The Biological Characteristics Study Of Antigen-extracted Cancellous Bone

Objective:Bone is an important part of the body,but its defect caused by trauma,infection,tumor,inflammation,congenital diseases and other causes is constantly threat to human health.Large-scale bone defects and repair of special bone defect of joint,facial and other parts remains a problem difficult to completely solve.Autologous bone or allogeneic are now usually used for bone defects,but they caused a lot of complications like pain,infection,body rejection.In addition,the sources of autologous bones are relatively less,leading to the limited quantity available.The research of bone tissue engineering offers hope to solve this problem.The construction of tissue engineered bone include three important factors:seed cells,scaffold materials and construction method.Currently,people often use osteoblasts or mesenchymal cells(like MSCs) for seed cells,the method of get and culture these two kinds of cells is mature,but the injury caused by the method and limited cell sources limit the clinical application.Ideal scaffold for bone tissue engineering should meet the following characteristics:good biocompatibility and safety,good biological activity, proper mechanical strength, appropriate porosity etc.A good scaffold should not only to meet the forward conditions,but also has wide sources.Antigen-extracted xenogeneic cancellous bone matrix is easy to be obtained,it has pore structure like human bone and bone morphogenetic protein as well as other osteogenesis factors,its beneficial for osteoblasts's adhesion, proliferation and transfer of material.Thus it catches more and more attention.In this research,we used traditional method plus Supercritical Carbon Dioxide Fluid Technology to make antigen-extracted xenogeneic cancellous bone matrix for cultivation with human adipose stem cells (hADSCs) and differentiate them into osteoblasts,and then investigate the feasibility of using it for bone tissue engineering scaffold material and its biological characteristics.Methods:1.Fresh pig distal femur trabecular bone was treated by Supercritical Carbon Dioxide Fluid Technology and retreated by absolute alcohol,chloroform,methanol,H2O2,HCl ,then determined it by Thermogravimetry-Differential thermal analysis,SEM and calculation of porosity.2.hADSCs were cultivated from human fat aspirates,then the passage 5 hADSCs were induced to differentiate to the osteoblasts and adipogenic cells in vitro,and cultured with bone matrix,one group was cultured with hADSCs and then induced them to differentiate to the osteoblasts,another one was only cultivated with uninduced hADSCs.This process is divided into two groups,one group was cultivated in vitro staticly,another was cultivated in Bellco roller bottle.The growth of cells on the carrier was observed through inverted phase contrast microscope and SEM. Results:1. The treated cancellous bone which is removed of fat and protein ,retains the pore structure of bone tissue.2.Observed by inverted phase contrast microscope and scanning electron microscope, hADSCs and osteogenic induced cells on the bone vector grew well, alkaline phosphatase expression was positive in the osteogenesis induction group, mineralized nodule formation.Conclusion:The treated pig cancellous bone has a proper pore diameter and porosity and has no significant adverse effects for the growing of the hADSCs and their differentiation into osteoblasts, it has good biocompatibility, can be used as a good bone tissue engineering scaffold material.