Effect Of Platelet-Rich Plasma Gel Factors Of Biological Effects In Vitro

Background and ObjectiveThe biological effects of platelet-rich plasma gel are affected by many factors, such asthe preparation method of platelet-rich plasma, blood platelet integrity, the choice ofanticoagulants and activators and so on. To find out the best method to prepare platelet-richplasma (PRP)and to compare the effects on the release of growth factors from platelet-richplasma gel with different combinations of anticoagulants and activators.Methods1．The whole blood was extracted from New Zealand rabbits to prepare the platelet-richplasma with three kinds of centrifuge methods:in group A,2400RPM centrifuge for10minutes、3600RPM centrifuge for15minutes；in group B,2000RPM centrifuge for10minutes、3000RPM centrifuge for15minutes；in group C,3000RPM centrifuge for10minutes、4000RPM centrifuge for10minutes. Compare the platelet counts and percentrecovery of platelet to determine an ideal centrifuge.2．The whole blood was extracted from the New Zealand rabbit,and respectively withEDTA and HS anticoagulant citrate dextrose. Use an ideal centrifuge to prepare theplatelet-rich plasma, The platelets were counted before and after centrifugation, and thepercent recovery of platelet was counted in each group.The platelet-rich plasma was activatedwith bovine thrombin and type I collagen in different groups. Transforming growth factor-β1(TGF-β1) and platelet-derived growth factor-AB (PDGF-AB) were measured in theplatelet-rich plasma gel of all the groups using the enzyme linked immunosorbent assay at2hours,1,3,5days after the platelet-rich plasma was activated, and release modes andconcentration of these two growth factors were compared at the same time.Results1． Platelet counting in group A is1315.14×109/L, the platelet concentration of groupA is approximately585%of whole blood,and the percent recovery of platelet in group A is70.28%; Platelet counting in group B is1005.32×109/L, the platelet concentration of group B is approximately448%of whole blood,and the percent recovery of platelet in group B is53.73%; Platelet counting in group C is850.12×109/L, the platelet concentration of group Cis approximately378%of whole blood,and the percent recovery of platelet in group C is45.53%.The average platelet concentration was higher in group A (1315.14×109/L) than ingroup B(1005.32×109/L)andC(850.12×109/L),and the percent recovery of platelet by the Amethod was observed to be significantly higher when compared to other methods, thesignificant differences were noted between A、BandC group(P<0.05).2．The platelet concentration of PRP prepared with EDTA as an anticoagulant isapproximately580%of whole blood, the percent recovery of platelet is69.81%; The plateletconcentration of PRP prepared with HS as an anticoagulant is approximately413%of wholeblood, the percent recovery of platelet is49.61%.The cumulative release of TGF-b1andPDGF-AB was maximum in the EDTA-type I collagen group, and the significant differenceswere noted between the EDTA-type I collagen group and the other groups（P<0.05）.The useof type I collagen as an activator resulted in sustained release of TGF-b1and PDGF-AB, andthe release of TGF-b1was in a time-dependent manner (r=0.873); while the aliquots of PRPclotted with thrombin had an immediate release of TGF-b1and PDGF-AB（P>0.05）.Conclusion1．2400RPM centrifuge for10minutes、3600RPM centrifuge for15minutes may be thebest PRP preparation method.2．The cumulative release of TGF-b1and PDGF-AB was maximum in the EDTA-type Icollagen group, and the release mode of the growth factors have a sustained slow steadyrelease patten.The concentration and the release modes of the growth factors in theEDTA-type I collagen group is more conducive to the seed cells of tissue engineering nucleuspulposus differentiated into nucleus pulposus-like cells and the treatment of intervertebraldisc degeneration.Conclusion summary2400RPM centrifuge for10minutes、3600RPM centrifuge for15minutes may be theideal PRP preparation method,the cumulative release of TGF-b1and PDGF-AB wasmaximum and the sustained release pattern of the growth factors in the EDTA-type Icollagen group make the biological effect of the platelet-rich plasma gel get more sufficientplay, and may be more conducive to the seed cells of tissue engineering nucleus pulposus differentiated into nucleus pulposus-like cells and the treatment of intervertebral discdegeneration.