Research On Isolation, Culture And Differentiation Potential Of Beijing Fatty Chicken Adipose-derived Stem Cells

In the 90th of 20 century, the world set off an upsurge of stem cell research, more and more researcheres payed their attention to the proliferation and differentiation characteristics of stem cells. Because of the unique advantages, the research value of adipose-derived stem cells (ADSCs) is gradually exposed in the field of tissue engineering. In this study, we isolated ADSCs from adipose tissues of Beijing fatty chicken, and optimized the culture conditions. We also explored the biological characteristics and the differentiation potency of ADSCs. The results were following:1. The ADSCs were isolated from 1-day old Beijing fatty chicken adipose tissues by 0.1% collagenase digestion. The suitable culture condition for ADSCs was DMEM/F-12 + 10% FBS + 10 ng/ml bFGF + 1% B-27 + 2 mM glutamine + 100 IU/ml penicillin + 100μg/ml streptomycin.2. The ADSCs expanded easily and exhibited a fibroblast-like morphplogy. In primary culture, many hemocytes mixed with ADSCs, after 3-4 passages, the hemocytes were detached and the cells were purified. The growth curves of different passages all showed typical"S"shape. The cell growth experienced the processes of latentperiod, logarithmic phase and plateau phase. The cells were sub-cultured to the 15th passage. Before the 10th passage, cells maintained stable morphological characteristics and multiplication capacity. From the 12th passage, the intracellular vacuoles could be observed. The cells sub-cultured to the 15th passage showed representative senescent appearance, and could not be sub-culture anymore.3. We studied the influences of freezing conservation and recovery on cell viability. The results showed that cryopreservation did not affect cell viability in P1 and P3 cells (P > 0.05) but would decrease the viability in P5 and P7 cells (0.01 < P < 0.05), which demonstrated that the cell passage should be considered and the composition of frozen stock solution should be optimized.4. We detected the surface antigen markers of ADSCs by immunofluorescence staining and RT-PCR experiment. The results showed that ADSCs expressed the MSCs markers of CD29, CD44, CD71 and CD73 while did not express endothelial cells specific marker CD 31.5. Under the appropriate effect of inducing factors and time, Beijing fatty chicken ADSCs could be induced to differentiate into osteoblasts, adipocytes, cardiac myocytes and even neurocytesTo sum up, the ADSCs isolated from Beijing fatty chicken adipose tisseus had a high reproductive capacity and plasticity in vitro. In view of the easy to obtain, result in minimal patient discomfort, and yet be capable of yielding cells substantial enough, ADSCs can be used as seed cells for the preservation of endangered species and for clinical treatment.