Effect Of Platelet-rich Plasma (PRP) On Proliferation Of Rat Adipose-Derived Stem Cells In Vitro

Objective: To explore the methods for the isolation and expansion of adipose-derivedstem cells (ADSCs) from SD rat. To observe proliferation characteristics, surfaceantigens and the differentiation potential.Methods: ADSCs cells were separated from SD rat’s adipose tissue located atbilateral groin and digested by collagenaseⅠ.The ADSCs were purified by adherencescreening method. Growth curve of ADSCs was recorded. Surface antigens wereindentified by flow cytometry. We used the third passage ADSCs to induce intoadipocytes by adipogenic inducing fluid and identified by Oil Red O staining. Weuesd the third passage ADSCs to induce into osteoblasts by osteogenic inducing fluidand identified by Alizarin Red staining.Results: Large amount of ADSCs with high purity were obtained by type Ⅰcollagenase digestion method and adherence screening method. The ADSCs exhibitedfusiform or polygonal shape and proliferated rapidly. Growth curve show S-shape. The surface antigens of the ADSCs based on flow cytometry showed CD45(-),CD106(-), CD90(+). The third passage ADSCs were induced withadipogenic/osteogenic inducing fluid and observed red stained lipid droplet ofadipocytes in the cytoplasm on Oil Red O staining and calcium nodes on Alizarin Redstaining.Conclusions: The ADSCs isolated from SD rat’s adipose tissue can get a high purityand proliferated rapidly. The purity can be better by subculture. ADSCs expressspecific surface antigens, has differentiation potential, comly with the characteristicsof MSCs. Objective: To explore the effects of platelet-rich plasma (PRP) on the proliferation ofADSCs in vitro. To find suitable concentration and mechanism, provide theoriesfoundation for application of PRP on clinical fat transplatation.Methods:1. The PRP were obtained by two-step centrifugation and used to cultureADSCs. There are three groups: control group, non-activated PRP group, activatedPRP group (five concentration gradients:1%PRP、5%PRP、10%PRP、15%PRP、20%PRP). The proliferation of ADSCs were analyzed by CCK-8method at1st,2nd,3rd,4th,5th,6th,7th, the OD was detected by enzyme mark instrument at450nmwavelength. The suitable concentration was5%and10%PRP.2. DNA synthesis and cell cycle related proteins were analyzed. There are two groups: control group,10%PRP group The DNA synthesis was observed by PI method and flow cytometry at48hours after PRP treatment. The expression of cyclinD1and p27Kip1were detected bywestern blot.Results: The morphology of ADSCs after PRP treatment showed no obviousdifference with control cells. It showed activated-PRP could promote the proliferationof ADSCs, the strength of proliferation was correlated to the treatment time. Therewas obvious dose-dependent effect under the suitable concentration range and highdose showed inhibitory effect.10%PRP increased the DNA synthesis of cellularS-phase and G2/M-phase and decreased G0/G1-phase after. PRP could promoteG0/G1-phase into S-phase.10%PRP increased the expression of cyclinD1and p27Kip1obviously.Conclusion: PRP could promote cell proliferation and fissiparism of ADSCs byregulating DNA synthesis and the expression of cell cycle related proteins.