Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (36): 6681-6684.doi: 10.3969/j.issn.1673-8225.2010.36.007

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Isolation, cultivation and chondrogenic differentiation of adipose-derived stromal cells in vitro

Zhao Da-qing 1,2, Li Qing2, Ma Yu2, Deng Zhi-hong1, Qiao Li1, Lu Lian-jun1, Qiu Jian-hua1   

  1. 1 Department of Otorhinolaryngology-Head and Neck Surgery, 2 Department of Pathology, Xijing Hospital, Fourth Military Medical University of Chinese PLA, Xi’an  710032, Shaanxi Province, China
  • Online:2010-09-03 Published:2010-09-03
  • Contact: Qiu Jian-hua, Professor, Doctoral supervisor, Department of Otorhinolaryngology-Head and Neck Surgery, Xijing Hospital, Fourth Military Medical University of Chinese PLA, Xi’an 710032, Shaanxi Province, China qiujh @fmmu.edu.cn.
  • About author:Zhao Da-qing☆, Doctor, Associate professor, Department of Otorhinolaryngology-Head and Neck Surgery; Department of Pathology, Xijing Hospital, Fourth Military Medical University of Chinese PLA, Xi’an 710032, Shaanxi Province, China zhaodq @fmmu.edu.cn
  • Supported by:

    the National Natural Science Foundation of China, No. 30772261*, 30671087*; Chinese Army Medical Research Council, No. 06MA233*

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Abstract:

BACKGROUND: Tissue engineering has been investigated as a potential means for generation of replacement cartilage. But the ideal seed cell which is still a problem has to be further solved.
OBJECTIVE: To observe the solation and cultivation of adipose-derived stromal cells (ADSCs), and to investigate its chondrogenic differentiation ability in vitro.
METHODS: Subcutaneous fatty tissue from New Zealand rabbits were digested with collagenase. The obtained ADSCs were cultured in the chondrogenic induced medium. Incubation and passage of ADSCs were observed under inverted microscope. The feasibility of ADSCs cartilage induction was detected by immunohistochemistry, Alcian Blue stain, and picrosirius red stain. Proliferation of ADSCs was observed by MTT methods.
RESULTS AND CONCLUSION: ADSCs could be isolated and cultured in vitro. After cultured in chondrogenic medium, the cells differentiated toward cartilage cells and secreted the specific cartilaginous matrices acid mucopolysaccharide and type Ⅱ collagen. Primary cultured ADSCs were spindle-shaped, with oval nucleus. At day 3, ADSCs were in the exponential growth phase, and the population doubling time was about 55 hours. At day 4, cell growth was stable. At day 9, the cells were observed aging symptoms. Cartilage-induced cell proliferation rate increased significantly, while with the incubation time prolonging, cell volume increased significantly, from long fusiform shape into large, multi-polygonal pseudopodia, furthermore nuclei were also larger, and nucleolus was clear. At day 2 after incubation, ADSCs were in the exponential growth phase, and the population doubling time was about 30 hours. At days 8, the cell growth was stable. After the 15th generation, cell proliferation slowed down and cells were observed signs of aging. This suggested that ADSCs differentiated into chondrocytes under a certain degree; therefore, ADSCs could be served as seed cell for cartilaginous tissue engineering.

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