EXPERIMENTAL STUDY ON CHONDROGENIC DIFFERENTIATION OF ADIPOSE-DERIVED STEM CELLS CO-CULTURED WITH CHONDROCYTES_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

XU Haiwei ¹ , XU Baoshan ¹ , YANG Qiang ¹ , LI Xiulan ² , ZHANG Yang ² , ZHANG Chunqiu ³ , WU Yaohong ¹ , CUI Li ²

1Department of Spinal Surgery, 2Cell Engineering Laboratory of Orthopaedic Institute, Tianjin Hospital, Tianjin, 300211, P.R.China;;
3School of Mechanical Engineering, Tianjin University of Technology. Corresponding author: XU Baoshan, E-mail: xubshn@yahoo.com.cn;;
YANG Qiang, E-mail: yangqiang1980@126.com;

Corresponding author：

Keywords：

Adipose-derived stem cells; Chondrocytes; Co-culture; Differentiation; Rabbit

DOI：

10.7507/1002-1892.20130043

Video：

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Objective To observe the chondrogenic differentiation of adipose-derived stem cells (ADSCs) by co-culturing chondrocytes and ADSCs. Methods ADSCs and chondrocytes were isolated and cultured from 8 healthy 4-month-old New Zealand rabbits (male or female, weighing 2.2-2.7 kg). ADSCs and chondrocytes at passage 2 were used. The 1 mL chondrocytes at concentration 2 × 104/mL and 1 mL ADSCs at concentration 2 × 104/mL were seeded on the upper layer and lower layer of Transwell 6-well plates separately in the experimental group, while ADSCs were cultured alone in the control group. The morphology changes of the induced ADSCs were observed by inverted phase contrast microscope. The glycosaminoglycan and collagen type II synthesized by the induced ADSCs were detected with toluidine blue staining and immunohistochemistry staining. The mRNA expressions of collagen type II, aggrecan, and SOX9 were detected with real-time fluorescent quantitative PCR. Results ADSCs in the experimental group gradually became chondrocytes-like in morphology and manifested as round; while ADSCs in the control group manifested as long spindle in morphology with whirlool growth pattern. At 14 days after co-culturing, the results of toluidine blue staining and immunohistochemistry staining were positive in the experimental group, while the results were negative in the control group. The results of real-time fluorescent quantitative PCR indicated that the expression levels of collagen type II, aggrecan, and SOX9 mRNA in the experimental group (1.43 ± 0.07, 2.13 ± 0.08, and 1.08 ± 0.08) were significantly higher than those in the control group (0.04 ± 0.03, 0.13 ± 0.04, and 0.10 ± 0.02) (P lt; 0.05). Conclusion ADSCs can differentiate into chondrocytes-like after co-culturing with chondrocytes.

Citation： XU Haiwei,XU Baoshan,YANG Qiang,LI Xiulan,ZHANG Yang,ZHANG Chunqiu,WU Yaohong,CUI Li. EXPERIMENTAL STUDY ON CHONDROGENIC DIFFERENTIATION OF ADIPOSE-DERIVED STEM CELLS CO-CULTURED WITH CHONDROCYTES. Chinese Journal of Reparative and Reconstructive Surgery, 2013, 27(2): 193-198. doi: 10.7507/1002-1892.20130043 Copy

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2.	Khan WS, Tew SR, Adesida AB, et al. Human infrapatellar fat pad-derived stem cells express the pericyte marker 3G5 and show enhanced chondrogenesis after expansion in fibroblast growth factor-2. Arthritis Res Ther, 2008, 10(4): R74.
3.	Merceron C, Vinatier C, Portron S, et al. Differential effects of hypoxia on osteochondrogenic potential of human adipose-derived stem cells. Am J Physiol Cell Physiol, 2010, 298(2): C355-364.
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7.	Fan J, Gong Y, Ren L, et al. In vitro engineered cartilage using synovium-derived mesenchymal stem cells with injectable gellan hydrogels. Acta Biomater, 2010, 6(3): 1178-1185.
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14.	Hildner F, Albrecht C, Gabriel C, et al. State of the art and future perspectives of articular cartilage regeneration: a focus on adipose-derived stem cells and platelet-derived products. J Tissue Eng Regen Med, 2011, 5(4): e36-51.
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17.	Li TZ, Kim JH, Cho HH, et al. Therapeutic potential of bone-marrow-derived mesenchymal stem cells differentiated with growth-factor-free coculture method in liver-injured rats. Tissue Eng Part A, 2010, 16(8): 2649-2659.
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19.	徐志刚. 脂肪间充质干细胞与自体髓核细胞共培养中相互作用分子机制初步研究. 武汉: 华中科技大学, 2010.
20.	张洁, 蒋海越, 何乐仁, 等. 残耳软骨细胞与脂肪干细胞共培养体内构建软骨的实验研究. 组织工程与重建外科杂志, 2011, 7(2): 75-79.

