Blockade of Metallothioneins 1 and 2 Increases Skeletal Muscle Mass and Strength

Abstract

Metallothioneins are proteins that are involved in intracellular zinc storage and transport. Their expression levels have been reported to be elevated in several settings of skeletal muscle atrophy. We therefore investigated the effect of metallothionein blockade on skeletal muscle anabolism in vitro and in vivo. We found that concomitant abrogation of metallothioneins 1 and 2 results in activation of the Akt pathway and increases in myotube size, in type IIb fiber hypertrophy, and ultimately in muscle strength. Importantly, the beneficial effects of metallothionein blockade on muscle mass and function was also observed in the setting of glucocorticoid addition, which is a strong atrophy-inducing stimulus. Given the blockade of atrophy and the preservation of strength in atrophy-inducing settings, these results suggest that blockade of metallothioneins 1 and 2 constitutes a promising approach for the treatment of conditions which result in muscle atrophy.KEYWORDS muscle metabolism S keletal muscle hypertrophy is characterized in the adult mammal by an increase in the size of preexisting myofibers. The induction of hypertrophy involves an activation of the pathways that increase protein synthesis and inhibition of cellular signaling, which induces protein degradation. Hypertrophy can be induced by the activation of Akt, through multiple potential inputs (1). Akt induces hypertrophy in part by activating the mTOR/70S6 kinase pathway. In addition, Akt inhibits protein degradation, by phosphorylating and therefore blocking Foxo1 and Foxo3-transcription factors which are required for the upregulation of the E3 ubiquitin ligases MuRF1 and MAFbx, which help mediate protein turnover during muscle atrophy (2-4). Therefore, activation of Akt constitutes a critical signaling node to increase muscle hypertrophy and block muscle atrophy (1).Mammalian metallothioneins (MTs) belong to a family of cysteine-rich, metalbinding proteins. In rodents, four MT isoforms have been identified: the two major isoforms, MT-1 and MT-2, are ubiquitously expressed, while MT-3 and MT-4 show tissue specific expression in the central nervous system and squamous epithelia, respectively. In humans, multiple isoforms have been reported for MT-1 (MT-1A, MT-1B, MT-1E, MT-1F, MT-1G, MT-1H, MT-1M, and MT-1X), while no splice variants are documented for MT-2, MT-3, or MT-4 (5; for a review, see reference 6).MTs play a role in cellular zinc homeostasis, mitochondrial function (7), defense against oxidative stress (8), and defense against inflammation (5). Moreover, several reports and a recent review highlight a role of metallothioneins in cancer (9), aging (10), and the onset of particular central nervous system diseases (11).

Pharmacological Dual Inhibition of Tumor and Tumor-Induced Functional Limitations in a Transgenic Model of Breast Cancer

“…These include Pgc-1β, which is induced by MyoD and alternate NF-κB, and is a negative regulator of canonical NF-κB signaling in skeletal muscle (25,32), Hoxa9, which is typically elevated in aging satellite cells of muscle and its expression is repressed by NF-κB (33,34), and metallothioneins 1 and 2 (Mt1 and Mt2), which control skeletal muscle mass and strength (35). Pgc-1β and Hoxa9 but Mt1 and Mt2 mRNA levels were lower in the skeletal muscle of tumor-bearing mice, which were reversed upon DMAPT treatment (Fig.…”

Upregulation of ZIP14 and Altered Zinc Homeostasis in Muscles in Pancreatic Cancer Cachexia

“…Thus, Zip14 was the only zinc influx transporter gene that was significantly induced in cachectic muscles from both models (Figure 2A and Figure S2). Metallothioneins (Mt), such as Mt1 and Mt2, are zinc-inducible genes that encode cysteine-rich metal-binding proteins and serve as markers for elevated intracellular zinc ion concentration in cells [24,25]. In line with Zip14 induction and increased zinc levels, cachectic muscles showed significant upregulation of both Mt1 and Mt2 in the Pan02 (Mt1: p = 0.0072; Mt2: p = 0.0016) and FC1242 (Mt1: p = 0.0070; Mt2: p = 0.0295) models compared to their respective controls ( Figure 2C).…”

Inhibition of mTORC1 signaling in aged rats counteracts the decline in muscle mass and reverses multiple parameters of muscle signaling associated with sarcopenia

“…Additionally, expression of the metallothioneins, MT1 and MT2, increases during atrophy and is elevated in sarcopenic muscle (45). Genetic silencing of these genes promotes muscle hypertrophy in vivo (45).…”

“…Additionally, expression of the metallothioneins, MT1 and MT2, increases during atrophy and is elevated in sarcopenic muscle (45). Genetic silencing of these genes promotes muscle hypertrophy in vivo (45). Interestingly, RAD001 treatment suppressed the MT1 gene expression level in old muscles, while levels of MT2 though variable within either treatment group, remained unchanged between groups (Fig.…”

“…In addition to MuRF1, the metallothionein MT1 was downregulated by mTORC1 inhibition. We had previously shown that this is a high-fidelity marker of atrophy, and knocking out the MT genes in mice causes muscle hypertrophy (45). This finding too is consistent with the increase in mass observed in the present study, and provides further mechanistic rationale.…”