Experimental Study On Enhancing The Ability Of Rat Adipose - Derived Mesenchymal Stem Cells To Transform Into Islet - Like Cells

Objective:(1) The separation, extraction and culture of rat adipose mesenchymal stem cells.(2) To study the effects of injuried pancreatic extract and cytokines on the indudced differentiation of rat adipose mesenchymal stem cells(AMSCs) to islet-like cells.Methods:(1) The adipose tissues were separated from rat by the surgical operation under aseptic condition. The collagenase and trypsin digestion were used to extract the AMSCs from tissues and cells’ purification.(2)It consisted of four groups: control group, cell drug induced group, mixed induced group and injuried pancreatic extract group. Different cytokines and injuried pancreatic extract were added to different groups. The cell was induced with different methods for 11 days. Cells’ morphology changes were observed and recorded with inverted microscope. Insulin-positive cells were identified by dithizone(DTZ) staining and the amount of insulin of each group after being stimulated in the high glucose environment were measured through enzyme-linked immunosorbent assay. The related gene expression was detected with real-time quantitative polymerase chain reaction(RT-PCR) and immunofluorescence staining.Results:(1) The rat AMSCs attached to the plate. The primary cells were nearly round. The spindle cell type or cell clusters appeared after 36-48 h. Cell clusters grew around and the gap among cells decreased gradually in the next 7-8 days, finally, the cells arranged like whirlpool. At about 10 days, the cell growth density reached70%-80% of the bottom of bottle. The cells’ purification became higher than before after trypsin digestion.(2) The cells of induced groups gathered into clusters and their shapes became round. DTZ staining positive cells appeared. Insulin was secreted by the cells under high glucose environment, the insulin level and the related genes expression of mixed induced group were higher than those of other induced groups.There were no changes on the indexes mentioned above.Conclusion:(1) The AMSCs can be collected through collagenase digestion and trypsin digestion was used for cells’ purification.(2) The injuried pancreatic extract can promote AMSCs to differentiate into islet-like cells and the transformation ability of AMSCs can be improved by it.