1. Feng G, Wan Y, Balian G, et al. Adenovirus-mediated expression of growth and differentiation factor-5 promotes chondrogenesis of adipose stem cells. Growth Factors, 2008, 26(3): 132-142.
2. Khan WS, Tew SR, Adesida AB, et al. Human infrapatellar fat pad-derived stem cells express the pericyte marker 3G5 and show enhanced chondrogenesis after expansion in fibroblast growth factor-2. Arthritis Res Ther, 2008, 10(4): R74.
3. Merceron C, Vinatier C, Portron S, et al. Differential effects of hypoxia on osteochondrogenic potential of human adipose-derived stem cells. Am J Physiol Cell Physiol, 2010, 298(2): C355-364.
4. Hamid AA, Idrus RB, Saim AB, et al. Characterization of human adipose-derived stem cells and expression of chondrogenic genes during induction of cartilage differentiation. Clinics (Sao Paulo), 2012, 67(2): 99-106.
5. Diekman BO, Rowland CR, Lennon DP, et al. Chondrogenesis of adult stem cells from adipose tissue and bone marrow: induction by growth factors and cartilage-derived matrix. Tissue Eng Part A, 2010, 16(2): 523-533.
6. Li JW, Guo XL, He CL, et al. In vitro chondrogenesis of the goat bone marrow mesenchymal stem cells directed by chondrocytes in monolayer and 3-dimetional indirect co-culture system. Chin Med J (Engl), 2011, 124(19): 3080-3086.
7. Fan J, Gong Y, Ren L, et al. In vitro engineered cartilage using synovium-derived mesenchymal stem cells with injectable gellan hydrogels. Acta Biomater, 2010, 6(3): 1178-1185.
8. Jiang J, Li J, Hao X, et al. Use of autologous chondrocytes and bioinert perforated chambers to tissue engineer cartilage in vivo. J Surg Res, 2012, 173(1): e27-32.
9. Vasiliadis HS, Lindahl A, Georgoulis AD, et al. Malalignment and cartilage lesions in the patellofemoral joint treated with autologous chondrocyte implantation. Knee Surg Sports Traumatol Arthrosc, 2011, 19(3): 452-457.
10. Dai J, Wang X, Shen G. Cotransplantation of autologous bone marrow stromal cells and chondrocytes as a novel therapy for reconstruction of condylar cartilage. Med Hypotheses, 2011, 77(1): 132-133.
11. 杨强, 彭江, 卢世璧, 等. 软骨脱细胞基质多孔支架与骨髓基质干细胞体外构建组织工程软骨的研究. 中华医学杂志, 2011, 91(17): 1161-1166.
12. Dhinsa BS, Adesida AB. Current clinical therapies for cartilage repair, their limitation and the role of stem cells. Curr Stem Cell Res Ther, 2012, 7(2): 143-148.
13. 聂绪强, 陈怀红, 唐宁, 等. 脂肪源性干细胞研究及其应用进展. 中国修复重建外科杂志, 2011, 25(7): 854-858.
14. Hildner F, Albrecht C, Gabriel C, et al. State of the art and future perspectives of articular cartilage regeneration: a focus on adipose-derived stem cells and platelet-derived products. J Tissue Eng Regen Med, 2011, 5(4): e36-51.
15. 张明, 周哲, 周娟, 等. 人脂肪干细胞与猪尿路上皮细胞隔离共培养后向尿路上皮样细胞分化. 中华临床医师杂志(电子版), 2012, 6(8): 109-112.
16. Chao KC, Chao KF, Fu YS, et al. Islet-like clusters derived from mesenchymal stem cells in Wharton’s Jelly of the human umbilical cord for transplantation to control type 1 diabetes. PLoS One, 2008, 3(1): e1451.
17. Li TZ, Kim JH, Cho HH, et al. Therapeutic potential of bone-marrow-derived mesenchymal stem cells differentiated with growth-factor-free coculture method in liver-injured rats. Tissue Eng Part A, 2010, 16(8): 2649-2659.
18. 于鹏. 兔骨髓间充质干细胞与髓核细胞共培养体系的实验研究. 武汉: 华中科技大学, 2010.
19. 徐志刚. 脂肪间充质干细胞与自体髓核细胞共培养中相互作用分子机制初步研究. 武汉: 华中科技大学, 2010.
20. 张洁, 蒋海越, 何乐仁, 等. 残耳软骨细胞与脂肪干细胞共培养体内构建软骨的实验研究. 组织工程与重建外科杂志, 2011, 7(2): 75-79.

Chinese Journal of Reparative and Reconstructive Surgery

EXPERIMENTAL STUDY ON CHONDROGENIC DIFFERENTIATION OF ADIPOSE-DERIVED STEM CELLS CO-CULTURED WITH CHONDROCYTES

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