Sains Malaysiana 50(7)(2021): 1987-1996
http://doi.org/10.17576/jsm-2021-5007-13
The Combination of bFGF and
Hydrocortisone is a Better Alternative Compared to 5-Azacytidine for
Cardiomyogenic Differentiation of Bone Marrow and Adipose Stem Cells
(Gabungan bFGF dan Hidrokortison adalah Alternatif yang Lebih Baik Berbanding dengan 5-Azasitidin bagi Perbezaan Kardomiogen Sumsum Tulang dan Sel Stem Adipos)
NADIAH
SULAIMAN1*, NUR QISYA AFIFAH VERONICA SAINIK1,2, SHAMSUL
BIN SULAIMAN1, PEZHMAN HAFEZ1, NG MIN HWEI1 & RUSZYMAH BT HJ IDRUS1,2
1Tissue Engineering Centre, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Yaacob Latiff, Bandar Tun Razak, 56000 Cheras, Kuala
Lumpur, Federal Territory, Malaysia
2Department of Physiology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Yaacob Latiff, Bandar Tun Razak, 56000 Cheras, Kuala
Lumpur, Federal Territory, Malaysia
Received:
27 April 2020/Accepted: 13 November 2020
ABSTRACT
Stem cells can be differentiated into
cardiomyocytes by induction with 5-azacytidine (5-aza) but its carcinogenicity
is of concern for future translational application. Alternatively, growth
factors and hormones such as basic fibroblast growth factor (bFGF) and hydrocortisone have been reported to act as a
therapeutic inducer for cardiomyocytes differentiation. In this study, we aim
to investigate the ability of bFGF and hydrocortisone
in combination to stimulate the differentiation of mesenchymal stem cells (MSC)
into cardiomyocytes lineage. Sheep adipose tissue stem cell (ATSC) and bone
marrow stem cell (BMSC) were isolated, cultured and induced with the three
groups of induction factors; 5-aza alone, the combination of hydrocortisone and bFGF and all three factors in combination for
cardiomyogenic differentiation. Morphological, protein and functional ability
of both ATSC and BMSC were observed and analysed to
confirm cardiomyocyte differentiation. Viability of BMSC and ATSC in each
treated group was significantly higher (P < 0.05) on both cells after
treated with 10 nM of bFGF and 50 μM of hydrocortisone. Cardiomyocyte
proteins; α-Sarcomeric actin (αSA) and Phospolamban (Plb) was detected
in both ATSC and BMSC exposed to induction factors but not in the control
negative group. Both ATSC and BMSC without induction factors showed only minute
cell number possesses αSA and Plb. Calcium ion
(Ca2+) spark was observed in primary heart cells. Similarly, Ca2+ spark was also detected in induced ATSC and BMSC, proving some functionality of
induced cells. In conclusion, bFGF and hydrocortisone
are safer induction factor compared to the currently used 5-aza as both showed
higher viability after induction, therefore more cells are available for future
use in cardiac tissue engineering.
Keywords: 5-Azacytidine; basic Fibroblast Growth Factor; cardiomyocytes
differentiation; hydrocortisone; stem cells
ABSTRAK
Sel induk boleh
dibezakan menjadi kardiomiosit dengan aruhan 5-azasitidin (5-aza) tetapi sifat
karsinogeniknya menimbulkan kerisauan bagi kegunaan klinikal pada masa hadapan.
