Application Of Cell Sheet On Structural Allograft Healing Of Bone Defects

Critical sized bone defect is a major challenge in the field of orthopaedic surgery.Processed allografts remain an attractive substitute for bone grafting.It has long been recognized that there are several fundamental differences between a live autograft and a processed allograft including: allografts don’t contain living cells that can produce new bone;allografts don’t contain growth factors and osteogenic substances;and allografts are lack of appropriate angiogenesis.Therefore,if the chondrogenic,osteogenic and angiogenesis property of bone-marrow derived stromal cells(BMSC)could be enhanced in the bone remodeling,bone defect healing during skeletal repair can be promoted.In this paper,according to the needs of bone repair,we hope that BMSC Cell Sheets act as an engineered periosteum that promotes bone callus formation during allograft healing and integration in a preclinical mouse model of critical sized femoral allograft.Additionally,it is reported that intermittent PTH could increase not only the bone mass but also small vessels in the sites of new bone formation.Therefore,our study hope that transient Jagged1-mediated NOTCH signaling could promote human BMSC(h BMSC)expansion in vivo,while preserving their differentiation capacity,and explore the effect of allografts wrapped with NOTCH-mediated h BMSC Cell Sheet combined with PTH on the allograft healing.Additionally,the preliminary study of human-induced pluripotent stem cell(hi PSC)was carried out to determine its possibllity in the field of criticized bone defect.The purpose of this study is to improve the effect of allograft on bone defect repairing,which is divided into two parts.The first part,targeting the Notch pathway of h BMSC to form Cell Sheet combined with PTH to promote the repairing of bone defect with allograft bone.In this part,we target the activation of Notch h BMSC signaling pathway in vitro,and construct the allograft wrapped by Cell Sheet.At the same time,the experimental model of bilateral ovarian resection(OVX)was constructed to study the effect and mechanism of Cell Sheet combined with PTH on the healing of critical bone defect.The second part is the function of human induced stem cells(hi PSC)Cell Sheet for the early basic research of bone defect repair.We obtain the transfected hi PSC in vitro,identify the specific markers,and develop hi PSC Cell Sheet.And the ability of hi PSC-MSC to differentiate into chondrocytes and osteoblasts was verified by experiments in vivo and in vitro.In vivo and in vitro experimental results show that Cell Sheet formed by targeting Notch signaling pathway promotes the formation of cartilage,osteogenesis and angiogenesis to accelerate the healing of bone defect with allograft bone.PTH can enhance the healing of bone defect with allograft bone through the increase of CFU number of BMSC,BMSC osteogenic and chondrogenic differentiation,angiogenesis and the activity of osteoclasts to accelerate callus remodeling to promote the repairing of the bone defect.At the same time,hi PSC with osteogenic and chondrogenic differentiation potential,is a potential source of seed cells,which can be used as Cell Sheet to promote bone defect repair.We use concept of tissue engineering,h BMSC,allograft bone and growth factor binding,and the signaling transduction,in vitro we target Notch ligand Jagged1 to activate Notch signaling of h BMSC.Then in vivo we demonstrate that the Notch signaling activation of h BMSC Cell Sheet combined with PTH can promote allograft bone repairing of bone defect.This method provides a favorable approach and insight for the treatment of massive allograft healing.At the same time,through in vivo and in vitro experiments,we suggest that hi PSC has the potential to replace h BMSC as Cell Sheet to promote bone defect repairing.