Allen DL, Cleary AS, Speaker KJ, Lindsay SF, Uyenishi J, Reed JM, Madden MC, Mehan RS. Myostatin, activin receptor IIb, and follistatin-like-3 gene expression are altered in adipose tissue and skeletal muscle of obese mice. Am J Physiol Endocrinol Metab 294: E918-E927, 2008. First published March 11, 2008 doi:10.1152/ajpendo.00798.2007 is a secreted growth inhibitor expressed in muscle and adipose. We sought to determine whether expression of MSTN, its receptor activin RIIb (ActRIIb), or its binding protein follistatin-like-3 (FSTL3) are altered in subcutaneous or visceral adipose or in skeletal muscle in response to obesity. MSTN and ActRIIb mRNA levels were low in subcutaneous (SQF) and visceral fat (VF) from wild-type mice but were 50-to 100-fold higher in both SQF and VF from ob/ob compared with wild-type mice. FSTL3 mRNA levels were increased in SQF but decreased in VF in ob/ob compared with wild-type mice. Moreover, MSTN mRNA levels were twofold greater in tibialis anterior (TA) from ob/ob mice, whereas ActRIIb and FSTL3 mRNA levels were unchanged. MSTN mRNA levels were also increased in TA and SQF from mice on a high-fat diet. Injection of ob/ob mice with recombinant leptin caused FSTL3 mRNA levels to decrease in both VF and SQF in ob/ob mice; MSTN and ActRIIb mRNA levels tended to decrease only in VF. Finally, MSTN mRNA levels and promoter activity were low in adipogenic 3T3-L1 cells, but an MSTN promoter-reporter construct was activated in 3T3-L1 cells by cotransfection with the adipogenic transcription factors SREBP-1c, C/EBP␣, and PPAR␥. These results demonstrate that expression of MSTN and its associated binding proteins can be modulated in adipose tissue and skeletal muscle by chronic obesity and suggest that alterations in their expression may contribute to the changes in growth and metabolism of lean and fat tissues occurring during obesity. leptin; 3T3-L1 cells; visceral adipose tissue; subcutaneous adipose tissue FACTORS THAT AFFECT THE GROWTH of skeletal muscle or of adipose tissue can have profound effects on overall health and viability (18). One such factor regulating growth of skeletal muscle and adipose is myostatin (MSTN). MSTN is a member of the activin/transforming growth factor-␤ (TGF␤)/bone morphogenetic protein (BMP) family of secreted signaling factors that binds to the activin receptor type II family members, most notably activin receptor IIb (ActRIIb; Ref. 20), and inhibits skeletal muscle growth through repression of proliferation, differentiation, and protein synthesis (33, 37). Inactivating mutations to the MSTN gene result in a hypermuscular phenotype in mice, cows, and humans (15,25,36), whereas muscle-specific overexpression of MSTN in transgenic mice results in decreased muscle mass (31).In addition, several lines of evidence have suggested a role for MSTN in the regulation of adipose tissue growth in addition to its obvious effects on muscle growth. First, in their initial paper identifying the MSTN gene, McPherron et al. (25) stated that MSTN mRNA could be detected by ...

During food deprivation (FD), skeletal muscle protein is broken down to produce amino acids for hepatic gluconeogenesis to maintain blood glucose levels. However, it is unclear what role, if any, the secreted antigrowth factor myostatin (MSTN) plays in the muscle atrophy induced by FD. We therefore examined expression and function of MSTN in FD in mice. Two days of FD significantly decreased muscle mass and protein content and increased mRNA levels of ubiquitin ligases MuRF-1 and atrogin-1 in fast-twitch tibialis anterior (TA) muscle but not slow-twitch soleus (Sol) muscle, while 2 days of refeeding returned these to fed values in TA. MSTN mRNA levels were significantly increased approximately threefold by 2 days, but not 1 day, of FD and returned to fed levels with 2 days of refeeding in TA but were not significantly affected by FD or refeeding in Sol. TA mass decreased to a similar amount after 1 day of FD in wild-type mice and mice null for the MSTN gene but was decreased to a greater amount in wild-type than MSTN-null mice by 2 days of FD. In addition, blood glucose levels decreased and corticosterone levels increased to a greater extent in MSTN-null mice after 2 days of FD, but surprisingly muscle MuRF-1 and atrogin-1 mRNA levels were not affected by the lack of MSTN during FD. Similarly, changes in hepatic enzyme expression in response to FD were identical between wild-type and MSTN-null mice. Our data are consistent with the hypothesis that MSTN is dispensable for the initial atrophy occurring in response to FD but attenuates the decrease in fast-twitch muscle mass during prolonged FD.

