Skip to main content
SpringerLink
Log in

Removal of serum factors by charcoal treatment promotes adipogenesis via a MAPK-dependent pathway

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

In vitro differentiation of the progenitor cells or preadipocytes into adipocytes is usually achieved by adding an adipogenic mixture (isobutylmethylxanthine, dexamethasone, and insulin, IDI) to medium supplemented with fetal bovine serum (FBS). To study the effects of steroid hormones in vitro, endogenous hormones, growth factors and cytokines are removed by charcoal stripping of serum. However, the effects of charcoal-stripped serum (CS-FBS) per se on adipogenesis have been ignored. Here, we showed that alkaline phosphate activity and nodule formation of osteoprogenitor KS483 cells were lower in CS-FBS than in FBS. Concurrently, abundant amounts of adipocytes were only observed in KS483 cells cultured with CS-FBS, irrespective of the brands of serum used. Inhibition of the p42/44 MAPK pathway by its specific inhibitor PD98059 increased adipogenesis of KS483 cells with FBS, whereas activation of this signalling pathway by EGF blocked adipogenesis of these cells with CS-FBS. Furthermore, the p42/44 MAPK phosphorylation of KS483 cells cultured with CS-FBS was decreased compared with FBS. We concluded that charcoal-stripping of serum removed stimulators of the MAPK signalling pathway and in turn led to downregulation of osteogenesis and upregulation of adipogenesis. Interestingly, the adipogenic mixture IDI stimulated adipogenesis of KS483 cells cultured with CS-FBS, but not with FBS. Furthermore, differential effects of genistein on adipogenesis were observed in KS483 cells cultured with FBS or CS-FBS in combination with IDI. Our results showed that charcoal stripping of serum affected the commitment of KS483 cells and therefore differentially regulated adipogenesis influenced by IDI alone and in combination with genistein. (Mol Cell Biochem 268: 159–167, 2005)

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Gregoire FM, Smas CM, Sul HS: Understanding adipocyte differentiation. Physiol Rev 78(3): 783–809, 1998

    CAS  PubMed  Google Scholar 

  2. Rosen ED, Walkey CJ, Puigserver P, Spiegelman BM: Transcriptional regulation of adipogenesis. Genes Dev 14(11): 1293–1307, 2000

    CAS  Google Scholar 

  3. Rangwala SM, Lazar MA: Transcriptional control of adipogenesis. Annu Rev Nutr 20: 535–559, 2000

    Article  Google Scholar 

  4. Gimble JM, Robinson CE, Wu X, Kelly KA, Rodriguez BR, Kliewer SA, Lehmann JM, Morris DC: Peroxisome proliferator-activated receptor-gamma activation by thiazolidinediones induces adipogenesis in bone marrow stromal cells. Mol Pharmacol 50(5): 1087–1094, 1996

    CAS  PubMed  Google Scholar 

  5. Chen D, Ji X, Harris MA, Feng JQ, Karsenty G, Celeste AJ, Rosen V, Mundy GR, Harris SE: Differential roles for bone morphogenetic protein (BMP) receptor type IB and IA in differentiation and specification of mesenchymal precursor cells to osteoblast and adipocyte lineages. J. Cell Biol 142(1): 295–305, 1998

    Article  Google Scholar 

  6. Pittenger MF, Mackay AM, Beck SC, Jaiswal RK, Douglas R, Mosca JD, Moorman MA, Simonetti DW, Craig S, Marshak DR: Multilineage potential of adult human mesenchymal stem cells. Science 284(5411): 143–147, 1999

    Article  CAS  PubMed  Google Scholar 

  7. Aakvaag A, Utaaker E, Thorsen T, Lea OA, Lahooti H: Growth control of human mammary cancer cells (MCF-7 cells) in culture: Effect of estradiol and growth factors in serum-containing medium. Cancer Res 50(24): 7806–7810, 1990

    Google Scholar 

  8. Puccinelli JM, Omura TH, Strege DW, Jeffrey JJ, Partridge NC: A serum factor promotes collagenase synthesis by an osteoblastic cell line. J Cell Physiol 147(3): 505–513, 1991

    Google Scholar 

  9. Yohay DA, Zhang J, Thrailkill KM, Arthur JM, Quarles LD: Role of serum in the developmental expression of alkaline phosphatase in MC3T3-E1 osteoblasts. J Cell Physiol 158(3): 467–475, 1994

