Proteins are synthesized from messenger RNA (mRNA) which carries the nucleic acid code for the protein’s amino acid sequence, a process called protein translation. Each protein is encoded by a unique mRNA that is transcribed from the cell’s genomic DNA. Translation of a protein begins with the formation of the ribosomal complex on the mRNA. This ribosomal complex is composed of ribosomal RNAs (rRNAs), ribosome proteins, initiation factors, and transfer RNAs and it binds to a methionine codon at the mRNA’s initiation site. Additional amino acids are added to the initiator methionine to produce a growing peptide. The processive translation process continues down the mRNA until the ribosomal complex encounters a termination codon on the mRNA and the fully formed nascent peptide and the ribosomal complex dissociate.

A cell’s capacity to synthesize proteins is determined by multiple factors. Formation of the ribosomal complex is a tightly regulated process. Epigenetic and transcriptional mechanisms modulate the levels of ribosomal proteins and rRNAs. Signals internal and external to the cell also influence the rate of protein translation. The mechanistic target of rapamycin protein, mTORC1, is a key integrator of these signals. mTORC1 is a kinase that phosphorylates two key translational proteins—ribosomal protein S6 kinase-1 and eukaryotic initiation factor 4E binding protein (eIF4E-BP). eIF4E is normally inhibited by eIF4E-BP. Phosphorylation of eIF4E-BP allows eIF4E to join the ribosome-mRNA preinitiation complex. S6 facilitates the initiation and elongation steps of protein translation and phosphorylation enhances its activities.

Leucine and insulin/insulin-like growth factor 1 (IGF-1) are key modulators that positively influence protein synthesis through mTORC1, albeit by different signaling pathways (Fig. 1). Leucine activates mTORC1 via Rheb signaling whereas insulin and IGF-1 activate mTORC1 via Akt-induced phosphorylation. In CKD, the circulating and intracellular levels of leucine in skeletal muscle are reduced and likely contribute to a lower rate of translation [11,17]. Insulin and IGF-1 signaling through Akt in muscle is also impaired [18]. In theory, the resulting anabolic resistant state should reduce mTORC1 signaling and protein synthesis. However, measuring the activation state of mTORC1 in skeletal muscles is difficult and few, if any, attempts have been made to do so in patients or animals with CKD.

A recent 2020 paper by Zhang et al. [19] suggests the assumption that anabolic resistance is a major cause of reduced protein synthesis in muscle during CKD may be an oversimplification and that multidimensional causes are responsible. Zhang et al. [19] found that nucleolar protein 66 (NO66) was increased in muscle biopsies from dialysis patients and in muscles of mice with CKD. This was a notable finding because NO66 is a chromatin-modifying demethylase that modulates histone methylation, a modification associated with gene expression. Knockout of NO66 in the skeletal muscles of mice with CKD resulted in a higher rate of protein synthesis than in normal mice with CKD. NO66 was shown to suppress ribosomal biogenesis by a mechanism that requires demethylation activity. Inflammatory cytokines like tumor necrosis factor (TNF) and interleukin 6 (IL-6) were responsible for increased expression of NO66 in skeletal muscle via activation of nuclear factor kappa B (NF-κB). Thus, protein synthesis is likely to be negatively impacted by both a reduction in ribosomal function as well as impaired anabolic signaling in skeletal muscle during CKD (Fig. 1).

Cellular and contractile proteins in skeletal muscles have different turnover rates, depending on function and structure. Myofibrillar proteins have long half-lives whereas signaling and other metabolic enzymes turn over much more rapidly. A variety of proteases and several proteolytic systems are responsible for degrading cellular proteins in skeletal muscles. For this review, only the proteolytic systems that are known to be upregulated during CKD will be discussed (Fig. 2).

Muscles are constantly in use throughout the day. This leads to damage to the myofibers that in healthy individuals get repaired through regeneration (Fig. 1). This repair process involves macrophage-dependent removal of the damaged portions of the myofibrils and repair of the site [34]. The regenerative process is sensitive to IGF-1 and occurs in sequential steps. First, localized muscle stem cells (MSCs), which are also called satellite cells and located under the basal lamina of the muscles, become activated. The activation signals are mediators produced by local macrophages and endothelial and immune cells within the muscle fibers at the site. The activated MSCs then migrate to the repair site in the myofiber where they proliferate, differentiate into myotubes, and fuse with other myotubes and myofibers. In addition, multinucleated myotubes can fuse together to produce new fibers. In mice with CKD, satellite cell function and muscle regeneration are defective [19,35]. Markers of satellite cell activation (i.e., MyoD, myogenin), proliferation (i.e., bromodeoxyuridine incorporation), and differentiation (i.e., embryonic myosin heavy chain) were reduced. At least one signal for this regenerative dysfunction would seem to be impaired IGF-1 signaling. Deletion of the IGF-1 receptor from satellite cells recapitulated the CKD-related defects in their proliferation, differentiation, and fusion [35].

