Analysis of Gene Expression and Morphology of P19 Cells Cultured in an Apatite-Fiber Scaffold

Abstract:

Heart disease is the second most common cause of mortality in Japan. Most cases of late stage heart failure can only be effectively treated by a heart transplant. Cardiac tissue engineering is emerging both as a new approach for improving the treatment of heart failure and for developing new cardiac drugs. Apatite-fiber scaffold (AFS) was originally designed as a substitute material for bone. AFS contains two sizes of pores and is appropriate for the three dimensional proliferation and differentiation of osteoblasts. To establish engineered heart tissue, a pluripotent embryonal carcinoma cell line, P19.CL6, was cultured in AFS. P19.CL6 cells seeded into AFS proliferated well. Generally, cardiac differentiation of P19.CL6 cells is induced by treating suspension-cultured cells with dimethyl sulfoxide (DMSO), after which the cells form spheroids. However, our results showed that P19.CL6 cells cultured in AFS differentiated into myocytes without forming spheroidal aggregates, and could be cultured for at least one month. Thus, we conclude that AFS is a good candidate as a scaffold for cardiac tissue engineering.

• Periodical: Key Engineering Materials (Volumes 529-530)
• Pages: 370-373
• DOI: https://doi.org/10.4028/www.scientific.net/KEM.529-530.370
• Online since: November 2012
• Authors: Hide Ishii, Yuya Mukai, Mamoru Aizawa, Nobuyuki Kanzawa
• Keywords: Apatite-Fiber Scaffold (AFS), Cardiac Tissue Engineering, P19.CL6 Cells, RT-PCR

References

[1] K. Dracup, J.A. Walden, L.W. Stevenson, M.L. Brecht, Quality-Of-Life in Patients with Advanced Heart-Failure, Journal of Heart and Lung Transplantation 11 (1992) 273-279.

[2] M. Aizawa, A.E. Porter, S.M. Best, W. Bonfield, Ultrastructural observation of single-crystal apatite fibres, Biomaterials 26 (2005) 3427-3433.

DOI: 10.1016/j.biomaterials.2004.09.044

[3] M. Aizawa, T. Terado, F.S. Howell, K. Itatani, Preparation of spherical apatite particles by the homogeneous precipitation method in the presence of magnesium ions and their ion-exchange properties, Materials Research Bulletin 34 (1999).

DOI: 10.1016/s0025-5408(99)00118-x

[4] M. Honda, T.J. Fujimi, S. Izumi, K. Izawa, M. Aizawa, H. Morisue, T. Tsuchiya, N. Kanzawa, Topographical analyses of proliferation and differentiation of osteoblasts in micro- and macropores of apatite-fiber scaffold, Journal of Biomedical Materials Research Part A 94A (2010).

DOI: 10.1002/jbm.a.32779

[5] A. HabaraOhkubo, Differentiation of beating cardiac muscle cells from a derivative of P19 embryonal carcinoma cells, Cell Structure and Function 21 (1996) 101-110.

DOI: 10.1247/csf.21.101

[6] M.D. Abramoff, P.J. Magalhaes, S.J. Ram, Image processing with ImageJ, Biophotonics International 11 (2004) 36-42.

[7] K. Monzen, I. Shiojima, Y. Hiroi, S. Kudoh, T. Oka, E. Takimoto, D. Hayashi, T. Hosoda, A. Habara-Ohkubo, T. Nakaoka, T. Fujita, Y. Yazaki, I. Komuro, Bone morphogenetic proteins induce cardiomyocyte differentiation through the mitogen-activated protein kinase kinase kinase TAK1 and cardiac transcription factors Csx/Nkx-2. 5 and GATA-4, Molecular and Cellular Biology 19 (1999).

DOI: 10.1128/mcb.19.10.7096

[8] G.E. Lyons, S. Schiaffino, D. Sassoon, P. Barton, M. Buckingham, Developmental Regulation of Myosin Gene-Expression in Mouse Cardiac-Muscle, Journal of Cell Biology 111 (1990) 2427-2436.

DOI: 10.1083/jcb.111.6.2427

[9] H.J. Evans, J.K. Sweet, R.L. Price, M. Yost, R.L. Goodwin, Novel 3D culture system for study of cardiac myocyte development, American Journal of Physiology-Heart and Circulatory Physiology 285 (2003) H570-H578.

DOI: 10.1152/ajpheart.01027.2002

[10] M. Rumpler, A. Woesz, F. Varga, I. Manjubala, K. Klaushofer, P. Fratzl, Three-dimensional growth behavior of osteoblasts on biomimetic hydroxylapatite scaffolds, Journal of Biomedical Materials Research Part A 81A (2007) 40-50.

DOI: 10.1002/jbm.a.30940