    Google Scholar 

  10. Lindquist DL, de Alarcon PA: Charcoal-dextran treatment of fetal bovine serum removes an inhibitor of human CFU-megakaryocytes. Exp Hematol 15(3): 234–238, 1987

    Google Scholar 

  11. Beresford JN, Bennett JH, Devlin C, Leboy PS, Owen ME: Evidence for an inverse relationship between the differentiation of adipocytic and osteogenic cells in rat marrow stromal cell cultures. J Cell Sci 102(Pt 2): 341–351, 1992

    Google Scholar 

  12. Nuttall ME, Patton AJ, Olivera DL, Nadeau DP, Gowen M: Human trabecular bone cells are able to express both osteoblastic and adipocytic phenotype: implications for osteopenic disorders. J. Bone Miner Res 13(3): 371–382, 1998

    Google Scholar 

  13. Loftus TM, Lane MD: Modulating the transcriptional control of adipogenesis. Curr Opin Genet Dev 7(5): 603–608, 1997

    Google Scholar 

  14. Rosen ED, Hsu CH, Wang X, Sakai S, Freeman MW, Gonzalez FJ, Spiegelman BM: C/EBPalpha induces adipogenesis through PPARgamma: A unified pathway. Genes Dev 16(1): 22–26, 2002

    Google Scholar 

  15. Fajas L, Fruchart JC, Auwerx J: Transcriptional control of adipogenesis. Curr Opin Cell Biol 10(2): 165–173, 1998

    Article  CAS  PubMed  Google Scholar 

  16. Ren D, Collingwood TN, Rebar EJ, Wolffe AP, Camp HS: PPARgamma knockdown by engineered transcription factors: Exogenous PPARgamma2 but not PPARgamma1 reactivates adipogenesis. Genes Dev 16(1): 27–32, 2002

    Google Scholar 

  17. Lazar MA: Becoming fat. Genes Dev 16(1): 1–5, 2002

    Google Scholar 

  18. Lehmann JM, Lenhard JM, Oliver BB, Ringold GM, Kliewer SA: Peroxisome proliferator-activated receptors alpha and gamma are activated by indomethacin and other non-steroidal anti-inflammatory drugs. J Biol Chem 272(6): 3406–3410, 1997

    Article  CAS  PubMed  Google Scholar 

  19. Willson TM, Lambert MH, Kliewer SA: Peroxisome proliferator-activated receptor gamma and metabolic disease. Annu Rev Biochem 70: 341–367, 2001

    Article  CAS  PubMed  Google Scholar 

  20. Camp HS, Tafuri SR: Regulation of peroxisome proliferator-activated receptor gamma activity by mitogen-activated protein kinase. J Biol Chem 272(16): 10811–10816, 1997

    Google Scholar 

  21. Hu E, Kim JB, Sarraf P, Spiegelman, BM: Inhibition of adipogenesis through MAP kinase-mediated phosphorylation of PPARgamma. Science 274(5295): 2100–2103, 1996

    Article  Google Scholar 

  22. Adams M, Reginato MJ, Shao D, Lazar MA, Chatterjee VK: Transcriptional activation by peroxisome proliferator-activated receptor gamma is inhibited by phosphorylation at a consensus mitogen-activated protein kinase site. J Biol Chem 272(8): 5128–5132, 1997

    Google Scholar 

  23. Chan GK, Deckelbaum RA, Bolivar I, Goltzman D, Karaplis AC: PTHrP inhibits adipocyte differentiation by down-regulating PPARgamma activity via a MAPK-dependent pathway. Endocrinology 142(11): 4900–4909, 2001

    Google Scholar 

  24. Dang ZC, Audinot V, Papapoulos SE, Boutin JA, Lowik CW: Peroxisome proliferator-activated receptor gamma (PPARgamma) as a molecular target for the soy phytoestrogen genistein. J Biol Chem 278(2): 962–967, 2003

    Article  CAS  PubMed  Google Scholar 

  25. Wang H, Malbon CC: G(s)alpha repression of adipogenesis via Syk. J Biol Chem 274(45): 32159–32166, 1999

    Google Scholar 

  26. Harmon AW, Patel YM, Harp JB, Harmon AW, Harp JB: Genistein inhibits CCAAT/enhancer-binding protein beta (C/EBPbeta) activity and 3T3-L1 adipogenesis by increasing C/EBP homologous protein expression. Biochem J 367(Pt 1): 203–208, 2002

