Effect of Androstenedione on Adipogenesis in Murine C3H10T1/2 Mesenchymal Cells

Pandurangan Ramaraj^1*, Jorge N. Artaza², Indrani Sinha-Hikim², Wayne E. Taylor^2,3

¹Department of Biochemistry, Kirksville College of Osteopathic Medicine, A.T. Still University of Health Sciences, Kirksville, MO, USA.
²Division of Endocrinology, Charles R. Drew University of Medicine and Science, Los Angeles, CA, USA.
³Department of Health and Life Sciences, Charles R. Drew University of Medicine and Science, Los Angeles, CA, USA.

DOI: 10.4236/ojemd.2015.52002                

Abstract

Clinical trials of weak androgen androstenedione (AD) administered at a high concentration, showed an increase in muscle mass in men like strong androgens testosterone (T) and dihydrotestosterone (DHT), but did not show any inhibitory effect on fat mass unlike strong androgens. This observation prompted us to check the in-vitro effect of AD on adipogenesis using mouse mesenchymal multipotent cells (C3H10T1/2), which can differentiate into both myoblasts and adipocytes. Results indicated that AD inhibited adipogenesis at 10 nM, 100 nM and 1 μM concentrations, but not at 10 μM concentration. AD did not inhibit adipogenesis at 10 μM concentration and also did not inhibitmyogenesis at 10 μM concentration. Addition of bicalutamide, an androgen receptor (AR) antagonist decreased myogenesis and increased adipogenesis, indicating that the effect of AD was mediated through AR. Another weak androgen dehydroepiandrosterone (DHEA) also showed the same pattern of adipogenesis in 10T1/2 cells. AD also showed a similar pattern of adipogenesis in 3T3-L1 preadipocyte cells. Thus, the in-vitro results of AD on adipogenesis correlated with the in-vivo results of AD on fat-mass from clinical trials and suggested a possible difference in biological action between weak androgens (AD, DHEA) and strong androgens (T, DHT) on adipogenesis. Since the biological action of AD was mediated through AR, this physiological difference onadipogenesis could be due to the nature (partial agonist/antagonist) of AD binding to AR.

