Myostatin (MSTN) is a transforming growth factor-ß (TGF-ß) family member that plays a critical role in regulating skeletal muscle mass . Mice engineered to carry a deletion of the Mstn gene have about a doubling of skeletal muscle mass throughout the body as a result of a combination of muscle fiber hyperplasia and hypertrophy . Moreover, loss of myostatin activity resulting either from postnatal inactivation of the Mstn gene ,  or following administration of various myostatin inhibitors to wild type adult mice – can also lead to significant muscle growth. Hence, myostatin appears to play as least two distinct roles, one to regulate the number of muscle fibers that are formed during development and a second to regulate growth of muscle fibers postnatally. The function of myostatin appears to have been conserved across species, as inactivating mutations in the myostatin gene have been demonstrated to cause increased muscling in cattle – , sheep , dogs  and humans . As a result, there has been considerable effort directed at developing strategies to modulate myostatin activity in clinical settings where enhancing muscle growth may be beneficial. In this regard, loss of myostatin activity has been demonstrated to improve muscle mass and function in dystrophic mice – and to have beneficial effects on fat and glucose metabolism in mouse models of obesity and type II diabetes .
Myostatin is synthesized as a precursor protein that undergoes proteolytic processing to generate an N-terminal propeptide and a C-terminal dimer, which is the biologically active species. Following proteolytic processing, the propeptide remains bound to the C-terminal dimer and maintains it in an inactive, latent complex , , , which represents one of the major forms of myostatin that circulates in the blood , . In addition to the propeptide, other binding proteins are capable of regulating myostatin activity in vitro, including follistatin , , FLRG , and Gasp-1 . We previously showed that follistatin can also block myostatin activity in vivo; specifically, we showed that follistatin can ameliorate the cachexia induced by high level expression of myostatin in nude mice  and that transgenic mice expressing follistatin in muscle have dramatic increases in muscle mass . Here, I show that overexpression of follistatin can also cause substantial muscle growth in mice lacking myostatin, demonstrating that other TGF-ß related ligands normally cooperate with myostatin to suppress muscle growth and that the capacity for enhancing muscle growth by targeting this signaling pathway is much larger than previously appreciated.
Increased muscle mass in transgenic mice expressing FLRG
Previous studies have identified several proteins that are normally found in a complex with myostatin in the blood , . One of these is the follistatin related protein, FLRG, which has been demonstrated to be capable of inhibiting myostatin activity in vitro. To determine whether FLRG can also inhibit myostatin activity in vivo, I generated a construct in which the FLRG coding sequence was placed downstream of a myosin light chain promoter/enhancer. From pronuclear injections of this construct, a total of four transgenic mouse lines (Z111A, Z111B, Z116A, and Z116B) were obtained containing independently segregating insertion sites. Each of these four transgenic lines was backcrossed at least 6 times to C57 BL/6 mice prior to analysis in order to control for genetic background effects. Northern analysis revealed that in three of these lines the transgene was expressed in skeletal muscles but not in any of the non-skeletal muscle tissues examined (Figure 1); in the fourth line, Z111B, the expression of the transgene was below the level of detection in these blots. As shown in Table 1, all four lines exhibited significant increases in muscle weights compared to wild type control mice. These increases were observed in all four muscles that were examined as well as in both sexes. Moreover, the rank order of magnitude of these increases correlated with the rank order of expression levels of the transgene; in the highest-expressing line, Z116A, muscle weights were increased by 57–81% in females and 87–116% in males compared to wild type mice. Hence, FLRG is capable of increasing muscle growth in a dose-dependent manner when expressed as a transgene in skeletal muscle.
The research was funded by grants from the NIH and the Muscular Dystrophy Association and by a gift from Merck Research Laboratories.
See http://www.jhu.edu/sejinlee/%20for%20more%20information for more information.
Citation: Lee S-J (2007) Quadrupling Muscle Mass in Mice by Targeting TGF-ß Signaling Pathways. PLoS ONE 2(8): e789. doi:10.1371/journal.pone.0000789
LINK TO THE PUBLISHED ARTICLE http://www.plosone.org/doi/pone.0000789
Source: Nick Zagorski
Johns Hopkins Medical Institutions