    Google Scholar 

  27. Yamashita T, Asano K, Takahashi N, Akatsu T, Udagawa N, Sasaki T, Martin TJ, Suda T: Cloning of an osteoblastic cell line involved in the formation of osteoclast-like cells. J Cell Physiol 145(3): 587–595, 1990

    Google Scholar 

  28. Yamashita T, Ishii H, Shimoda K, Sampath TK, Katagiri T, Wada M, Osawa T, Suda T: Subcloning of three osteoblastic cell lines with distinct differentiation phenotypes from the mouse osteoblastic cell line KS-4. Bone 19(5): 429–436, 1996

    Google Scholar 

  29. Deckers MM, Karperien M, van der Bent C, Yamashita T, Papapoulos SE, Lowik CW: Expression of vascular endothelial growth factors and their receptors during osteoblast differentiation. Endocrinology 141(5): 1667–1674, 2000

    Google Scholar 

  30. Dang ZC, van Bezooijen RL, Karperien M, Papapoulos SE, Lowik CW: Exposure of KS483 cells to estrogen enhances osteogenesis and inhibits adipogenesis. J Bone Miner Res 17(3): 394–405, 2002

    CAS  PubMed  Google Scholar 

  31. Labarca C, Paigen K: A simple, rapid, and sensitive DNA assay procedure. Anal Biochem 102(2): 344–352, 1980

    CAS  PubMed  Google Scholar 

  32. Schiller PC, D’Ippolito G, Brambilla R, Roos BA, Howard GA: Inhibition of gap-junctional communication induces the trans-differentiation of osteoblasts to an adipocytic phenotype in vitro. J Biol Chem 276(17): 14133–14138, 2001

    Google Scholar 

  33. Xiao G, Jiang D, Thomas P, Benson MD, Guan K, Karsenty G, Franceschi RT: MAPK pathways activate and phosphorylate the osteoblast-specific transcription factor, Cbfa1. J Biol Chem 275(6): 4453–4459, 2000

    Google Scholar 

  34. Lai CF, Chaudhary L, Fausto A, Halstead LR, Ory DS, Avioli LV, Cheng SL: Erk is essential for growth, differentiation, integrin expression, and cell function in human osteoblastic cells. J Biol Chem 276(17): 14443–14450, 2001

    Google Scholar 

  35. Parhami F, Jackson SM, Tintut Y, Le V, Balucan JP, Territo M, Demer LL: Atherogenic diet and minimally oxidized low density lipoprotein inhibit osteogenic and promote adipogenic differentiation of marrow stromal cells. J Bone Miner Res 14(12): 2067–2078, 1999

    Google Scholar 

  36. Diascro DD Jr, Vogel RL, Johnson TE, Witherup KM, Pitzenberger SM, Rutledge SJ, Prescott DJ, Rodan GA, Schmidt A: High fatty acid content in rabbit serum is responsible for the differentiation of osteoblasts into adipocyte-like cells. J Bone Miner Res 13(1): 96–106, 1998

    Google Scholar 

  37. Hauner H, Rohrig K, Petruschke T: Effects of epidermal growth factor (EGF), platelet-derived growth factor (PDGF) and fibroblast growth factor (FGF) on human adipocyte development and function. Eur J Clin Invest 25(2): 90–96, 1995

    Google Scholar 

  38. Dennis JE, Merriam A, Awadallah A, Yoo JU, Johnstone B, Caplan AI: A quadripotential mesenchymal progenitor cell isolated from the marrow of an adult mouse. J Bone Miner Res 14(5): 700–709, 1999

    Google Scholar 

  39. Liu X, Malbon CC, Wang HY: Identification of amino acid residues of Gsalpha critical to repression of adipogenesis. J Biol Chem 273(19): 11685–11694, 1998

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Endocrinology and Metabolic Diseases, Leiden University Medical Center, Leiden, The Netherlands

    Z. C. Dang & C. W. G. M. Lowik

Authors
  1. Z. C. Dang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C. W. G. M. Lowik
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Z. C. Dang.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Dang, Z.C., Lowik, C.W.G.M. Removal of serum factors by charcoal treatment promotes adipogenesis via a MAPK-dependent pathway. Mol Cell Biochem 268, 159–167 (2005). https://doi.org/10.1007/s11010-005-3857-7

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11010-005-3857-7

Key words

Search

Navigation