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The Journal of Physiological Sciences

, Volume 68, Issue 2, pp 137–151 | Cite as

Sphingosine-1-phosphate pretreatment amends hypoxia-induced metabolic dysfunction and impairment of myogenic potential in differentiating C2C12 myoblasts by stimulating viability, calcium homeostasis and energy generation

  • Babita Rahar
  • Sonam Chawla
  • Sanjay Pandey
  • Anant Narayan Bhatt
  • Shweta SaxenaEmail author
Original Paper

Abstract

Sphingosine-1-phosphate (S1P) has a role in transpiration in patho-physiological signaling in skeletal muscles. The present study evaluated the pre-conditioning efficacy of S1P in facilitating differentiation of C2C12 myoblasts under a normoxic/hypoxic cell culture environment. Under normoxia, exogenous S1P significantly promoted C2C12 differentiation as evident from morphometric descriptors and differentiation markers of the mature myotubes, but it could facilitate only partial recovery from hypoxia-induced compromised differentiation. Pretreatment of S1P optimized the myokine secretion, intracellular calcium release and energy generation by boosting the aerobic/anaerobic metabolism and mitochondrial mass. In the hypoxia-exposed cells, there was derangement of the S1PR1–3 expression patterns, while the same could be largely restored with S1P pretreatment. This is being proposed as a plausible underlying mechanism for the observed pro-myogenic efficacy of exogenous S1P preconditioning. The present findings are an invaluable addition to the existing knowledge on the pro-myogenic potential of S1P and may prove beneficial in the field of hypoxia-related myo-pathologies.

Keywords

Normoxia Hypoxia S1P Differentiation Myogenesis 

Notes

Compliance with ethical standards

Funding

The authors are thankful to the Director of DIPAS for supporting this study. This work was funded by the Defence Research and Development Organisation (DRDO), India [Grant No. S & T-09 DIP-251 A 2. 3 (AB)]. The authors acknowledge DIPAS, DRDO, the University Grants Commission (UGC) and Council of Scientific and Industrial Research (CSIR), India, for providing the necessary facilities and funding for this study.

Conflict of interest

The authors have no conflict of interest.

Ethical approval

The article does not encompass studies conducted with human participants or animals.

Supplementary material

12576_2016_518_MOESM1_ESM.doc (9.4 mb)
Supplementary material 1 (DOC 9660 kb)

