Isolation,Identification,and Biological Characteristics of Mouse Adipose-derived Stromal Cells

【Objective】 To establish an effective way for isolation and expansion of mouse adipose-derived stromal cells(ADSC),and to observe the biological characteristics of mouse ADSC.【Methods】 ADSC were obtained from adipose of BALB/C mouse,isolated and purified in vitro to determine cell morphology,immunophenotype,differentiation potentiality,clone formation ability,growth kinetics and immunoloregulation characteristics in the lymphocyte proliferation inhibition test: T lymphocytes(1 × 105) were cultured in the presence of ConA(ConA was 20 μg/mL),and ADSC of different number interacted with activated T lymphocytes respectively.Experiment was divided into 8 groups by the different number of ADSC: A group as positive control(ConA+T lymphocytes),B group(ADSC 1 × 103+ConA+T lymphocytes),C group(ADSC 1 × 104+ConA+T lymphocytes),D group(ADSC 2 × 104+ConA+T lymphocytes),E group as negative control(T lymphocytes),F group(ADSC1 × 103),G group(ADSC1 × 104),H group(ADSC 2 × 104),F,G,H were reference groups.The ability of T lymphocyte proliferation was measured by cck-8 method.【Results】 ADSC showed plastic-adherent,fibroblastic-like morphologic characteristics,cell proliferation was inadequate and cell morphology changed to flat,enlarged shapes with the passage increasing.ADSC at passage 3 highly expressed CD29 and CD44,moderately expressed Sca-1,weakly expressed CD34,CD45,and CD11b.Three passaged ADSC could be induced to differentiate into osteoblasts and adipocytes,and had strong ability to shape colony forming unit-fibroblasts(CFU-Fs).Doubling time of three,five,and eight passaged ADSC was(22 ± 3)h,(24 ± 3)h,and(30 ± 3)h,respectively.The proliferation rate after passage 5 slowed down gradually.ADSC could suppress T lymphocyte proliferation,the inhibition ratios of B,C,and D group were 20.78%,47.94%,and 60.70%,respectively,which was similar to bone marrow mesenchymal stem cells(BMSC).【Conclusion】 ADSC can be isolated and expanded effectively by using adherent culture,they possess general characteristics of mesenchymal stem cells,in addition,they are easier to be purified and their proliferation rate is faster compared with BMSC,so ADSC may be a good multipotential cell candidate for the future cell replacement therapy.