Co-culture Of Human Adipose-derived Stem Cells And Mature Adipocytes

Just like bone marrow mesenchymal stem cells(BM-MSCs),aidpose derived stem cells(ADSCs) have the capacity of multi-differentiation.However,ADSCs can be isolated easily with abundant resources and little injury to patients.Thus more and more researchers became interested in the study of ADSCs.One of the important applications of ADSCs is Tissue Engineering.It's very clear that ADSCs are the most suitable seed cells for the construction of engineering adipose tissues.Thus,it is meaningful to study the adipose differentiation of ADSCs.In this paper,ADSCs and mature adipocytes were isolated from human liposuctioned adipose tissue.Observation in cell morphology,Oil Red O assay,trypan blue stain,the continual culture of mature adipocytes from adipose tissue,were taken to the mature adipocytes used in experiments;Observation in cell morphology,cck-8 for growth curve,Oil Red O assay after differentiating into adipocytes,hoechst fluorescence stain and cells surface maker expression by flow cytometry were taken to the ADSCs isolated from adipose tissue in our tests.In this study,the differentiation of human adipose-derived stem cells into adipocytes was studied by means of coculturing them with human mature adipocytes.Then we used transwell to carry out the indirect co-culture of human adipose-derived stem cells(in up-chamber) and human mature adipocytes(in bottom-chamber),with 4 different kinds of ratios of stem cells to mature adipocytes,as 1:5,1:1,2:1 and 5:1.Oil Red O Assay and Trypan Blue Assay were taken after this co-culture for the evaluation of adipose differentiation of human adipose-derived stem cells.Induction of ADSCs with adipose differentiation reagents was taken as positive control, and culture of ADSCs with normal medium was taken as negative control.It was proved that human adipose-derived stem cells could not differentiate into mature adipocytes within 8 days co-culture.It's suggested that two reasons may exist for this result. The first one may be that this period of co-culture was too short to induce ADSCs into mature adipocytes;the other one may be that it couldn't make ADSCs differentiate into mature adipocytes,only by this co-culture way.To verify the first cause above,the period of coculture was prolonged to 20 days.And flow cytometry,hoechst,PI assay of ADSCs after coculturing with mature adipocytes were used in this parts of experiment.It was still not that definite differentiation of ADSCs into mature adipocytes happened after 20 days coculture.But the ADSC number expressing CD105 after coculturing with mature adipocytes decreased.It may implicate that ADSCs after coculture model has been committed to adipocytes in some extent of differentiation.The indirect co-culture of ADSCs and mature adipocytes for adipose differentiation of stem cells was carried out for the first time within this study.The result of this paper maybe an implication of further development of adipose differentiation,and will be helpful for the research and application of adipose tissue engineering.