Sebagai alternatif, faktor pertumbuhan dan pelbagai jenis hormon seperti faktor
pertumbuhan fibroblas asas (bFGF) dan hidrokortison dilaporkan boleh bertindak
sebagai pemacu terapi untuk pembezaan kardiomiosit. Kajian ini bertujuan untuk
mengkaji kemampuan bFGF dan hidrokortison secara gabungan untuk merangsang
pembezaan MSC kepada leluhur kardiomiosit. Sel dasar lemak (ATSC) dan tulang
sum-sum kambing (BMSC) diasingkan, dikultur dan diaruh dengan tiga kumpulan
faktor aruhan; 5-aza sahaja, gabungan hidrokortison dan bFGF dan ketiga-tiga
faktor gabungan untuk pembezaan kardiomogenik. Perubahan morfologi, protein dan
fungsi kedua-dua ATSC dan BMSC dikaji dan dianalisis untuk mengesahkan
pembezaan leluhur kardiomiosit. Perkembangan ATSC dan BMSC pada setiap kumpulan
yang dirawat jauh lebih tinggi (P <0.05) pada kedua-dua sel setelah dirawat
dengan 10 nM bFGF dan 50 μM hidrokortison. Protein kardiomiosit,
α-Sarcomeric actin (αSA) dan Phospolamban (Plb) dikesan pada
kedua-dua ATSC dan BMSC yang terdedah kepada faktor aruhan tetapi tidak dalam
kawalan negatif. Kedua-dua sel, ATSC dan BMSC tanpa faktor aruhan menunjukkan
hanya sebilangan kecil sel mempunyai αSA dan Plb. Percikan ion kalsium (Ca2
+) diperhatikan pada sel jantung primer, yang turut dikesan pada ATSC dan
BMSC yang diinduksi. Maka, sel yang diinduksi sedikit sebanyak berfungsi
seperti kardiomiosit. Kesimpulannya, bFGF dan hidrokortison adalah faktor
aruhan yang lebih selamat berbanding dengan 5-Aza yang digunakan sekarang. Hal
ini demikian adalah kerana kedua-duanya menunjukkan perkembangan yang lebih
tinggi selepas aruhan, oleh itu lebih banyak sel tersedia untuk kegunaan pada
peringkat klinikal.
Kata kunci:
5-azasitidin; faktor tumbesaran fibroblas asas; hidrokortison; pembezaan
kardiomiosit; sel dasar
REFERENCES
Akin, B.L., Hurley, T.D., Chen, Z. & Jones, L.R. 2013.
The structural basis for phospholamban inhibition of the calcium pump in
sarcoplasmic reticulum. Journal of Biological Chemistry 288(42): 30181-30191.
Bird, S.D., Doevendans, P. A., van Rooijen, M.A., de la
Riviere, A.B., Hassink, R.J., Passier, R. & Mummery, C.L. 2003. The human
adult cardiomyocyte phenotype. Cardiovascular Research 58(2): 423-434.
Brade, T., Pane, L.S., Moretti, A., Chien, K.R. &
Laugwitz, K.L. 2013. Embryonic heart progenitors and cardiogenesis. Cold
Spring Harbor Perspectives in Medicine 3(10): a013847.
Choi, Y.S., Dusting, G.J., Stubbs, S., Arunothayaraj, S.,
Han, X.L., Collas, P., Morrison, W.A. & Dilley, R.J. 2010. Differentiation
of human adipose-derived stem cells into beating cardiomyocytes. Journal of
Cellular and Molecular Medicine 14(4): 878-889.
Clifford, D.M., Fisher, S.A., Brunskill, S.J., Doree, C.,
Mathur, A., Watt, S. & Martin-Rendon, E. 2012. Stem cell treatment for
acute myocardial infarction. Cochrane Database of Systematic Reviews (2): CD006536.
Colenci, R., da Silva Assunção, L.R., Bomfim, S.R.M., de
Assis Golim, M., Deffune, E. & Oliveira, S.H.P. 2014. Bone marrow
mesenchymal stem cells stimulated by bFGF up-regulated protein expression in
comparison with periodontal fibroblasts in
vitro. Archives of Oral Biology 59(3): 268-276.
Giugliano, G.R., Giugliano, R.P., Michael Gibson, C. &
Kuntz, R.E. 2003. Meta-analysis of corticosteroid treatment in acute myocardial
infarction. American Journal of Cardiology 91(9): 1055-1059.
Guo, X., Bai, Y., Zhang, L., Zhang, B., Zagidullin, N., Carvalho,
K., Du, Z. & Cai, B. 2018. Cardiomyocyte differentiation of mesenchymal
stem cells from bone marrow: New regulators and its implications. Stem Cell
Research and Therapy 9(1): 44.
