Investigation Of Biological Characteristics Of Tissue Engineered Periosteum After Cryopreservation

Objective:To observe the biological characteristics of cryopreserved tissue-engineered periosteum and to examine the osteogenic potential.Methods:The bone marrow mesenchymal stem cells(BMSCs)of New Zealand rabbit were first cultivated and osteogenically induced as seeding cells in vitro,then co-cultivated with porcine small intestinal submucosa(SIS).Other BMSCs were not osteogenically induced and co-cultivated with SIS as well.They were both cryopreserved at -196℃in liquid nitrogen for 30 days banking in cryoprotectant, which contains 10%DMSO,50%fetal calf serum by programmed cooling procedure. Subsequently,the biological characteristics of cryopreserved tissue-engineered periosteum were analyzed by scan electronic microscopy(SEM),MTT and ALP detection after resuscitation.In the meantime,the quantity of typeⅠcollagen and osteocalcin secreted by the tissue-engineered periosteum(TEP)was detected by western blotting.Rusults:The growth status of BMSCs remains good after resuscitation.The quantity of typeⅠcollagen,osteocalcin and ALP secreted by the two kinds of TEP were higher than SIS.The quantity of typeⅠcollagen in the TEP which was constructed with BMSCs and SIS(M1 group)was not significantly different from the TEP which was constructed with osteoblasts and SIS(M2 group).(P＞0.05)Other wise, the quantity of osteocalcin and ALP in the M2 group was all significantly higher than the M1 group.(P＜0.05)Conclusion:The biology activity of tissue engineered periosteum remained stable and the quantity of typeⅠcollagen,ALP and osteocalcin was secreted at a certain level.The tissue-engineered periosteum showed a highly osteogenetic activity. The expression level of osteogenetic activity in M2 group was higher than M1 group.