• 1 華中科技大學(xué)同濟(jì)醫(yī)學(xué)院附屬同濟(jì)醫(yī)院腫瘤科（武漢，430030）;;
  • 中國(guó)醫(yī)學(xué)科學(xué)院 北京協(xié)和醫(yī)學(xué)院北京協(xié)和醫(yī)院 2 血液內(nèi)科，3 心內(nèi)科;

目的 分析hBMSCs 經(jīng)5- 氮雜胞苷（5-azacytidine，5-aza）誘導(dǎo)向心肌樣細(xì)胞分化過(guò)程中基因表達(dá)譜
的改變。 方法 取胸外科非血液病患者手術(shù)中廢棄的肋骨骨髓分離培養(yǎng)hBMSCs，5-aza 誘導(dǎo)第2 代hBMSCs。取誘導(dǎo)
前及誘導(dǎo)后的細(xì)胞，免疫細(xì)胞化學(xué)法檢測(cè)α-actin、肌鈣蛋白T（cardiac troponin T，cTnT）和連接蛋白43（connexin 43）表
達(dá)，流式細(xì)胞儀檢測(cè)cTnT 陽(yáng)性細(xì)胞百分率；利用人表達(dá)譜基因芯片技術(shù)篩選分化過(guò)程中的差異表達(dá)基因，對(duì)部分基因進(jìn)
行功能分類和分層聚類分析。 結(jié)果 hBMSCs 經(jīng)5-aza 誘導(dǎo)后，部分呈肌細(xì)胞樣形態(tài)。免疫細(xì)胞化學(xué)染色檢測(cè)示誘導(dǎo)
前細(xì)胞α-actin、cTnT 染色呈弱陽(yáng)性，connexin 43 染色呈陰性；誘導(dǎo)后3 周細(xì)胞α-actin、cTnT、connexin 43 染色均呈陽(yáng)性。
流式細(xì)胞儀檢測(cè)示誘導(dǎo)前cTnT 陽(yáng)性細(xì)胞百分率為7.43% ± 0.02%，誘導(dǎo)后3 周為49.64% ± 0.05%。分化過(guò)程中共檢測(cè)到
1 814 個(gè)顯著差異表達(dá)基因，對(duì)其中647 個(gè)基因分層聚類，聚為5 類，生物功能包括信號(hào)傳導(dǎo)、細(xì)胞代謝、增殖分化、發(fā)育以
及形態(tài)發(fā)生等。 結(jié)論 hBMSCs 經(jīng)5-aza 誘導(dǎo)向心肌樣細(xì)胞分化過(guò)程受信號(hào)傳導(dǎo)通路、轉(zhuǎn)錄基因、生長(zhǎng)因子等多種因素
在不同時(shí)間點(diǎn)上的共同調(diào)控。

引用本文： 楊琳 ,沈悌,陳連鳳,曹欣欣,賴晉智. BMSCs 向心肌分化的基因表達(dá)譜分析. 中國(guó)修復(fù)重建外科雜志, 2012, 26(5): 607-611. doi: 復(fù)制

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2. Wollert KC, Meyer GP, Lotz J, et al. Intracoronary autologous bone-marrow cell transfer after myocardial infarction: the BOOST randomised controlled clinical trial. Lancet, 2004, 364(9429): 141-148.
3. Britten MB, Abolmaali ND, Assmus B, et al. Infarct remodeling after intracoronary progenitor cell treatment in patients with acute myocardial infarction (TOPCARE-AMI): mechanistic insights from serial contrast-enhanced magnetic resonance imaging. Circulation, 2003, 108(18): 2212-2218.
4. Dimmeler S, Zeiher AM, Schneider MD. Unchain my heart: the scientific foundations of cardiac repair. J Clin Invest, 2005, 115(3): 572-583.