Kaptoge, S., Pennells, L., De Bacquer, D., Cooney, M.T., Kavousi, M., Stevens, G., Riley, L.M., Savin, S., Khan, T., Altay, S., Amouyel, P., Assmann, G., Bell, S., Ben-Shlomo, Y., Berkman, L., Beulens, J.W., Björkelund, C., Blaha, M., Blazer, D.G., Bolton, T., Beaglehole, R.B., Brenner, H., Brunner, E.J., Casiglia, E., Chamnan, P., Choi, Y-H., Chowdry, R., Coady, S., Crespo, C.J., Cushman, M., Dagenais, G.R., D'Agostino Sr., R.B., Daimon, M., Davidson, K.W., Engström, G., Ford, I., Gallacher, J., Gansevoort, R.T., Gaziano, T.A., Giampaoli, S., Grandits, G., Grimsgaard, S., Grobbee, D.E., Gudnason, V., Guo, Q., Tolonen, H., Humphries, S., Iso, H., Jukema, J.W., Kauhanen, J., Kengne, A.P., Khalili, D., Koenig, W., Kromhout, D., Krumholz, H., Lam, T.H., Laughlin, G., Ibañez, A.M., Meade, T.W., Moons, K.G.M., Nietert, P.J., Ninomiya, T., Nordestgaard, B.G., O'Donnell, C., Palmieri, L., Patel, A., Perel, P., Price, J.F., Providencia, R., Ridker, P.M., Rodriguez, B., Rosengren, A., Roussel, R., Sakurai, M., Salomaa, V., Sato, S., Schöttker, B., Shara, N., Shaw, J.E., Shin, H-C., Simons, L.A., Sofianopoulou, E., Sundström, J., Völzke, H., Wallace, R.B., Wareham, N.J., Willeit, P., Wood, D., Wood, A., Zhao, D., Woodward, M., Danaei, G., Roth, G., Mendis, S., Onuma, O., Varghese, C., Ezzati, M., Graham, I., Jackson, R., Danesh, J., Angelantonio, E.D. 2019. World Health
Organization cardiovascular disease risk charts: Revised models to estimate
risk in 21 global regions. The Lancet Global Health 7(10): e1332-e1345.
Kawai, T., Takahashi, T., Esaki, M., Ushikoshi, H., Nagano,
S., Fujiwara, H. & Kosai, K.I. 2004. Efficient cardiomyogenic
differentiation of embryonic stem cell by fibroblast growth factor 2 and bone
morphogenetic protein 2. Circulation Journal 68(7): 691-702.
Kelecsényi, Z., Spencer, D.L. & Caspary, W.J. 2000.
Molecular analysis of 5-azacytidine-induced variants in mammalian cells. Mutagenesis 15(1): 25-31.
Knot, H.J., Laher, I., Sobie, E.A., Guatimosim, S.,
Gomez-Viquez, L., Hartmann, H., Song, L.S., Lederer, W.J., Graier, W.F., Malli,
R., Frieden, M. & Petersen, O.H. 2005. Twenty years of calcium imaging:
Cell physiology to dye for. Molecular Interventions 5(2): 112-127.
Kuhn, N.Z. & Tuan, R.S. 2010. Regulation of stemness and
stem cell niche of mesenchymal stem cells: Implications in tumorigenesis and
metastasis. Journal of Cellular Physiology 222(2): 268-277.
Leon, B.M. 2015. Diabetes and cardiovascular disease:
Epidemiology, biological mechanisms, treatment recommendations and future
research. World Journal of Diabetes 6(13): 1246-1258.
Li, K., Li, S.Z., Zhang, Y.L. & Wang, X.Z. 2011. The effects
of dan-shen root on cardiomyogenic differentiation of human placenta-derived
mesenchymal stem cells. Biochemical and Biophysical Research Communications 415(1): 147-151.
Li, W., Wu, D., Niu, Z., Jiang, D., Ma, H., He, H., Zuo, X.,
Xie, X. & He, Y. 2016. 5-azacytidine suppresses EC9706 cell proliferation
and metastasis by upregulating the expression of SOX17 and CDH1. International
Journal of Molecular Medicine 38(4): 1047-1054.