Satellite cell senescence was recently identified as a contributory process to CKD-induced dysfunctional muscle regeneration, function, and muscle atrophy [36]. Senescence refers to as an irreversible state in which cells no longer divide and proliferate. When satellite cells become senescent, they can no longer participate in the muscle regeneration process. Despite their inability to divide and proliferate, senescent cells retain a capacity to produce and secrete various growth factors, proinflammatory cytokines, and chemokines which can negatively impact other nonsenescent cells. This cellular capacity is referred to as a senescence-associated secretory phenotype or SASP. Senescent cells are frequently also resistant to apoptosis. Senescence can be induced by one of two conditions; telomere erosion such as is associated with aging or chronic cellular stresses such as DNA damage or oxidative stress. Stress-induced premature senescence can result from activation of either the p21 or the p16^INK4a signaling pathways (Fig. 3). Both pathways inhibit the cyclin-dependent kinase which regulates cell cycle progression.

Muscle content of three senescence markers—senescence-associated β-galactosidase (SA-β-gal), p21, and p16^INK4a—was increased in muscles of mice with CKD and was negatively correlated with weights of the gastrocnemius muscles [36]. Administration of a senolytics cocktail of dasatinib plus quercetin, compounds that induce apoptosis in senescent cells but not in proliferating or quiescent cells, led to a decline in the amounts of SA-β-gal, p21, and p16^INK4a. The levels of proinflammatory cytokines, such as TNF-α and IL-6, in the muscle of CKD mice were reduced by the treatment. Markers of protein degradation (i.e., MuRF1 and atrogin-1) also declined whereas muscle function, as measured as grip strength, and average muscle fiber size increased. Notably, addition of uremic mouse serum to cultured satellite cells increased the number of p21⁺ cells. Co-addition of the senolytics cocktail plus uremic serum to satellite cells decreased the number of p21⁺ cells by increasing their apoptosis. Overexpression of constitutively activated FOXO1 in satellite cells also increased the number of p21⁺ cells but the action persisted despite addition of senolytics. These findings with p21 indicate that it is activated in satellite cells when FOXO activity is increased.

In summary, CKD impairs the ability of muscle to repair itself. Multiple factors, including an inefficient regenerative process and an increase in satellite cell senescence, contribute to the dysfunction. Regenerative medicine and therapies with senolytics are important continuing areas of investigation in muscle biology and CKD.

Insulin and IGF-1 are critical anabolic hormones that support the health of skeletal muscle. Activation of their receptors triggers cell signaling through insulin receptor substrate-1 (IRS-1) which docks with the receptors. IRS-1 also interacts and activates phosphatidyl-3 phosphate kinase (PI3K), which in turn, phosphorylates and activates Akt (Fig. 4). Akt serves as a signaling nexus. It activates mTOR which positively regulates protein synthesis as described earlier. Akt also acts as a suppressor of protein degradation by negatively regulating the FOXO transcription factors that control atrogene expression.

Studies involving patients and animals with CKD have repeatedly found that insulin/IGF-1 signaling is impaired. The conditions underlying these defects are numerous and include dialysis, inflammation, metabolic acidosis, increased glucocorticoid production, and others [37]. Bailey et al. [18] found that IRS-1-associated PI3K kinase activity in muscle was decreased by CKD. Later studies by Thomas et al. [38] identified a membrane-associated glycoprotein, signal regulatory protein-α (SIRP-α), that is upregulated in muscle during CKD. SIRP-α interacts with the insulin/IGF-1 receptors and IRS-1. Overexpression of SIRP-α in cultured muscle cells suppressed insulin signaling whereas knockdown of SIRP-α in muscle cells overcame a cytokine-induced suppression of insulin signaling and increase in protein degradation. Other investigators have reported that phosphatase and tensin homolog (PTEN), a phosphatase that inactivates Akt, was increased in the muscles of mice with CKD [39]. When PTEN was knocked out specifically in skeletal muscle, the CKD-induced activation of protein degradation was dampened [40]. To summarize, CKD impairs insulin/IGF-1 anabolic signaling in muscle. Evidence indicates that multiple mechanisms are responsible for the aberrant insulin/IGF-1 signaling that increases FOXO-mediated atrogene responses such as MuRF1, atrogin-1, and ATG proteins and ultimately, accelerates protein degradation.

