Osteocalcin is necessary and sufficient to maintain muscle mass in older mice

Abstract

Objective: A decrease in muscle protein turnover and therefore in muscle mass is a hallmark of aging. Because the circulating levels of the bone-derived hormone osteocalcin decline steeply during aging in mice, monkeys and humans we asked here whether this hormone might regulate muscle mass as mice age.

Methods: We examined muscle mass and strength in mice lacking osteocalcin (Ocn-/-) or its receptor in all cells (Gprc6a-/-) or specifically in myofibers (Gprc6a Mck -/-) as well as in 9 month-old WT mice receiving exogenous osteocalcin for 28 days. We also examined protein synthesis in WT and Gprc6a-/- mouse myotubes treated with osteocalcin.

Results: We show that osteocalcin signaling in myofibers is necessary to maintain muscle mass in older mice in part because it promotes protein synthesis in myotubes without affecting protein breakdown. We further show that treatment with exogenous osteocalcin for 28 days is sufficient to increase muscle mass of 9-month-old WT mice.

Conclusion: This study uncovers that osteocalcin is necessary and sufficient to prevent age-related muscle loss in mice.

Keywords: Aging; Muscle mass; Osteocalcin.

Figure 1

Osteocalcin is necessary to maintain muscle mass in adult mice. (A) Weight of hindlimb muscles and (B) body weight in 6 month-old Ocn−/− female mice and WT littermates. (C) Representative histology and measurement of the cross section area (CSA) of the muscle fibers in 6 month-old Ocn−/− female mice and WT littermates. (D) Muscle strength determined as the maximal grip force in 12 month-old Ocn−/− female mice and WT littermates.

Figure 2

Osteocalcin signaling in myofibers is necessary to maintain muscle mass in older mice. (A) Weight of hindlimb muscles and (B) body weight in 12 month-old Gprc6a−/− female mice and WT littermates. (C) Weight of hindlimb muscles and (D) body weight in 12 month-old Gprc6a_Mck−/− female mice and WT littermates. (E) Weight of hindlimb muscles and (F) body weight in 12 month-old compound mutant mice Ocn+/−;Gprc6a_Mck+/− female mice and control littermates (control group includes WT, Gprc6af/f, Gprc6af/+ and Mck-Cre transgenic mice).

Figure 3

Osteocalcin increases protein synthesis in WT myotubes. (A) Urine levels of 3-methylhistidine (3MH) in Ocn−/− female mice and WT littermates. (B) Protein synthesis in WT mouse myotubes as measured by ³H-Tyrosine incorporation into cellular protein and in (C) Gprc6a−/− myotubes treated for 2 h with different dose of osteocalcin (Ocn). (D) Phosphorylation of S6K1 in WT mouse myotubes treated for 30 min with different dose of Ocn. (E) Phosphorylation of S6K1 in WT and raptor−/− mouse myotubes treated for 30 min with Ocn (10 ng/ml). (F) Phosphorylation of mTOR and S6K1 in WT mouse myotubes treated for 30 min with Ocn (10 ng/ml) in the presence or absence of the mTOR inhibitor Torin 1.

Figure 4

Exogenous osteocalcin is sufficient to increase muscle mass in older mice. (A) Weight of hindlimb muscles and (B) body weight of 10 month-old WT female mice receiving vehicle or osteocalcin (Ocn) for 28 days. (C) Representative histology and measurement of the cross section area (CSA) of the muscle fibers in 10 month-old WT female mice receiving vehicle or Ocn for 28 days. (D) Circulating Ocn levels in 10 month-old WT female mice receiving vehicle or Ocn for 28 days. (E) Muscle strength determined as the maximal grip force in 10 month-old WT female mice receiving vehicle or Ocn for 28 days.