5. Ling SK, Wang R, Dai ZQ, et al. Pretreatment of rat bone marrow mesenchymal stem cells with a combination of hypergravity and 5-azacytidine enhances therapeutic efficacy for myocardial infarction. Biotechnol Prog, 2011, 27(2): 473-482.
6. Xu W, Zhang X, Qian H, et al. Mesenchymal stem cells from adult human bone marrow differentiates into a cardiomyocyte phenotype in vitro. Exp Biol Medi (Maywood), 2004, 229(7): 623-631.
7. Makino S, Fukuda K, Miyoshi S, et al. Cardiomyocytes can be generated from marrow stromal cells in vitro. J Clin Invest, 1999, 103(5): 697-705.
8. Cohen ED, Tian Y, Morrisey EE. Wnt signaling: an essential regulator of cardiovascular differentiation, morphogenesis and progenitor self-renewal. Development, 2008, 135(5): 789-798.
9. Flaherty MP, Abdel-Latif A, Li Q, et al. Noncanonical Wnt11 signaling is sufficient to induce cardiomyogenic differentiation in unfractionated bone marrow mononuclear cells. Circulation, 2008, 117(17): 2241-2252.
10. Qyang Y, Martin-Puig S, Chiravuri M, et al. The renewal and differentiation of isl1+ cardiovascular progenitors are controlled by a Wnt/beta-catenin pathway. Cell Stem Cell, 2007, 1(2): 165-179.
11. Zhu W, Shiojima I, Ito Y, et al. IGFBP-4 is an inhibitor of canonical Wnt signalling required for cardiogenesis. Nature, 2008, 454(7202): 345-349.
12. Schneider VA, Mercola M. Wnt antagonism initiates cardiogenesis in Xenopus laevis. Genes Dev, 2001, 15(3): 304-315.
13. Tzahor E, Lassar AB. Wnt signals from the neural tube block ectopic cardiogenesis. Genes Dev, 2001, 15(3): 255-260.
14. Li Y, Hiroi Y, Ngoy S, et al. Notch1 in bone marrow-derived cells mediates cardiac repair after myocardial infarction. Circulation, 2011, 123(8): 866-876.
15. Crispino JD, Lodish MB, Thurberg BL, et al. Proper coronary vascular development and heart morphogenesis depend on interaction of GATA-4 with FOG cofactors. Genes Dev, 2001, 15(7): 839-844.
16. Zhao R, Watt AJ, Battle MA, et al. Loss of both GATA4 and GATA6 blocks cardiac myocyte differentiation and results in acardia in mice. Dev Biol, 2008, 317(2): 614-619.
17. Sachinidis A, Kolossov E, Fleischmann BK, et al. Generation of cardiomyocytes from embryonic stem cells experimental studies. Herz, 2002, 27(7): 589-597.
18. Yang HS, Bhang SH, Kim IK, et al. In situ cardiomyogenic differentiation of implanted bone marrow mononuclear cells by local delivery of transforming growth factor-β1. Cell Transplant, 2011. [Epub ahead of print].
19. Wan CR, Chung S, Kamm RD. Differentiation of embryonic stem cells into cardiomyocytes in a compliant microfluidic system. Ann Biomed Eng, 2011, 39(6): 1840-1847.
20. Afouda BA, Hoppler S. Different requirements for GATA factors in cardiogenesis are mediated by non-canonical Wnt signaling. Dev Dyn, 2011, 240(3): 649-662.
  1. 1. Passier R, van Laake LW, Mummery CL. Stem-cell-based therapy and lessons from the heart. Nature, 2008, 453(7193): 322-329.
  2. 2. Wollert KC, Meyer GP, Lotz J, et al. Intracoronary autologous bone-marrow cell transfer after myocardial infarction: the BOOST randomised controlled clinical trial. Lancet, 2004, 364(9429): 141-148.
  3. 3. Britten MB, Abolmaali ND, Assmus B, et al. Infarct remodeling after intracoronary progenitor cell treatment in patients with acute myocardial infarction (TOPCARE-AMI): mechanistic insights from serial contrast-enhanced magnetic resonance imaging. Circulation, 2003, 108(18): 2212-2218.