Normal daily metabolism generates acids that are excreted by healthy kidneys in the urine. In CKD, this urinary excretion of acids is reduced, resulting in an accumulation of acids in the blood, a condition called metabolic acidosis. Studies involving patients or animals with CKD have consistently shown how metabolic acidosis is linked to the loss of muscle mass and how correction of CKD-related metabolic acidosis improves muscle protein turnover as well as other measures of nutritional status [8,11,15,41–43]. In rats with ammonium chloride-induced acidosis, physiological levels of glucocorticoids were necessary for accelerating protein degradation; however, the same levels of steroids were insufficient to elevate protein degradation in adrenalectomized nonacidotic rats [43]. Similarly, in a study of patients with CKD, arterial [HCO₃] was inversely correlated with the rate of forearm protein degradation [41]. Interestingly, the subjects’ plasma cortisol levels were in the normal range; however, the cortisol levels were also inversely correlated with their arterial [HCO₃]. These correlations are reminiscent of the synergistic relationship between acidosis and glucocorticoids in animal studies.

How do acidosis and glucocorticoids increase protein degradation in muscle? One obvious way for glucocorticoids to influence protein degradation is through their transcription factor functionality. Indeed, pharmacological doses of glucocorticoids increase the expression of several atrogenes including MuRF1 and myostatin [44,45]. Interestingly, these genes have binding sites for both the GR and the FOXO transcription factors suggesting the transcription factors might act synergistically. Inhibition of either the FOXO proteins or GR is sufficient to attenuate muscle loss in rats and cultured myotubes. Hu et al. [23] also reported that both glucocorticoids and insulin/IGF-1 resistance are necessary to activate protein degradation in skeletal muscles of rodents. They found that GR directly interacts with PI3K to reduce signaling via IRS-1, which results in greater FOXO activity. This nongenomic mechanism of GR is an important aspect of the atrophy process as muscle loss does not occur in adrenalectomized rodents with either metabolic acidosis or insulinopenia [46,47]. Acidosis also seems to exert actions on insulin signaling that are independent of the GR. In dogs, ammonium chloride-induced acidosis caused insulin resistance [48]. In a cultured myotube model, incubation in acidified media (pH 7.1) produced defective IRS-1-associated PI3K and Akt activities [49]. Media acidification also increased the expression of mRNAs several components of the ubiquitin-proteasome system [50]. In summary, considerable evidence indicates that acidosis and glucocorticoids have a synergistic relationship that negatively impacts insulin/IGF-1 signaling and influences atrogene expression via multiple mechanisms.

Inflammation is a frequent phenotypic aspect of CKD that is strongly correlated with increased patient mortality and progression of CKD [51,52]. Acidosis, infections and sepsis, oxidative stress and mitochondrial dysfunction, obesity, premature aging and senescence, and end-stage dialysis are among the many conditions that contribute to a chronic inflammatory state in CKD.

Some inflammation is necessary for healthy muscle repair and regeneration; however, muscle atrophy occurs when levels of inflammatory cytokines are elevated above physiologic levels. Release of low levels of IL-6 and other cytokines from the muscle initiates the recruitment of M1 macrophages to the injury site where they help to clear the damaged tissue [53]. These cells are replaced by M2 macrophages and, when combined with an increase in muscle IGF-1 production, facilitate myogenesis and repair. Mice with CKD that were treated with a single session of acupuncture with electrical stimulation exhibited a post-session increase in IL-6 from muscle, relative to mice with CKD alone. Markers of myogenesis and the rate of protein degradation were also improved significantly after the treatment [54].

In other studies, elevated levels of inflammatory cytokines, especially IL-6 and TNFα, were found to induce insulin resistance, increase atrogene expression, and accelerate the rate of muscle protein degradation in rodents [55–58]. Pharmacologic antagonism of these cytokines reduced both atrogene expression and protein breakdown. In muscle biopsies from late-stage nondiabetic CKD patients, TNF-α was elevated and phosphorylated Akt was decreased, relative to control patients [59]. Similarly, forearm release of IL-6 and plasma IL-6 were elevated in ESRD patients on hemodialysis and their forearm net protein balance was correlated with the release of IL-6 [60]. In summary, evidence from experimental animals as well as CKD and ESRD patients strongly suggest that elevated levels of inflammatory cytokines in the muscle milieu are strongly linked to negative protein turnover.

