The Differentiation Of Adipose-derived Stem Cells In Hydrogel In Vitro

Objective: It has been proved that adipose tissue can serve as an abundant, accessible and rich source of adult stem cells with multipotent properties which was suitable for tissue engineering and regenerative medical applications. There has been increased interest in adipose derived stem cells (ADSCs) for tissue engineering applications.In this part,methods for the isolation, expansion of ADSCs are presented and described in detail. While this part study at the isolation of ADSCs from rat adipose tissue, the procedure can be applied to adipose tissues from other species with minimal modifications. At last of this part we will do some research to describe the Phenotypic characteristics of ADSCs. Materials and Methods: ADSCs cells were obtained from rat's adipose tissue located at groin and digested by collagenase.The variation of morphology was observed by inverted phase contrast microscope.The cellular proliferation activity was assessed.At last flow cytometry was used to analyse and the expressions of immunophenotype.Results: Our study show that there was a population of cells with high potential to proliferate in the adipose tissue of rat.The primarily cultured cells exhibited a fibroblast-like morphology after culturing.ADSCs also showed characteristic like other stem cells.The morphous and phcnotype of ADSCs were similar to bone marrow derived stromal cells(BMSC).The immmophenotype phenotype of the ADSCs based on flow cytomctry shows CD29(+),CD44(+),CD90(+),CD45(-),CD49d(-),CD56(-).Conclusion:(1) ADSCs derived from SD rat' groin can get a high purity. (2)The purity of ADSCs can be improved by subculture. (3) ADSCs express specific surface molecules, with the characteristics of mesenchymal stem cells, has strong self-renewal ability and differentiation potential. Objective: To verify the multipotential differentiation of mesenchymal characteristics of rat ADSCs, cells were subjected to differentiation in conditions known to induce adipogenic and osteogenic lineages. To discuss the osteogenic and adipogenic differentiation ability of ADSCs with or without hydrogel in vitro.Materials and Methods: In two-dimensional manner to cultured rat adipose stem cells with certain induction medium and observe the osteogenic and adipogenic ability of ADSCs.To seed ADSCs in hydrogel and culture in three-dimensional way. Then use the same induction medium as before and observe the adipogenic and osteogenic differentiation ability.Results: In the two-dimensional culture environment, After 14 d of adipogenic, the ADSCs shows the ability to differentiation into adipocyte which full of lipide, after 14 d of osteogenic induction, emergence of osteoblasts with osteogenesis. Training in the three-dimensional environment, the osteogenic and adipogenic capacity still exists, but need a longer time to complete.Conclusion: 1. ADSCs cultured in two-dimensional environment have the adipogenic and osteogenic ability; ADSCs have characteristics of the multipotential differentiation of mesenchymal.2. ADSCs and hydrogel has good compatibility, ADSCs cultured in hydrogel have the ability with proliferation and division;3. ADSCs cultured in three-dimensional environment (hydrogel) has good osteogenic and adipogenic ability. Objective: To observe the effect of olfactory ensheathing cells and hydrogel on the differentiation of adipose derived stem cells into OEC-like cells and test the Neurogenesis ability of ADSCs in peptide hydrogel polymer scaffolds with a special cultue condition.Materials and Methods: In order to obserbe the effect of OECs and hydrogel on the differentiation of ADSCs we use Transwell Insert cell culture plate for co-culture experiments. Adipose derived stem cells were placed in the upper chamber with hydrogel and OECs in the lower. In control group there was only adipose derived stem cells in the upper chamber. After co-culture for 12 d the adipose derived stem cells were tested if they express the antigenic phenotypes (P75) of OECs.To test neurogenesis of adipose-derived stem cells (ADSCs) in hydrogel,ADSCs were isolated from rats, seeded in the peptide hydrogel polymer scaffolds and cultured in Neurobasal (NB) media supplemented with B27, basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). After 10 days in culture, some cells were expanded into clonal populations in which the expression of both Nestin and Brdu was detected but only the expression of Brdu was detected in the cells which were not expanded into clonal populations.Results: Our results showed that Olfactory ensheathing cells and hydrogel promoted differentiation of adipose derived stem cells into OEC-like cells. The proportion of OEC-like cells grew in experimental group. Our data suggested that ADSCs in peptide hydrogel polymer scaffolds can be induced to differentiate into cells which were characterized by the expression of the neuronal associated marker and the ability to undergo self-renewal and self-propagation.Conclusion: 1. Olfactory ensheathing cells and hydrogel promote differentiation of adipose stem cells into OEC-like cells. The outcome is better compared with control group.2. peptide hydrogel has an excellent biocompatibility with ADSCs and ADSCs has a neural differentiation capacity in peptide hydrogel in appropriate induction conditions.