The Study Of The Impact Of A New Type Of Hydrogels With PRF On Chondrogenic Differentiation Of BMSCs

Objective In order to provide a new hydrogels with platelet-rich fibrin(PRF)to repair of cartilage defect in clinical,this study explored the physicochemical and biological properties,and observed the impact of that hydrogels on chondrogenic differentiation of BMSCs.Method The chitosan(CS)thermosensitive hydrogels with Genipin was firstly prepared.2%(w/v)chitosan solution,56%(w/v)?-glycerophosphate(?-GP)solution and 0.05%(w/v)or 0.1%(w/v)genipin solution were mixed with different proportion to get the CS/GP mixed solution and CS/GP/genipin mixed solution.The gel-formation time was measured at 37?,observed mechanical strength and determined the degradation of these samples through in vitro experiment.After the hydrogels made into freeze-drying scaffold,the micro structure of that hydrogels were observed under scanning electron microscope(SEM),the structure of these samples used Fourier Transform Infrared Spectrometer(FTIR),and the swelling ratio are determined.Then preparation of PRF by once centrifugation,ground the PRF that through cryodesiccation intopowder,and added the PRF powder in the mixed solution Observed the new type hydrogels with PRF are able to delhyed release the TGF-?1 and IGF-1 in vitro experiment Its cytotoxicity of leach liquor of freeze-dried scaffold was detection by Cell Counting Kit-8.Finally,observed the growth of BMSCs thatwere cultured on the surface of the hydrogels in the 12-we In plate by fluoresceindiacetate(FDA)fluorescent staining.Chodrogenic induction of BMSCs after were cultured in the 24-well plate,added the new type of hydrogels,chitosan thermosensitive hydrogels and empty control at the same time.Obeserved the impact of the new type of hydrogels on chondrogenic differentiation by HE staining,safranin O-fast green staining alcian blue staining and immunohistochemical staining of type ? collagen after 2 weeks of cell culture.The amout of Glycosaminoglycans(GAGs)in the supernatant was quantitatively determind by Alcian blue colorimetric assay.The amount of type ? collagen in supernatant was quantitated by ELISA.Result The gel-formation time of the volume ratio of 6:1:1 of the CS/GP/Genip in was about 8 min at 37 ?.The mechanical strength and swelling ratio of that hydrogels respectively are 5 level and(404.62±105.39)%,and it can complete degradation about 7 weeks in vitro experiment at 37?.The SEM images indcate that the inside of hydrogels present spongy porous network structure.The result of FTIR showed that Genipin and CS had crosslinked copolymer.The ability of delayed release of TGF-?1 and IGF-1 showed that the new type of hydrogel with PRF and PRF to be equal,there are no statistical significance in the difference(P>0.05).The new type of hydrogel with PRF have a good biocompatibility with BMSCs.The BMSCs seeded on the surface of the hydrogels presented good proliferation by FDA fluorescent staining.For the group of the new type of hydrogels,the result of HE staining,safranin O-fast green staining,alcian blue staining and immunohistochemical staining of type ? collagen were positive,and the other two group were weakly positive.There are GAGs and type ? collagen in the supernatant of three groups.But there are statistical significance between the group of the new type of hydrogel and the other groups for the difference in secretion of GAGs and type ? collagen(P<0.05).Conclusion The volume ratio of 6:1:1 of the CS/GP/Genipin should be able to form the hydrogel that the physicochemical properties is satisfectory.The new type of hydrogel with PRF able to delayed release the growth fector througba certain speed.That hydrogel had a good cytocompatibility with BMSCs,and able to promote choldrogenic differentiation to a certain degree.There is prospect in the field of cartilage tissue engineering.