References

  1. 1.
    Giaccia AJ, Simon MC, Johnson R (2004) The biology of hypoxia: the role of oxygen sensing in development, normal function, and disease. Genes Dev. doi: 10.1101/gad.1243304 CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Kikusato M, Toyomizu M (2015) Moderate dependence of reactive oxygen species production on membrane potential in avian muscle mitochondria oxidizing glycerol 3-phosphate. J Physiol Sci. doi: 10.1007/s12576-015-0395-2 CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Murray Andrew J (2009) Metabolic adaptation of skeletal muscle to high altitude hypoxia: how new technologies could resolve the controversies. Genome Med. doi: 10.1186/gm117 CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Grocott M, Montgomery H, Vercueil A (2007) High-altitude physiology and pathophysiology: implications and relevance for intensive care medicine. Crit Care. doi: 10.1186/cc5142 CrossRefPubMedPubMedCentralGoogle Scholar
  5. 5.
    Nakai N, Fujita R, Kawano F, Takahashi K, Ohira T, Shibaguchi T, Nakata K, Ohira Y (2014) Retardation of C2C12 myoblast cell proliferation by exposure to low-temperature atmospheric plasma. J Physiol Sci. doi: 10.1007/s12576-014-0328-5 CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Karalaki M, Fili S, Philippou A, Koutsilieris M (2009) Muscle regeneration: cellular and molecular events. In Vivo. 23:79–96Google Scholar
  7. 7.
    Chaillou T, Koulmann N, Meunier A, Pugnière P, McCarthy JJ, Beaudry M, Bigard X (2014) Ambient hypoxia enhances the loss of muscle mass after extensive injury. Pflugers Arch. doi: 10.1007/s00424-013-1336-7 CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Nimker C, Kaur G, Revo A, Chaudhary P, Bansal A (2015) Ethyl 3,4-dihydroxy benzoate, a unique preconditioning agent for alleviating hypoxia-mediated oxidative damage in L6 myoblasts cells. J Physiol Sci. doi: 10.1007/s12576-014-0348-1 CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Di Carlo A, De Mori R, Martelli F, Pompilio G, Capogrossi MC, Germani A (2004) Hypoxia inhibits myogenic differentiation through accelerated MyoD degradation. J Biol Chem. doi: 10.1074/jbc.M313931200 CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Hoppeler H, Kleinert E, Schlegel C, Claassen H, Howald H, Kayar SR, Cerretelli P (1990) Morphological adaptations of human skeletal muscle to chronic hypoxia. Int J Sports Med Suppl. doi: 10.1055/s-2007-1024846 CrossRefGoogle Scholar
  11. 11.
    Attanasio S, Snell J (2009) Therapeutic angiogenesis in the management of critical limb ischemia: current concepts and review. Cardiol Rev. doi: 10.1097/CRD.0b013e318199e9b7 CrossRefGoogle Scholar
  12. 12.
    Chalfant CE, Spiegel S (2005) Sphingosine 1-phosphate and ceramide 1-phosphate: expanding roles in cell signaling. J Cell Sci. doi: 10.1242/jcs.02637 CrossRefGoogle Scholar
  13. 13.
    Donati C, Cencetti F, Bruni P (2013) Sphingosine 1-phosphate axis: a new leader actor in skeletal muscle biology. Front Physiol. doi: 10.3389/fphys.2013.00338 CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Chawla S, Sahni C, Tulsawani R, Singh M, Saraswat D, Bansal A, Saxena S (2014) Exogenous sphingosine 1-phosphate protects murine splenocytes against hypoxia-induced injury. Lipids. doi: 10.1007/s11745-013-3860-9 CrossRefGoogle Scholar
  15. 15.
    Chawla S, Rahar B, Singh M, Bansal A, Saraswat D, Saxena S (2014) Exogenous sphingosine-1-phosphate boosts acclimatization in rats exposed to acute hypobaric hypoxia: assessment of haematological and metabolic effects. PLoS One. doi: 10.1371/journal.pone.0098025 CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Chawla S, Rahar B, Saxena S (2016) S1P prophylaxis mitigates acute hypobaric hypoxia-induced molecular, biochemical, and metabolic disturbances: a preclinical report. IUBMB Life. doi: 10.1002/iub.1489 CrossRefGoogle Scholar
  17. 17.
    Burattini S, Ferri P, Battistelli M, Curci R, Luchetti F, Falcieri E (2004) C2C12 murine myoblasts as a model of skeletal muscle development: morpho-functional characterization. Eur J Histochem 48:223–233PubMedPubMedCentralGoogle Scholar
  18. 18.
    Donati C, Meacci E, Nuti F, Becciolini L, Farnararo M, Bruni P (2005) Sphingosine-1-phosphate regulates myogenic differentiation: a major role for S1P2 receptor. Faseb J. doi: 10.1096/fj.04-1780fje CrossRefGoogle Scholar
  19. 19.
    Bradford MM (1976) A rapid and sensitive for the quantitation of microgram quantitites of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  20. 20.
