• 中國醫(yī)科大學(xué)附屬第一醫(yī)院普通外科教研室器官移植科(遼寧沈陽 110001);

目的  觀察在不同培養(yǎng)模式下大鼠胰腺導(dǎo)管來源干細(xì)胞(PDSC)的生物學(xué)特性。
方法  以大鼠胰腺組織為材料,采用動力性三維培養(yǎng)及傳統(tǒng)二維培養(yǎng)方法進(jìn)行細(xì)胞培養(yǎng),獲得原代PDSC,并連續(xù)傳代,純化PDSC,比較動力性三維培養(yǎng)及傳統(tǒng)二維培養(yǎng)細(xì)胞間連接及細(xì)胞凋亡周期的差異。
結(jié)果  動力性三維培養(yǎng)的大鼠PDSC在培養(yǎng)第2天即可沿三維支架貼壁生長,在第7天即可形成細(xì)胞團(tuán),并沿三維支架多向貼壁生長; 二維培養(yǎng)的PDSC在培養(yǎng)第5天貼壁生長。三維培養(yǎng)與二維培養(yǎng)相比,二維培養(yǎng)的細(xì)胞間連接少見, 三維培養(yǎng)的細(xì)胞連接結(jié)構(gòu)豐富。三維培養(yǎng)與二維培養(yǎng)相比,三維培養(yǎng)的G1期細(xì)胞增多,G2期細(xì)胞相對減少(P<0.01),三維培養(yǎng)早期自發(fā)凋亡細(xì)胞比例相對減少(P<0.05),晚期自發(fā)凋亡與壞死細(xì)胞比例兩種培養(yǎng)方式間的差異無統(tǒng)計(jì)學(xué)意義(P>0.05)。
結(jié)論  動力性三維細(xì)胞培養(yǎng)較傳統(tǒng)二維細(xì)胞培養(yǎng),在細(xì)胞貼壁時間、細(xì)胞連接以及細(xì)胞凋亡方面具有明顯的優(yōu)勢。

引用本文: 陳旭春,程穎,楊蕾,李弘,劉永鋒. 三維及二維培養(yǎng)大鼠胰腺導(dǎo)管來源干細(xì)胞的體外對比實(shí)驗(yàn)研究. 中國普外基礎(chǔ)與臨床雜志, 2012, 19(7): 722-726. doi: 復(fù)制