Myostatin, which is also known as growth/differentiation factor-8 or GDF-8, was discovered in 1997 as a cytokine produced by muscle fibers (i.e., a myokine), that suppresses muscle growth [61,62]. It belongs to the transforming growth factor-β (TGF-β) protein superfamily. Humans and animals with naturally occurring loss of function mutations in the myostatin gene have extensive muscle development. The myokine is produced by myofibers as an inactive proprotein that becomes active when proteolytic cleaved by bone morphogenetic protein-1. When released into the bloodstream, active myostatin can bind to the activin receptors IIa and IIb (ActRIIs) on various cell types.

In patients and animals with CKD, reduced renal clearance and increased myostatin production in muscles result in an elevated circulating level of myostatin [63,64]. When the myokine binds to ActRIIs, the receptors activate downstream cachexia-inducing signaling pathways in skeletal muscle and adipose tissue. In muscle, myostatin exerts at least three seemingly distinct actions. First, activation of the ActRIIs triggers STAT3 signaling which upregulates atrogenes expression, especially atrogin-1 and MuRF1 [64]. Second, myostatin is linked to insulin resistance in muscle and other tissues, although the responsible mechanism(s) in CKD remain unclear [65]. Third, the myokine retards myogenesis via a mechanism involving Pax-7 [66]. Satellite cell activation in adult muscle is also reduced by myostatin via a p21-dependent mechanism [67]. As noted earlier, dysfunctional myogenesis and satellite cell activation (i.e., quiescence) have been noted in CKD, suggesting that myostatin may be one of the mediators responsible for these effects [35,36].

Activin A is another member of the TGF-β protein superfamily that is closely related to myostatin. It is produced by many cells throughout the body, including some kidney cell types. It also binds to ActRIIa and ActIIb receptors and activates the same pathways as myostatin. Recently, increased production of activin A in kidney fibroblast and cells of the juxtaglomerular apparatus in mice with CKD was linked to the development of muscle loss [68]. In the same report, CKD-induced muscle atrophy in multiple mouse models was effectively prevented by antagonizing the actions of activin A by two methods—systemic blockade of ActRIIs via a receptor-ligand trap or downregulation of muscle-specific ActRII signaling using an adeno-associated virus. This study firmly established the existence of organ-to-organ crosstalk between the failing kidneys and skeletal muscle.

Given the strong links between both activin A and myostatin and muscle atrophy during CKD and other wasting conditions, myostatin/activin A signaling is an attractive therapeutic target to prevent muscle loss. In one preclinical study, the administration of an inhibitory myostatin peptibody to mice with CKD improved muscle mass and muscle protein synthesis while reducing muscle protein degradation, atrophy-related signaling, and muscle inflammation [69]. The treatment also attenuated the muscle fibrosis that developed during CKD [70]. Unfortunately, the outcomes of randomized clinical trials testing anti-myostatin therapies have been inconsistent. In the only study involving kidney diseases, an anti-myostatin peptibody (PINTA 745) treatment was stopped early due to a lack of efficacy. In two phase 2 trials of LY2495655, a myostatin-specific monoclonal antibody, inconsistent outcomes were reported. When LY2495655 was given to elderly patients (≥75 years old) after falls, the treatment group (24 weeks) experienced a small significant increase in muscle mass with some limited benefits in gait and stair climbing speeds [71]. In a different trial with arthroplasty patients given LY2495655, small increases in lean body mass were noted during the 12 weeks of treatment; however, the treatment effect was not sustained 12 weeks after treatments ended [72]. In a trial of domagrozumab, a humanized monoclonal antibody to myostatin, ambulatory boys with Duchenne muscular dystrophy were treated for 48 weeks [73]. Small but nonsignificant increases in muscle volume without functional benefit were reported. In a trial of diabetic patients given bimagrumab, a humanized monoclonal antibody that targets the activin type II receptors for 48 weeks, lean mass increased significantly although the study did not include any functional evaluation of muscle mass or strength [74]. In a 24-week trial of bimagrumab in arthroplasty patients, lean body mass increased significantly but no functional benefit was noted in the treated patients [75]. The reasons for the seemingly different outcomes between preclinical studies and patient clinical trials are unclear. There has been speculation that anti-myostatin therapies alone may not be effective since activins would likely escape the inhibition. It is also unclear whether the anti-activin receptor antibodies are equally effective against type IIA and IIB receptors. Nonetheless, this remains a promising area of investigation as the tools to target the myostatin/activin A pathway evolve.

Declining kidney function in CKD results in elevated levels of “toxic” molecules, including urea, creatinine, phosphorous, and organic acids such as indoxyl sulfate (IS) and p-cresyl sulfate. Recent evidence underscores how IS may serve as a signal for muscle wasting. In the intestines, bacteria degrade dietary tryptophan to indole which enters the circulation. In the liver, microsomal enzymes convert indole to IS which accumulates in the body during CKD due to a reduction in its clearance by the kidneys [76]. In several studies of patients with CKD or ESRD, handgrip strength was inversely related to the serum concentrations of IS [77,78].

