Study On The Application Of ADSCs Combined With 3-D PCL Scaffolds In The Model Of Knee Joint Cartilage Injury

ObjectiveThe incidence of knee joint cartilage injury is high,and it is difficult to heal itself after injury,so there is a wide range of research needs.The purpose of this study is to print PCL,which has been used in the clinical research of non knee joint cartilage repair,into nano scaffold by 3D printing,and to cultivate ADSCs in a special biaxial tissue regeneration system(tisxell system),and to use the cultured tissue-engineered cartilage to regenerate the knee joint injury animal model for treatment.According to the principle of tissue engineering,the knee joint injury was treated with regenerative cartilage tissue.In the research,quantity and purity of cell culture,differentiation,enrichment and proliferation as well as the outcome of tissue-engineered cartilage implanted into animal models,the feasibility of the tissue-engineered scheme was discussed,and the experimental data of the effect of the scaffold material and cell culture system were analyzed.MethodThis study is divided into three parts.In the first part,ADSCs were extracted from the patella of New Zealand rabbits by enzyme digestion and cultured in vitro.ADSCs were identified by flow cytometry with surface antigens CD44,CD90,CD45,CD34 and CD29.ADSCs were induced to differentiate into lipogenic,osteogenic and cartilaginous by adding differentiation medium,and detected by oil red O staining,alizarin red staining and toluidine blue staining.In the second part,ADSCs and 3D printing PCL scaffolds were enriched by tisxell regeneration system.MTT method was used to detect the total amount of cell expansion.Hematoxylin eosin staining(HE)was used to detect the distribution and enrichment of ADSCs in 3D printed PCL scaffolds.In the third part,There were four groups of six New Zealand white rabbits.All the 24 rabbitswere divided randomly.They were blank control group;model group;ADSCs treatment group;3-D stent ADSCs treatment group.At 42 and 84 days after operation,the cartilage repair of knee joint was observed and evaluated by gross observation and histological staining,and the effect of repairing cartilage defect was compared between ADSCs treatment group and 3-D stent ADSCs treatment group.ResultsADSCs from the knee jiont were well isolated and cultured.CD44,CD90 and CD29 were positive for the phenotype of MSCs.The percentage of positive cells was over 95%.CD45,CD34,these endothelial cell phenotype antigens were negative,the positive cells accounted for less than 2%.It shows that the purity of ADSCs we separated meets the requirements.Oil red O staining,alizarin red staining and toluidine blue staining showed that ADSCs had good lipogenesis and osteogenesis and differentiation potential of cartilage.After enrichment and culture by tisxell regeneration system,it was found that ADSCs grew intensively in 3D printed PCL scaffolds,and the total number of cells reached grade 108,which met the requirements of in vivo inoculation experiment.After 6 weeks and 12 weeks,ADSCs have abillity to repair the articular cartilage injury,but after tisxell regeneration system enriches ADSCs in 3D printing PCL scaffold,more transparent cartilage is formed in the treatment group,which has a better effect on the later functional recovery of knee joint.ConclusionADSCs under the patella has the potential of better chondrogenic differentiation,and can effectively repair the knee joint cartilage injury model.3D printing PCL scaffolds have good mechanical properties and pore size,which can better enrich and culture ADSCs,and better induce ADSCs to differentiate into hyaline cartilage tissue,which has a better effect on functional recovery of knee cartilage injury.Tixell regeneration system and improved culture medium can shorten the culture time of tissue-engineered cartilage from 3-4 weeks to 12 days.The study provides a new tissue engineering scheme for later clinical application research.