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9. 黃子星, 劉洋洋, 宋彬. 胰島移植的分子影像示蹤與監(jiān)測 [J].中國普外基礎(chǔ)與臨床雜志, 2012, 19(2):140-145.[ 10] Hori Y, Rulifson IC, Tsai BC, et al. Growth inhibitors promote differentiation of insulin-producing tissue from embryonic stem cells [J]. Proc Natl Acad Sci USA, 2002, 99(25):16105-16110.
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13. Zulewski H, Abraha EJ, Gerlach MJ, et al. Multipotential Nestin-positive stem cells isolated from adult pancreatic islets differentiates ex vivo into pancreatic endocrine, exocrine, and hepatic phenotypes [J]. Diabetes, 2001, 50(3):521-533.
14. Behrous D, Laertis I, Bruce M, et al. Human islet-derived precursor cells are mesenchymal stromal cells that differentiate and mature to hormone-expressing cells in vivo [J]. Stem Cells,2007, 25(12):3215-3222.
15. Petropavlovskaia M, Rosenberg L. Identification and characterization of small cells in the adult pancreas: potential progenitor cells? [J]. Cell Tissue Res, 2002, 310(1):51-58.
16. Seeberger KL, Dufour JM, Shapiro AM, et al. Expansion of mesenchymal stem cells from human pancreatic ductal epithelium [J]. Lab Invest, 2006, 86(2):141-153.
17. Ramiya V, Maraist M, Arfors KE, et al. Reversal of insulindependent diabetes using islets generated in vitro from pancreatic stem cells [J]. Nat Med, 2000, 6(3):278-282.
18. Chase LG, Ulloa-Montoya F, Kidder BL, et al. Islet-derived fibroblast-like cells are not derived via epithelial-mesenchymal transition from Pdx-1 or insulin-positive cells [J]. Diabetes,2007, 56(1):3-7.
19. Giselbrecht S, Gottwald E, Truckenmueller R, et al. Microfabrication of chip-sized scaffolds for three-dimensional cell cultivation [J]. J Vis Exp, 2008, 12(5):691-699.
20. 夏豪, 李庚山, 李建軍, 等. 三維培養(yǎng)內(nèi)皮細(xì)胞形成血管樣結(jié)構(gòu)的實(shí)驗(yàn)研究 [J]. 微循環(huán)學(xué)雜志, 2000, 10(2):5-7.
21. 徐強(qiáng), 胡亞, 廖泉, 等. 胰腺癌細(xì)胞在二維和三維培養(yǎng)系統(tǒng)中的生長特性研究 [J]. 中國普外基礎(chǔ)與臨床雜志, 2011,18(1):33-38.
22. 施森, 何延政, 劉勇, 等. 外周血來源內(nèi)皮祖細(xì)胞三維血管新生模型的建立及其特性分析 [J]. 中國普外基礎(chǔ)與臨床雜志,2009, 16(2):119-123.
23. 王萍, 孫立, 林森森, 等. 內(nèi)皮細(xì)胞三維培養(yǎng)芽生模型的構(gòu)建 [J]. 中國藥科大學(xué)學(xué)報(bào), 2012, 43(1):60-63.
  1. 1. Shapiro AM, Ricordi C, Hering BJ, et al. International trial of the Edmonton protocol for islet transplantation [J]. N Engl J Med, 2006,355(13):1318-1330.
  2. 2. Dorrell C, Grompe MT, Pan FC, et al. Isolation of mouse pancreatic alpha, beta, duct and acinar populations with cell surface markers [J]. Mol Cell Endocrinol, 2011, 339(1-2):144-150.
  3. 3. Li XY, Zhan XR, Lu C, et al. Mechanisms of hepatocyte growth factor-mediated signaling in differentiation of pancreatic ductal epithelial cells into insulin-producing cells [J]. Biochem Biophys Res Commun, 2010, 398(3):389-394.
  4. 4. Pittenger MF, Martin BJ. Mesenchymal stem cells and their potential as cardiac therapeutics [J]. Circ Res, 2004, 95(1):9-20.
  5. 5. Taylor-Fishwick DA, Pittenger GL. Harnessing the pancreatic stem cell [J]. Endocrinol Metab Clin North Am, 2010, 39(4):763-776.
  6. 6. Seo MJ, Suh SY, Bae YC, et al. Differentiation of human adipose stromal cells into hepatic lineage in vitro and in vivo [J].Biochem Biophys Res Commun, 2005, 328(1):258-264.
  7. 7. Peck AB, Yin L, Ramiya V. Animal models to study adult stem cell-derived, in vitro-generated islet implantation [J]. ILAR J,2004, 45(3):259-267.
  8. 8. 黃孝倫. 胰島細(xì)胞移植治療糖尿病的歷史、現(xiàn)狀與未來 [J].中國普外基礎(chǔ)與臨床雜志, 2012, 19(2): 125-128.
  9. 9. 黃子星, 劉洋洋, 宋彬. 胰島移植的分子影像示蹤與監(jiān)測 [J].中國普外基礎(chǔ)與臨床雜志, 2012, 19(2):140-145.[ 10] Hori Y, Rulifson IC, Tsai BC, et al. Growth inhibitors promote differentiation of insulin-producing tissue from embryonic stem cells [J]. Proc Natl Acad Sci USA, 2002, 99(25):16105-16110.
  10. 10. Bonner-Weir S, Weir GC. New sources of pancreatic beta-cells [J]. Nat Biotechnol, 2005, 23(7):857-861.
  11. 11. Grapin-Botton A. Ductal cells of the pancreas [J]. Int J Biochem Cell Biol, 2005, 37(3):504-510.
  12. 12. Lin HT, Chiou SH, Kao CL, et al. Characterization of pancreatic stemcells derived from adult human pancreas ducts by fluorescence activated cell sorting [J]. World J Gastroenterol,2006, 12(28):4529-4535.
  13. 13. Zulewski H, Abraha EJ, Gerlach MJ, et al. Multipotential Nestin-positive stem cells isolated from adult pancreatic islets differentiates ex vivo into pancreatic endocrine, exocrine, and hepatic phenotypes [J]. Diabetes, 2001, 50(3):521-533.
  14. 14. Behrous D, Laertis I, Bruce M, et al. Human islet-derived precursor cells are mesenchymal stromal cells that differentiate and mature to hormone-expressing cells in vivo [J]. Stem Cells,2007, 25(12):3215-3222.
  15. 15. Petropavlovskaia M, Rosenberg L. Identification and characterization of small cells in the adult pancreas: potential progenitor cells? [J]. Cell Tissue Res, 2002, 310(1):51-58.
  16. 16. Seeberger KL, Dufour JM, Shapiro AM, et al. Expansion of mesenchymal stem cells from human pancreatic ductal epithelium [J]. Lab Invest, 2006, 86(2):141-153.
  17. 17. Ramiya V, Maraist M, Arfors KE, et al. Reversal of insulindependent diabetes using islets generated in vitro from pancreatic stem cells [J]. Nat Med, 2000, 6(3):278-282.
  18. 18. Chase LG, Ulloa-Montoya F, Kidder BL, et al. Islet-derived fibroblast-like cells are not derived via epithelial-mesenchymal transition from Pdx-1 or insulin-positive cells [J]. Diabetes,2007, 56(1):3-7.
  19. 19. Giselbrecht S, Gottwald E, Truckenmueller R, et al. Microfabrication of chip-sized scaffolds for three-dimensional cell cultivation [J]. J Vis Exp, 2008, 12(5):691-699.
  20. 20. 夏豪, 李庚山, 李建軍, 等. 三維培養(yǎng)內(nèi)皮細(xì)胞形成血管樣結(jié)構(gòu)的實(shí)驗(yàn)研究 [J]. 微循環(huán)學(xué)雜志, 2000, 10(2):5-7.
  21. 21. 徐強(qiáng), 胡亞, 廖泉, 等. 胰腺癌細(xì)胞在二維和三維培養(yǎng)系統(tǒng)中的生長特性研究 [J]. 中國普外基礎(chǔ)與臨床雜志, 2011,18(1):33-38.
  22. 22. 施森, 何延政, 劉勇, 等. 外周血來源內(nèi)皮祖細(xì)胞三維血管新生模型的建立及其特性分析 [J]. 中國普外基礎(chǔ)與臨床雜志,2009, 16(2):119-123.
  23. 23. 王萍, 孫立, 林森森, 等. 內(nèi)皮細(xì)胞三維培養(yǎng)芽生模型的構(gòu)建 [J]. 中國藥科大學(xué)學(xué)報(bào), 2012, 43(1):60-63.