with the regulation of apoptosis, autophagy, and UPS (Sharma et al., 2020). Oxidative stress induces Nrf1 and AMPK, the master regulator of proteasome gene expression, and inhibits the anabolic pathway involving insulin, IGF-I, and Akt. These signaling pathways are vital in determining muscle mass as they regulate ubiquitin ligases, protein synthesis, and myogenesis through their downstream mediators FoxO, mTOR, and NF-κB (Kjøbsted et al., 2018).

FoxO, NF-κB, and Nrf1 commence the process of transcribing E3 ligases, such as MuRF-1, Atrogin-1, (Fig. 7) and NEDD4, which produce ubiquitin chains comprising a minimum of four ubiquitin moieties on substrate proteins through a complex thioester cascade, frequently connected at lysine 48, in order to be recognized by the proteasome and degradation (Bodine and Baehr, 2014). However, deubiquitinating enzymes (DUBs) counteract the ubiquitinylation of proteins and their degradation (Reyes-Turcu et al., 2009). However, prolonged oxidative stress and inflammation fuelled UPS and muscle tissue. Among the E3 ligases, Atrogin-1 and MuRF-1 (Fig. 7) exhibit a marked affinity for targeting various proteins ranging from structural and contractile proteins to those associated with energy metabolism and transcription factors for UPS-mediated degradation (Yadav et al., 2022). Moreover, early studies by Solomon and Goldberg highlighted the proteasome's efficiency in degrading individual myosin, actin, troponin, and tropomyosin components, yet it couldn't disassemble actomyosin complexes or intact myofibrils. This disassembly is facilitated by proteases such as calpains and caspases, which expedite myofibril breakdown (Solomon and Goldberg, 1996).

Interestingly, Foxo-mediated upregulation of specific of muscle atrophy and regulated by transcription (SMART) substantially worsens myofiber atrophy post-denervation, following a Smad-dependent mechanism by collateral activation of UPS and autophagy (Milan et al., 2015). In addition, muscle injury-specific elevation of oxidative stress and inflammation in the muscle amplifies the expression of MuRF-1, Atrogin-1, and mitochondrial E3 ubiquitin protein ligase 1 (Mul1) through TRAF6-driven FoxO3 and NF-κB transcription factors. The Mul1 triggers the ubiquitin-mediated degradation of mitochondrial mitofusion 2, fostering mitophagy (Singh et al., 2021). It is evident that MuRF-1 and Atrogin-1 are critical players in the degradation of muscle atrophy, but the expression of other subsidiary ligases like functional role of F-box and leucine-rich protein 22 (Fbxl22) further amplified their expression. Upregulation of Fbxl22 coincides with elevated levels of p62, a marker of autophagy, and a concomitant reduction in essential proteins like α-actinin, vimentin, and dystrophin. These changes, orchestrated by Fbxl22, appear to set the stage for the early initiation of muscle atrophy. Notably, Fbxl22's involvement is not limited to this preparatory phase alone. Its influence extends further as it contributes to the subsequent upregulation of MuRF-1 expression (Hughes et al., 2020). Similarly, FoxO-RANK-induced expression of Znf216 has limited capacity to mediate ubiquitination. However, specific shuttle proteins, such as Dsk2p and Rad23p, augment its efficacy. Despite this, no experimental evidence exists in the literature to establish a direct link between Znf216 and these shuttle proteins. UPS-mediated degradation is the final stage in protein catabolism activated by diverse upstream regulators (Hishiya et al., 2006). Therefore, control over these regulatory factors could offer a potential avenue to mitigate the ubiquitination of muscle proteins.

Peripheral nerve injury leads to muscle atrophy and dysfunction in denervated muscles. A previous study identified four stages of muscle atrophy: oxidative stress, inflammation, mitophagy, and atrophic fibrosis. Early targeting of inflammation and oxidative stress can delay muscle atrophy progression. Denervation causes an extensive depolarization of the sarcolemma that disrupts the delicate balance of calcium within the cell. This disruption has even extended to the mitochondria, imposing a hefty oxidative burden upon the myofibers. Increased cytoplasmic and mitochondrial calcium levels activate calcium-dependent enzymes like calpains and caspases, which cause proteolysis and induce apoptosis (Sharma et al., 2020). Further, the extended period of oxidative stress triggers the activation of AMPK and NRF1, thereby suppressing mTORC1 and stimulating FoxO3, respectively, subsequently leading to an elevation in the transcription of autophagy and UPS-associated genes. Oxidative stress concurrently stimulates MAPKs and inhibits Akt, which activates downstream transcription factors NF-κB and FoxO, further to boost the inflammation and expression of autophagy and UPS-associated genes (Thomson, 2018). Additionally, Foxo-mediated upregulation of SMART substantially worsens myofiber atrophy, following a Smad-dependent mechanism by collateral activation of UPS and autophagy (Milan et al., 2015).