    Kornberg A (1955) Lactate dehydrogenase of muscle. In: Colowick SP, Kaplan NO (eds) Methods in enzymology, vol 1. Acad Press, New York, pp 441–443CrossRefGoogle Scholar
  21. 21.
    Supowit SC, Harris BG (1976) A scarissuum hexokinase: purification and possible function in compartmentation of glucose 6-phosphate in muscle. Biochim Biophys Acta. doi: 10.1016/0005-2744(76)90007-3 CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Shepherd D, Garland PB (1966) Citrate synthase from liver. Biochem Biophys Res Commun. doi: 10.1016/0076-6879(69)13006-2 CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Miyake T, McDermott JC, Gramolili AO (2011) A method for the direct identification of differentiating muscle cells by a fluorescent mitochondrial dye. PLoS One. doi: 10.1371/journal.pone.0028628 CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Rasband WS (1997–2015) ImageJ. U. S. National Institutes of Health, Bethesda. http://imagej.nih.gov/ij/ (1997–2015)
  25. 25.
    Schmittgen TD, Livak KJ (2008) Analyzing real-time PCR data by the comparative C(T) method. Nat Protoc 3(6):1101–1108CrossRefGoogle Scholar
  26. 26.
    Burattini S, Ferri P, Battistelli M, Curci R, Luchetti F, Falcieri E (2004) C2C12 murine myoblasts as a model of skeletal muscle development: morpho-functional characterization. Eur J Histochem 48(3):223–233PubMedPubMedCentralGoogle Scholar
  27. 27.
    Ceafalan LC, Popescu BO, Hinescu ME (2014) Cellular players in skeletal muscle regeneration. Biomed Res Int. doi: 10.1155/2014/957014 CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Muñoz-Cánoves P, Scheele C, Pedersen BK, Serrano AL (2013) Interleukin-6 myokine signaling in skeletal muscle: a double-edged sword? FEBS J. doi: 10.1111/febs.12338 CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Li YP (2003) TNF-alpha is a mitogen in skeletal muscle. Am J Physiol Cell Physiol. doi: 10.1152/ajpcell.00453.2002 CrossRefGoogle Scholar
  30. 30.
    Tolosa L, Morlá M, Iglesias A, Busquets X, Lladó J, Olmos G (2005) IFN-gamma prevents TNF-alpha-induced apoptosis in C2C12 myotubes through down-regulation of TNF-R2 and increased NF-kappaB activity. Cell Signal. doi: 10.1016/j.cellsig.2005.02.001 CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Przybylski RJ, MacBride RG, Kirby AC (1989) Calcium regulation of skeletal myogenesis. I. Cell content critical to myotube formation. In Vitro Cell Dev Biol. doi: 10.1007/BF02623667 CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Willkomm L, Schubert S, Jung R, Elsen M, Borde J, Gehlert S, Suhr F, Bloch W (2014) Lactate regulates myogenesis in C2C12 myoblasts in vitro. Stem Cell Res. doi: 10.1016/j.scr.2014.03.004 CrossRefPubMedPubMedCentralGoogle Scholar
  33. 33.
    Yun Z, Lin Q, Giaccia AJ (2005) Adaptive myogenesis under hypoxia. Mol Cell Biol. doi: 10.1128/MCB.25.8.3040-3055.2005 CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Hara M, Tabata K, Suzuki T, Do MK, Mizunoya W, Nakamura M, Nishimura S, Tabata S, Ikeuchi Y, Sunagawa K, Anderson JE, Allen RE, Tatsumi R (2012) Calcium influx through a possible coupling of cation channels impacts skeletal muscle satellite cell activation in response to mechanical stretch. Am J Physiol Cell Physiol. doi: 10.1152/ajpcell.00068.2012 CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Donati C, Nincheri P, Cencetti F, Rapizzi E, Farnararo M, Bruni P (2007) Tumor necrosis factor-alpha exerts pro-myogenic action in C2C12 myoblasts via sphingosine kinase/S1P2 signaling. FEBS Lett. doi: 10.1016/j.febslet.2007.08.007 CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Rapizzi E, Donati C, Cencetti F, Nincheri P, Bruni P (2008) Sphingosine 1-phosphate differentially regulates proliferation of C2C12 reserve cells and myoblasts. Mol Cell Biochem. doi: 10.1007/s11010-008-9780-y CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Loh KC, Leong WI, Carlson ME, Oskouian B, Kumar A, Fyrst H, Zhang M, Proia RL, Hoffman EP, Saba JD (2012) Sphingosine-1-phosphate enhances satellite cell activation in dystrophic muscles through a S1PR2/STAT3 signaling pathway. PLoS One. doi: 10.1371/journal.pone.0037218 CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    Porter GA Jr, Makuck RF, Rivkees SA (2002) Reduction in intracellular calcium levels inhibits myoblast differentiation. J Biol Chem. doi: 10.1074/jbc.M203961200 CrossRefPubMedPubMedCentralGoogle Scholar
