| Background:Adipose-derived stem cells (ADSCs) were extracted from adipose tissue by Zuk in2001. It has been discovered that adipose-delivered stem cells can be induced todifferentiate toward adipose cells, cartilage cells, Gegenbaur’s cells, muscle cells,nerve cells, hepatocytes and islet cells with different culture media, which illustratesthe potential of differentiation and cross-lineage differentiation.Adipose-derived stem cells (ADSCs) have several advantages. Firstly, human bodycontains huge amount of ADSCs, therefore a sufficient source is available. Secondly,ADSCs can be accessed simply. Thirdly, adipose tissue keeps rich stem cells. Fourthly,ADSCs have a strong hyperplasia ability; hence they have a stable and same functionafter producing several generations. Finally, they belong to autotransplantation,therefore no reject reaction exist. Due to these merits, Adipose-derived stem cells(ADSCs) are gradually concerned as seeded cells in terms of tissue engineering. Soautologous fat transplantation has been widely used in plastic surgeries. However,their low rate of survive leads to the requirement of an excess of experiments andcertain complications. It has been found that their rate of survive can be enhanced byplatelet-rich plasma (PRP) in cell experiments and zoopery, but the mechanism ofaction cannot be clarified.Wnt3gene and Klotho gene are relative genes to stem cells multiplication. In thisexperiment, the promotion level of Wnt3and Klotho is examined by adding PRP intoADSCS.Objective:The objective of the experiment is to examine that PRP has effects on promotingWnt3gene and Klotho gene in rabbit ADSCs.Methods:1. Extract ADSCs in epididymis adipose tissue from a New Zealand rabbit thenidentify them by Morphology and inducing differentiation, cultivate them to thefourth generation. 2. Prepare platelet-rich plasma (PRP) and platelet-poor plasma (PPP) fromabdominal aortic blood in a New Zealand rabbit through traditional centrifugemethod.3. Cultivate ADSCs in PRP (experiment), in PPP (comparison) and in an emptyculture bottle with5%nutrient fluid respectively for24hours,48hours and72hours namely.4. Crack cells and extract RNA in each group, check promotion level of Wnt3geneand Klotho gene by RT-PCR.Results:1. The original ADSCs were ovally shaped, after4-6hours the cells started to stickto the edge and to become long shuttle shape in25cm2culture bottle.8-10hourslater80%of the cells stick to the edge. With the multiplication, the cells grew as awhorled spindle. After they became the third or fourth generation, the cells werepurified, and they followed Morphology when placing it upside down on themicroscope.2. Coloring them by oil red O and alizarin red namely successfully demonstratesthey are adipose cells and osteocyte, as well as their ability of multilineagedifferentiation.3. The amount of Wnt3gene and Klotho gene increased significantly showed byPT-PCR result.Conclusion:It can be proved that the cells are rabbit adipose-derived stem cells (ADSCs) bymorphologic analyzing and multilineage differentiating the cracked and in vitrocultivated cells. Prepare platelet-rich plasma (PRP) can improve the multiplication ofADSCS and enhance the promotion of Wnt3gene and Klotho gene. |
PDF Full Text Request