Loughrey, C.M., MacEachern, K.E., Neary, P. & Smith, G.L.
2002. The relationship between intracellular [Ca(2+)] and Ca(2+) wave
characteristics in permeabilised cardiomyocytes from the rabbit. The Journal
of Physiology 543(3): 859-870.
Ludman, P.F. 2018. Percutaneous coronary intervention. Medicine
(United Kingdom) 46(9): 547-554.
MacLennan, D.H. & Kranias, E.G. 2003. Phospholamban: A
crucial regulator of cardiac contractility. Nature Reviews Molecular Cell
Biology 4: 566-577.
Marques, S.R., Lee, Y., Poss, K.D. & Yelon, D. 2008.
Reiterative roles for FGF signaling in the establishment of size and proportion
of the zebrafish heart. Developmental Biology 321(2): 397-406.
Ogawa, R., Akita, S., Akaishi, S., Aramaki-Hattori, N., Dohi,
T., Hayashi, T., Kishi, K., Kono, T., Matsumura, H., Muneuchi, G., Murao, N.,
Nagao, M., Okabe, K., Shimizu, F., Tosa, M., Tosa, Y., Yamawaki, S., Ansai, S.,
Inazu, N., Kamo, T., Kazki, R. & Kuribayashi, S. 2019. Diagnosis and
treatment of keloids and hypertrophic scars - Japan scar workshop consensus
document 2018. Burns and Trauma 7: 39.
Olesen, C., Picard, M., Winther, A.M.L., Gyrup, C., Preben
Morth, J., Oxvig, C., Vuust Møller, J. & Nissen, P. 2007. The structural
basis of calcium transport by the calcium pump. Nature 450: 1036-1042.
Peter, A.K., Bjerke, M.A. & Leinwand, L.A. 2016. Biology
of the cardiac myocyte in heart disease. Molecular Biology of the Cell 27(14): 2149-2160.
Pittenger, M.F. & Martin, B.J. 2004. Mesenchymal stem
cells and their potential as cardiac therapeutics. Circulation Research 95(1):
9-20.
Segers, V.F.M. & Lee, R.T. 2008. Stem-cell therapy for
cardiac disease. Nature 451: 937-942.
Shipunova, N.N., Petinati, N.A. & Drize, N.I. 2013.
Effect of hydrocortisone on multipotent human mesenchymal stromal cells. Bulletin
of Experimental Biology and Medicine 155(1): 159-163.
Steinhauser, M.L. & Lee, R.T. 2011. Regeneration of the
heart. EMBO Molecular Medicine 3(12): 701-712.
Stern-Straeter, J., Bonaterra, G.A., Juritz, S., Birk, R.,
Goessler, U.R., Bieback, K., Bugert, P., Schultz, J., Hörmann, K., Kinscherf,
R. & Faber, A. 2014. Evaluation of the effects of different culture media
on the myogenic differentiation potential of adipose tissue- or bone
marrow-derived human mesenchymal stem cells. International Journal of
Molecular Medicine 33(1): 160-170.
Yin, Z., Ren, J. & Guo, W. 2015. Sarcomeric protein
isoform transitions in cardiac muscle: A journey to heart failure. Biochimica
et Biophysica Acta - Molecular Basis of Disease 1852(1): 47-52.
Yoon, Y.S., Park, J.S., Tkebuchava, T., Luedeman, C. &
Losordo, D.W. 2004. Unexpected severe calcification after transplantation of
bone marrow cells in acute myocardial infarction. Circulation 109(25):
3154-3157.
Zhang, Y., Mignone, J. & Robb Maclellan, W. 2015. Cardiac
regeneration and stem cells. Physiological Reviews 95(4): 1189-1204.
Zhao, Y., Li, T., Wei, X., Bianchi, G., Hu, J., Sanchez,
P.G., Xu, K., Zhang, P., Pittenger, M.F., Wu, Z.J. & Griffith, B.P. 2012.
Mesenchymal stem cell transplantation improves regional cardiac remodeling
following ovine infarction. STEM CELLS Translational Medicine 1(9):
685-695.
*Corresponding
author; email: nadiahsulaiman@ukm.edu.my
|