Keywords

Androstenedione, Adipogenesis, Myogenesis, Androgen Receptor, Bicalutamide

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	Snyder, P.J., Peachey, H., Hannoush, P., Berlin, J.A., Loh, L., Lenrow, D.A., Holmes, J.H., Dlewati, A., Santanna, J., Rosen, C.J. and Strom, B.L. (1999) Effect of Testosterone Treatment on Body Composition and Muscle Strength in Men over 65 Years of Age. The Journal of Clinical Endocrinology and Metabolism, 84, 2647-2653.
[2]	Bhasin, S., Storer, T.W., Berman, N., Callegari, C., Clevenger, B.A., Philips, J., Bunnell, T., Tricker, R., Shirazi, A. and Casaburi, R. (1996) The Effects of Supraphysiologic Doses of Testosterone on Muscle Size and Strength in Normal Men. The New England Journal of Medicine, 335, 1-7. http://dx.doi.org/10.1056/NEJM199607043350101
[3]	Bhasin, S., Woodhouse, L., Casaburi, R., Singh, A.B., Bhasin, D., Berman, N., Chen, X.H., Yarasheski, K.E., Magliano, L., Dzekov, C., Dzekov, J., Bross, R., Philips, J., Sinha-Hikim, I., Shen, R.Q. and Storer, T.W. (2001) Testosterone Dose-Response Relationships in Healthy Young Men. American Journal of Physiology—Endocrinology and Metabolism, 281, E1172-E1181.
[4]	Bhasin, S., Woodhouse, L. and Storer, T.W. (2001) Proof of the Effect of Testosterone on Skeletal Muscle. Journal of Endocrinology, 170, 27-38. http://dx.doi.org/10.1677/joe.0.1700027
[5]	Katznelson, L., Rosenthal, D.I., Rosol, M.S., Anderson, E.J., Hayden, D.L., Schoenfeld, D.A. and Klibanski, A. (1998) Using Quantitative CT to Assess Adipose Distribution in Adult Men with Acquired Hypogonadism. American Journal of Roentgenology, 170, 423-427. http://dx.doi.org/10.2214/ajr.170.2.9456958
[6]	Bhasin, S., Storer, T.W., Berman, N., Yarasheski, K.E., Philips, J., Clevenger, B., Lee, W.P., Bunnell, T.J. and Casaburi, R. (1997) Testossterone Replacement Increases Fat-Free Mass and Muscle Size in Hypogonadal Men. The Journal of Clinical Endocrinology and Metabolism, 82, 407-413.
[7]	Brodsky, I.G., Balagopal, P. and Nair, K.S. (1996) Effects of Testosterone Replacement on Muscle Mass and Muscl Protein Synthesis in Hypogonadal Men—A Clinical Research Center Study. The Journal of Clinical Endocrinology and Metabolism, 81, 3469-3475.
[8]	Snyder, P.J., Peachey, H., Berlin, J.A., Hannoush, P., Haddad, G., Dlewati, A., Santanna, J., Loh, L., Lenrow, D.A., Holmews, J.H., Kapoor, S.C., Atkinson, L.E. and Strom, B.L. (2000) Effects of Testosterone Replacement in Hypogonadal Men. The Journal of Clinical Endocrinology and Metabolism, 85, 2670-2677.
[9]	Wang, C., Swerdloff, R.S., Iranmanesh, A., Dobs, A., Snyder, P.J., Cunningham, G., Matsumoto, A.M., Weber, T., Berman, N. and Testosterone Gel Study Group (2000) Transdermal Testosterone Gel Improves Sexual Function, Mood, Muscle Strength and Body Composition Parameters in Hypogonadal Men. The Journal of Clinical Endocrinology and Metabolism, 85, 2839-2853.
[10]	Sinha-Hikim, I., Artaza, J.N., Woodhouse, L., Gonzalez-Cadavid, N., Singh, A.B., Lee, M.I., Storer, T.W., Casaburi, R., Shen, R.Q. and Bhasin, S. (2002) Testosterone-Induced Increase in Muscle Size in Healthy Men is Associated with Muscle Fiber Hypertrophy. The American Journal of Physiology: Endocrinology and Metabolism, 283, E154-E164.
[11]	Wilson, J.D. (1988) Androgen Abuse by Athletes. Endocrine Reviews, 9, 181-199. http://dx.doi.org/10.1210/edrv-9-2-181
[12]	Cowan, D.A. and Kicman, A.T. (1997) Doping in Sport and Society: Misuse, Analytical Tests and Legal Aspects. Clinical Chemistry, 43, 1261-1279.
[13]	Blaquier, J., Forchielli, E. and Dorfman, R.I. (1967) In Vitro Metabolism of Androgens in Whole Human Blood. Acta Endocrinologica, 55, 697-704.
[14]	Mahesh, V.B. and Greenblatt, R.B. (1962) The in Vivo Conversion of Dehydroepiandrosterone and Androstenedione to Testosterone in Humans. European Journal of Endocrinology, 41, 400-406.
