The Comparison With Proliferation Of Platelet Plasma And Platelet Lysate In Bone Marrow Stroma Cells

Objective Comparing platelet rich plasma (PRP) and platelet lysate (PL) on cultured rat bone marrow stem cells recharge (BMSCs) proliferation, cycle, explore the promotion of bone marrow mesenchymal stem cell effect.Method(1)The BMSCs were isolated from bone marrow in SD murine about a mouth old and cultured in defined medium in vitro. (2)12 weeks old SD rats, PRP obtained from heart blood by gradient centrifugation . 10% FBS high glucose DMEM complete medium as an ordinary group, platelet lysate were added to a final concentration of 1% ,3% conditioned medium group training BMSCs. BMSCs were identificated by CD44,CD34 ;inverted phase contrast microscope and MTT assay proliferation of BMSCs;flow cytometry assays cycle of BMSCs;PCNA immuno- cytochemistry and laser scanning confocal microscope identifies proliferation of BMSCs.Results 1. BMSCs was cultured successfully , BMSCs were detected on the surface marker CD44,CD34.CD44 is positive expression,CD34 is negative expression.2. BMSCs's appearance of spindle shape, fibroblast-like shape. The proliferation of BMSCs in 3% PRP and PL conditioned medium group is higher than in 1% PRP and PL group and higher than in general group. The cells grow density .3.The cell grow curve was by MTT method. Within 1-2days that was latency period. It was no obvious cell proliferation; Three days later, the cells grew fast obviously and came into logarithmic phase. After 7 days, the cells grew slowly and this phase was called plain phase. Cell growth curve is similar to "S" shape. By the statistical analysis, General and the concentration of 1%,3% PRP and PL medium on the proliferation of BMSCs was significant difference (P <0.05), the concentration of 1% ,3% PRP and PL medium on the proliferation of BMSCs was no significant difference (P> 0.05).4. Each groups of BMSCs cycle is detected by flow cytometry. The rate of Cells in G1 phase is more , the rate of Cells in S and G₂ phase is fewer at 1day. The proportion of each group cells decreased in G1 phase, increased in S , G₂ phase at 3,5day . The concentration of 1% ,3% PRP and PL G₁ phase of BMSCs was significantly lower than that the general (P <0.05). The concentration of 1% ,3% PRP and PL groups S,G₂ phase of BMSCs was significantly higher than that of general groups (P <0.05), the concentration of 3%PRP and PL ,1% PRP and PL G₁,S,G₂ phase of BMSCs was no significant difference (P> 0.05).5. There was positive expression in PCNA's immunohistochemistry .At 1,3, 5day,the expression of PCNA in concentration of 1% ,3% PRP and PL groups was significantly higher than that of general groups at 1, 3, 5day (P<0.05). The expression of PCNA in concentration of 3%PRP and PL ,1% PRP and PL groups was no significant difference (P> 0.05).6. BMSCs's PCNA expression in the experimental group showed the site of green fluorescent in laser scanning confocal microscope; 1d, 3d, 5d, 1%, 3% PRP and PL conditioned medium group were significantly higher than the ordinary complete medium group (P <0.05). 1% PRPand PL, 3% PRPand PL conditioned media group were not statistically significant (P> 0.05). PCNA immunohistochemistry's results and laser scanning confocal microscope's results are consistent.Conclusion: PRP and PL could promote the proliferation of BMSCs, both conditions's result is the same in quite concentration.1.In vitro culture conditions, different concentrations of PRP and PL are on the BMSCs had no effect on changes in cell morphology.2.PRPand PL could significantly increase the proliferation of cells, both under the same concentration of BMSCs on the growth curve and cell cycle changes were not significantly different. 3.PL and PRP can be significantly enhanced the expression of PCNA in BMSCs cells, both under the same concentration,they had no significant difference.