We recently demonstrated that mRNA levels of three members of the CCAAT/enhancer binding factor (C/EBP) family of transcription factors are increased in skeletal muscle following 12 days of spaceflight. In the present study, we further explored the expression of C/EBP-δ in atrophying fast skeletal muscle by examining its expression in muscle from food-deprived (FD) mice, and investigated its role in regulating the expression of the secreted antigrowth factor myostatin. C/EBP-δ mRNA and protein levels were significantly increased by 2 days of food deprivation in the tibialis anterior (TA) muscle, and expression of both myostatin and C/EBP-δ mRNA during food deprivation was attenuated by injection with the glucocorticoid inhibitor RU486. The increase in myostatin mRNA levels with food deprivation appears to be at least partially transcriptionally driven, since levels of myostatin pre-mRNA were significantly increased in the TA muscle. C/EBP-δ mRNA levels and promoter activity were significantly increased by transfection of C(2)C(12) myotubes with a glucocorticoid receptor construct and 24 h of treatment with the synthetic glucocorticoid dexamethasone. Furthermore, activity of the C/EBP-δ promoter was significantly increased with as little as 1 h of dexamethasone treatment, while activity of the mouse myostatin promoter was only significantly increased with longer treatment periods of 24 h or more. Activity of the myostatin promoter-reporter construct was significantly increased in C(2)C(12) myotubes by cotransfection with expression constructs for C/EBP-α, -β, and -δ, with C/EBP-δ having the greatest effect. The myostatin promoter contains two potential C/EBP binding sequences, a CCAAT box, and a C/EBP binding element (CBE). Mutation of the CCAAT box attenuated basal myostatin promoter activity but potentiated C/EBP-δ-activated myostatin promoter activity in C(2)C(12) myotubes in vitro, while mutation of the CBE abolished glucocorticoid receptor and C/EBP-δ responsiveness. The present results support a model in which glucocorticoid-induced increases in C/EBP-δ expression may contribute to myostatin transcription during atrophic states.

The authors recommend chronic suppressive therapy for management of endometriosis symptoms, particularly in the postoperative setting. Empiric treatment is appropriate for those patients without evidence of severe disease. Currently available option may not be effective for nor tolerated by all patients. Newer compounds, including gonadotropin-releasing antagonists and aromatase inhibitors combined with hormonal contraceptives, offer possible alternatives to currently available therapies.

Expression of the cytokine interleukin-6 (IL-6) by skeletal muscle is hugely increased in response to a single bout of endurance exercise, and this appears to be mediated by increases in intracellular calcium. We examined the effects of endurance exercise on IL-6 mRNA levels and promoter activity in skeletal muscle in vivo, and the role of the calcium-activated calcineurin signaling pathway on muscle IL-6 expression in vivo and in vitro. IL-6 mRNA levels in the mouse tibialis anterior (TA) were increased 2–10-fold by a single bout of treadmill exercise or by 3 days of voluntary wheel running. Moreover, an IL-6 promoter-driven luciferase transgene was activated in TA by both treadmill and wheel-running exercise and by injection with a calcineurin plasmid. Exercise also increased muscle mRNA expression of the calcineurin regulatory gene MCIP1, as did treatment of C2C12 myotubes with the calcium ionophore A23187. Cotransfection of C2C12 myotubes with a constitutively active calcineurin construct significantly increased while cotransfection with the calcineurin inhibitor CAIN inhibited activity of a mouse IL-6 promoter-reporter construct. Cotransfection with a myocyte enhancer-factor-2 (MEF-2) expression construct increased basal IL-6 promoter activity and augmented the effects of calcineurin cotransfection, while cotransfection with the MEF-2 antagonist MITR repressed calcineurin-activated IL-6 promoter activity in vitro. Surprisingly, cotransfection with a dominant-negative form of another calcineurin-activated transcription factor, nuclear factor activator of T cells (NFAT), greatly potentiated both basal and calcineurin-stimulated IL-6 promoter activity in C2C12 myotubes. Mutation of the MEF-2 DNA binding sites attenuated, while mutation of the NFAT DNA binding sites potentiated basal and calcineurin-activated IL-6 promoter activity. Finally, CREB and C/EBP were necessary for basal IL-6 promoter activity and sufficient to increase IL-6 promoter activity but had minimal roles in calcineurin-activated IL-6 promoter activity. Together, these results suggest that IL-6 transcription in skeletal muscle cells can be activated by a calcineurin-MEF-2 axis which is antagonized by NFAT.