[15]	Morales, A.J., Nolan, J.J., Nelson, J.C. and Yen, S.S.C. (1994) Effects of Replacement Dose of Dehydroepiandrosterone in Men and Women of Advanced Age. The Journal of Clinical Endocrinology & Metabolism, 78, 1360-1367.
[16]	http://www.deadiversion.usdoj.gov/fed_regs/rules/2005/fr1216.htm
[17]	Jasuja, R., Ramaraj, P., Mac, R.P., Singh, A.B., Storer, T.W., Artaza, J.N., Miller, A., Singh, R., Taylor, W.E., Lee, M.L., Davidson, T., Sinha-Hikim, I., Gonzalez-Cadavid, N. and Bhasin, S. (2005) Δ-4-Androstene-3,17-Dione Binds Androgen Receptor, Promotes Myogenesis in Vitro and Increases Serum Testosterone Levels, Fat-Free Mass, and Muscle Strength in Hypogonadal Men. The Journal of Clinical Endocrinology & Metabolism, 90, 855-863. http://dx.doi.org/10.1210/jc.2004-1577
[18]	Lassar, A.B., Peterson, B.M. and Weintraub, H. (1986) Transfection of a DNA Locus That Mediates the Conversion of 10T1/2 Fibroblasts to Myoblasts. Cell, 47, 649-656. http://dx.doi.org/10.1016/0092-8674(86)90507-6
[19]	Fischer, L., Boland, G. and Tuan, R.S. (2002) Wnt-3A Enhances Bone Morphogenic Protein-2-Mediated Chondrogenesis of Murine C3H10T1/2 Mesenchymal Cells. Journal of Biological Chemistry, 277, 30870-30878. http://dx.doi.org/10.1074/jbc.M109330200
[20]	Singh, R., Artaza, J.N., Taylor, W.E., Gonzalez-Cadavid, N.F. and Bhasin, S. (2003) Androgens Stimulate Myogenic Differentiation and Inhibit Adipogenesis in C3H10T1/2 Pluripotent Cells through an Androgen Receptor-Mediated Pathway. Endocrinology, 144, 5081-5088. http://dx.doi.org/10.1210/en.2003-0741
[21]	Singh, R., Artaza, J.N., Taylor, W.E., Braga, M., Yum, X., Gonzalez-Cadavid, N. and Bhasin, S. (2006) Testosterone Inhibits Adipogenic Differentiation in 3T3-L1 Cells; Nuclear Translocation of Androgen Receptor Complex with β-Catenin and T-Cell Factor 4 May Bypass Canonical Wnt Signaling to Down-Regulate Adipogenic Transcription Factors. Endocrinology, 147, 141-154. http://dx.doi.org/10.1210/en.2004-1649
[22]	Bennet, C.N., Ross, S.E., Longo, K.A., Bajnok, L., Hemati, N., Johnson, K.W., Harrison, S.D. and MacDougald, O.A. (2002) Regulation of Wnt Signaling during Adipogenesis. Journal of Biological Chemistry, 277, 30998-31004. http://dx.doi.org/10.1074/jbc.M204527200
[23]	Chen, F., Knecht, K., Leu, C., Rutledge, S.J., Scafonas, A., Gambone, C., Vogel, R., Zhang, H., Kasparcova, V., Bai, C., Harada, S., Schmidt, A., Reszka, A. and Freedman, L. (2004) Partial Agonist/Antagonist Properties of Androstenedione and 4-Androsten-3β,17β-Diol. The Journal of Steroid Biochemistry and Molecular Biology, 91, 247-257. http://dx.doi.org/10.1016/j.jsbmb.2004.04.009
[24]	Soto, A.M. and Sonnenschein, C. (1987) Cell Proliferation of Estrogen-Sensitive Cells: The Case for Negative Control. Endocrine Reviews, 8, 44-52. http://dx.doi.org/10.1210/edrv-8-1-44
[25]	Sonnenschein, C., Olea, N., Pasanen, M.E. and Soto, M.E. (1989) Negative Control of Cell Proliferation: Human Prostate Cancer Cell and Androgens. Cancer Research, 49, 3473-3481.
[26]	Launoit, Y., Veilleus, R., Dufour, M., Simrad, J. and Labrie, F. (1991) Characteristics of the Biphasic Action of Androgens and of the Potent Antiproliferative Effects of the New Pure Anti-Estrogen EM-139 on Cell Cycle Kinetic Parameters in LNCaP Human Prostatic Cancer Cells. Cancer Research, 51, 5165-5170.
[27]	Glickman, S.E., Frank, L.G., Davidson, J.M., Smith, E.R. and Siteri, P.K. (1987) Androstenedione May Organize or Activate Sex-Reversed Traits in Female Spotted Hyenas. Proceedings of the National Academy of Sciences of the United States of America, 84, 3444-3447. http://dx.doi.org/10.1073/pnas.84.10.3444
[28]	Kurk, H. (1972) The Spotted Hyena. University of Chicago Press, Chicago, 1-335.