Moreover, the musculoskeletal unit is comprised of the combined functioning of muscles and bones. When there is damage to the skeletal muscle, both functional and structural abnormalities in the bones can occur, and vice versa. Transforming growth factor beta type 1 (TGF-1β) is released during bone remodeling and resorption, which can lead to the development of muscle fibrosis and atrophy (Fig. 8). It is essential to consider that denervation not only causes significant damage to the muscles but also to the bones. Therefore, the large amounts of TGF-1β released during bone regeneration and remodeling could hinder muscle recovery following nerve injury (Mendias et al., 2012).

It is widely recognized that there is a concomitant initiation of compensatory processes in muscle after nerve damage in addition to the deterioration processes. Following denervation, the degradation of muscle collagen is impeded, resulting in the buildup of collagen. The activation of satellite cells (SCs) during extended periods of denervation upholds the integrity of muscle fibers. As a consequence of these processes, even after an extensive period of denervation, atrophic skeletal muscle fibers are able to endure (Agüera et al., 2019). Further, increased expression of Nrf2 and antioxidant system also help the atrophied muscle to endure a small extent (Bellezza et al., 2018). Strategies to reduce the atrophy of denervated muscle and enhance the compensatory activation of safeguarding mechanisms can yield advantageous outcomes in muscle recovery. Although our understanding of the multifaceted regulatory processes governing SkM atrophy after denervation is extensive, optimized therapeutic interventions must be explored to prevent post-denervation skeletal muscle atrophy to improve the quality of life of affected individuals.

Maintaining muscle homeostasis is essential for overall well-being and health. SkM atrophy, a condition characterized by the loss of muscle mass, not only diminishes individuals' quality of life but also increases morbidity and mortality rates, imposing substantial economic burdens on societies. Despite its profound impact, effective treatments for SkM atrophy remain elusive despite the extensive understanding of its mechanisms. The current strategies and treatments for SkM atrophy encompass a range of approaches. These include drug interventions, such as active substances derived from traditional herbal medicine, chemical drugs, antioxidants, enzyme inhibitors, and hormone drugs. Gene therapy, stem cell therapy, and exosome therapy have emerged as potential treatment modalities. Stem cells derived from skeletal muscle, non-myogenic stem cells, and exosomes are among the avenues explored in this area. Cytokine therapy has also shown promise in addressing muscle atrophy. Physical therapy options include electroacupuncture, electrical stimulation, optogenetic technology, heat therapy, and low-level laser therapy. Through protein, essential amino acids, creatine, β-hydroxy-β-methylbutyrate, and vitamin D, nutrition support is another approach to managing SkM atrophy. Finally, additional therapies encompass biomaterial adjuvant therapy, intestinal microbial regulation, and oxygen supplementation (Yadav et al., 2022).

Exercise therapy has proven to be the most effective intervention for mitigating SkM atrophy. However, its applicability is limited to specific patient populations, such as those with fractures or nerve damage, who may be unable to engage in physical activity. While nutrition support in managing muscle atrophy, cachexia, and sarcopenia is widely recognized, there is a need for more robust, randomized, and long-term clinical trials that are essential to validate the effectiveness of nutritional interventions on muscle metabolism and clinical outcomes. Also, electromechanical stimulation of the muscle lacking neural innervation exhibits favorable outcomes in mitigating the adverse effects of denervation-induced muscular wasting. Although it effectively preserves the dimensions of muscle fibers, it does not enhance the process of neural re-establishment after extended periods of denervation (Huang et al., 2023).

Currently, the pharmacological agents frequently employed for the management of skeletal muscle atrophy encompass bioactive components of conventional herbal medicine, pharmacological compounds, antioxidative agents, hormone medications, enzymes, or inhibitors of enzymes. Synthetic drugs such as torbafylline, formoterol, clenbuterol, megestrol acetate, and similar substances are employed in the treatment of muscle atrophy, with a specific focus on mitigating the effects of inflammation and oxidative stress (Table 1). However, the intervention with these pharmaceutical substances encounters variations among patients, the possibility of adverse drug reactions, and the necessity for continuous surveillance, thereby rendering it incapable of achieving a satisfactory clinical outcome regarding muscular performance, bulk, and general state of health (Yadav et al., 2022). For example, testosterone treatment amplifies the synthesis of muscle protein, and its influence on muscle tissue is regulated by nutrition and physical exertion. Numerous investigations have exhibited the advantageous consequences of testosterone supplementation on sarcopenia attributes, such as reductions in muscle volume and grip potency. Although the administration of testosterone and its derivatives has the potential to stimulate muscular hypertrophy and elevate muscular potency, its clinical ap