  4. 4. Dimmeler S, Zeiher AM, Schneider MD. Unchain my heart: the scientific foundations of cardiac repair. J Clin Invest, 2005, 115(3): 572-583.
  5. 5. Ling SK, Wang R, Dai ZQ, et al. Pretreatment of rat bone marrow mesenchymal stem cells with a combination of hypergravity and 5-azacytidine enhances therapeutic efficacy for myocardial infarction. Biotechnol Prog, 2011, 27(2): 473-482.
  6. 6. Xu W, Zhang X, Qian H, et al. Mesenchymal stem cells from adult human bone marrow differentiates into a cardiomyocyte phenotype in vitro. Exp Biol Medi (Maywood), 2004, 229(7): 623-631.
  7. 7. Makino S, Fukuda K, Miyoshi S, et al. Cardiomyocytes can be generated from marrow stromal cells in vitro. J Clin Invest, 1999, 103(5): 697-705.
  8. 8. Cohen ED, Tian Y, Morrisey EE. Wnt signaling: an essential regulator of cardiovascular differentiation, morphogenesis and progenitor self-renewal. Development, 2008, 135(5): 789-798.
  9. 9. Flaherty MP, Abdel-Latif A, Li Q, et al. Noncanonical Wnt11 signaling is sufficient to induce cardiomyogenic differentiation in unfractionated bone marrow mononuclear cells. Circulation, 2008, 117(17): 2241-2252.
  10. 10. Qyang Y, Martin-Puig S, Chiravuri M, et al. The renewal and differentiation of isl1+ cardiovascular progenitors are controlled by a Wnt/beta-catenin pathway. Cell Stem Cell, 2007, 1(2): 165-179.
  11. 11. Zhu W, Shiojima I, Ito Y, et al. IGFBP-4 is an inhibitor of canonical Wnt signalling required for cardiogenesis. Nature, 2008, 454(7202): 345-349.
  12. 12. Schneider VA, Mercola M. Wnt antagonism initiates cardiogenesis in Xenopus laevis. Genes Dev, 2001, 15(3): 304-315.
  13. 13. Tzahor E, Lassar AB. Wnt signals from the neural tube block ectopic cardiogenesis. Genes Dev, 2001, 15(3): 255-260.
  14. 14. Li Y, Hiroi Y, Ngoy S, et al. Notch1 in bone marrow-derived cells mediates cardiac repair after myocardial infarction. Circulation, 2011, 123(8): 866-876.
  15. 15. Crispino JD, Lodish MB, Thurberg BL, et al. Proper coronary vascular development and heart morphogenesis depend on interaction of GATA-4 with FOG cofactors. Genes Dev, 2001, 15(7): 839-844.
  16. 16. Zhao R, Watt AJ, Battle MA, et al. Loss of both GATA4 and GATA6 blocks cardiac myocyte differentiation and results in acardia in mice. Dev Biol, 2008, 317(2): 614-619.
  17. 17. Sachinidis A, Kolossov E, Fleischmann BK, et al. Generation of cardiomyocytes from embryonic stem cells experimental studies. Herz, 2002, 27(7): 589-597.
  18. 18. Yang HS, Bhang SH, Kim IK, et al. In situ cardiomyogenic differentiation of implanted bone marrow mononuclear cells by local delivery of transforming growth factor-β1. Cell Transplant, 2011. [Epub ahead of print].
  19. 19. Wan CR, Chung S, Kamm RD. Differentiation of embryonic stem cells into cardiomyocytes in a compliant microfluidic system. Ann Biomed Eng, 2011, 39(6): 1840-1847.
  20. 20. Afouda BA, Hoppler S. Different requirements for GATA factors in cardiogenesis are mediated by non-canonical Wnt signaling. Dev Dyn, 2011, 240(3): 649-662.