  39. 39.
    Kanatous SB, Mammen PP, Rosenberg PB, Martin CM, White MD, Dimaio JM, Huang G, Muallem S, Garry DJ (2009) Hypoxia reprograms calcium signaling and regulates myoglobin expression. Am J Physiol Cell Physiol. doi: 10.1152/ajpcell.00428.2008 CrossRefPubMedPubMedCentralGoogle Scholar
  40. 40.
    Meacci E, Cencetti F, Formigli L, Squecco R, Donati C, Tiribilli B, Quercioli F, ZecchiOrlandini S, Francini F, Bruni P (2002) Sphingosine 1-phosphate evokes calcium signals in C2C12 myoblasts via Edg3 and Edg5 receptors. Biochem J. doi: 10.1042/bj3620349 CrossRefPubMedPubMedCentralGoogle Scholar
  41. 41.
    Sassoli C, Formigli L, Bini F, Tani A, Squecco R, Battistini C, Zecchi-Orlandini S, Francini F, Meacci E (2011) Effects of S1P on skeletal muscle repair/regeneration during eccentric contraction. J Cell Mol Med. doi: 10.1111/j.1582-4934.2010.01250.x CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Koopman R, Ly CH, Ryall JG (2014) A metabolic link to skeletal muscle wasting and regeneration. Front Physiol. doi: 10.3389/fphys.2014.00032 CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Li W, Hu ZF, Chen B, Ni GX (2013) Response of C2C12 myoblasts to hypoxia: the relative roles of glucose and oxygen in adaptive cellular metabolism. Biomed Res Int. doi: 10.1155/2013/326346 CrossRefPubMedPubMedCentralGoogle Scholar
  44. 44.
    Leary SC, Battersby BJ, Hansford RG, Moyes CD (1998) Interactions between bioenergetics and mitochondrial biogenesis. Biochim Biophys Acta. doi: 10.1016/S0005-2728(98)00105-4 CrossRefGoogle Scholar
  45. 45.
    Yamada K (2016) Energetics of muscle contraction: further trials. J Physiol Sci. doi: 10.1007/s12576-016-0470-3 CrossRefPubMedPubMedCentralGoogle Scholar
  46. 46.
    Wheaton WW, Chandel NS (2011) Hypoxia. 2. Hypoxia regulates cellular metabolism. Am J Physiol Cell Physiol 300:C385–C393CrossRefGoogle Scholar
  47. 47.
    Seagroves TN, Ryan HE, Lu H, Wouters BG, Knapp M, Thibault P, Laderoute K, Johnson RS (2001) Transcription factor HIF-1 is a necessary mediator of the Pasteur effect in mammalian cells. Mol Cell Biol. doi: 10.1128/MCB.21.10.3436-3444.2001 CrossRefPubMedPubMedCentralGoogle Scholar
  48. 48.
    Hamilton DL, Beall C, Jeromson S, Chevtzoff C, Cuthbertson DJ, Ashford ML (2014) Kv1.3 inhibitors have differential effects on glucose uptake and AMPK activity in skeletal muscle cell lines and mouse ex vivo skeletal muscle. J Physiol. doi: 10.1007/s12576-013-0285-4 CrossRefGoogle Scholar
  49. 49.
    Wagatsuma A, Sakuma K (2013) Mitochondria as a potential regulator of myogenesis. Sci World. doi: 10.1155/2013/593267 CrossRefGoogle Scholar
  50. 50.
    Shen Z, Liu C, Liu P, Zhao J, Xu W (2014) Sphingosine 1-phosphate (S1P) promotes mitochondrial biogenesis in Hep G2 cells by activating Peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α). Cell Stress Chaperones. doi: 10.1007/s12192-013-0480-5 CrossRefPubMedPubMedCentralGoogle Scholar
  51. 51.
    Biswas G, Adebanjo OA, Freedman BD, Anandatheerthavarada HK, Vijayasarathy C, Zaidi M, Kotlikoff M, Avadhani NG (1999) Retrograde Ca2+ signaling in C2C12 skeletal myocytes in response to mitochondrial genetic and metabolic stress: a novel mode of inter-organelle crosstalk. EMBO J 18(3):522–533CrossRefGoogle Scholar
  52. 52.
    Ieronimakis N, Pantoja M, Hays AL, Dosey TL, Qi J, Fischer KA, Hoofnagle AN, Sadilek M, Chamberlain JS, Ruohola-Baker H, Reyes M (2013) Increased sphingosine-1-phosphate improves muscle regeneration in acutely injured mdx mice. Skelet Muscle. doi: 10.1186/2044-5040-3-20 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© The Physiological Society of Japan and Springer Japan 2017

Authors and Affiliations

  • Babita Rahar
    • 1
  • Sonam Chawla
    • 1
  • Sanjay Pandey
    • 2
  • Anant Narayan Bhatt
    • 2
  • Shweta Saxena
    • 3
    Email author
  1. 1.Experimental Biology Division, Defence Institute of Physiology and Allied Sciences (DIPAS)Defence Research and Development Organization (DRDO)DelhiIndia
  2. 2.Division of Metabolic and Cell Signaling Research, Institute of Nuclear Medicine and Allied Sciences (INMAS)Defence Research and Development Organization (DRDO)DelhiIndia
  3. 3.Medicinal and Aromatic Plant Division, Defence Institute of High Altitude Research (DIHAR)Defence Research and Development Organization (DRDO), Ministry of DefenceLeh-LadakhIndia

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