IS has been linked to the production of reactive oxygen species, oxidative stress, and inflammation in vascular and other tissues [76]. These signals are also linked to muscle atrophy. In studies of mice with adenine-induced CKD, IS accumulated in their skeletal muscles [79]. This accumulation was accompanied by structural abnormalities in the arrangement of skeletal muscle fibers, reduction in muscle fiber size and shifts in metabolites associated with glycolysis, energy production, and antioxidants. Administration of IS to cultured myotubes produced an increase in the production of inflammatory cytokines (i.e., TNF and IL-6) as well as myostatin and atrogin-1 [80]. When IS was added to cultured myoblasts, differentiation was blocked due to suppression of key myogenic regulatory factors such as myogenic regulatory factor 4 (MRF4/Myf6) that regulate key steps in myogenic differentiation [81]. The same study confirmed that MRF4/Myf6 was suppressed in the muscles of mice with CKD. In in vitro studies, adding IS to mitochondria isolated from skeletal muscle resulted in impaired electron transport, reduced energetics, and increased hydrogen peroxide production [82]. Together, these preclinical studies support the hypothesis that IS exerts wasting effects on skeletal muscle by negatively impacting both protein turnover in myofibers and by suppressing satellite cells involved in muscle repair and maintenance.

Efforts to develop compounds that slow progression of CKD by reducing the accumulation of IS and other uremic toxins date back to the previous century. In the late 20th century, an oral activated carbon compound, AST-120, was developed and an abundance of preclinical studies and clinical trials with the compound have been conducted with the compound. Administration of AST-120 to mice with CKD for 20 weeks resulted in reductions in the levels of IS, myostatin, and atrogin-1 in hindlimb skeletal muscles [83]. A corresponding increase in phosphorylated Akt was also noted. Levels of inflammatory cytokines in muscle were unchanged by the treatment, however, muscle weights were improved following AST-120 therapy. In a similar study involving the administration of AST-120 to CKD mice for 20 weeks, the compound reversed the reduction of exercise capacity, as measured by runtime and distance [84]. Notably, several measures of mitochondrial function and biogenesis in muscle were suppressed by CKD and were normalized to the control levels by treatment with AST-120.

A considerable number of clinical trials of AST-120 have been conducted in patients with CKD over the past 2–3 decades with variable outcomes. We will summarize the findings of two meta-analyses that have been conducted within the last 5 years; both reported limited positive overall actions of AST-120 in terms of slowing the progression of CKD and lowering the level of IS. A 2019 review analyzed data from five trials from 1982 to 2013 [85]. In the second review conducted in 2021, the authors analyzed data from 15 randomized clinical trials (3,763 subjects) with a duration of treatment consisting of between 7 days and 37 months [86]. In both reviews, the authors concluded that AST-120 may slow the decline in renal function and delay the onset of dialysis. No assessment of muscle function was included in either study. According to PubMed, only one clinical trial has been conducted that investigated the effects of AST-120 on skeletal muscle health [87]. The 2022 study involved 124 CKD patients who completed the 48-week AST-120 intervention; measures of muscle health included gait speed, hand grip strength, and body composition. No significant improvements were noted in any of the muscle-related outcomes although a trend towards improving sarcopenia was noted. Given the limited positive outcomes that have been reported for AST-120, the overall benefits of the therapy remain controversial [86]. The overall importance of uremic toxins in the pathophysiology of CKD, especially with regard to muscle wasting, remains an unanswered question.

CKD is a devastating and complex condition that impacts many organ systems of the body, including skeletal muscle. Loss of skeletal muscle mass is a frequent pathologic consequence of CKD that is referred to in the literature by various terms, depending on whether and what criteria are applied—muscle atrophy, wasting, sarcopenia, or PEW. The causes of reduced muscle mass and muscle function are multifactorial and stem from many catabolic signals present during CKD, including acidosis, inflammation, and uremic toxins. These extracellular signals alter intracellular signaling pathways that control muscle protein turnover and regeneration. The consequences of these phenotypic and metabolic changes are a state of protein and energy wasting. Therapies developed so far typically address one aspect of this disorder, leaving in place other aspects of the catabolic state. This perhaps explains why treatments have largely been unsuccessful. The most promising approaches to minimize muscle loss will likely need to address multiple aspects of the condition, much as modern multipronged cancer therapies frequently combat multiple critical cell targets.