Vitreous cryopreservation of tissue engineered bone composed of bone marrow mesenchymal stem cells and partially demineralized bone matrix.
Cryopreservation of tissue engineered products by maintaining their structure and function is a prerequisite for large-scale clinical applications. In this study, we examined the feasibility of cryopreservation of tissue engineered bone (TEB) composed of osteo-induced canine bone marrow mesenchymal stem cells (cBMSCs) and partially demineralized bone matrix (pDBM) scaffold by vitrification. A novel vitreous solution named as VS442 containing 40% dimethyl-sulfoxide (DMSO), 40% EuroCollins (EC) solution and 20% basic culture medium (BCM) was developed. After being cultured in vitro for 8 days, cell/scaffold complex in VS442 was subjected to vitreous preservation for 7 days and 3 months, respectively. Cell viability, proliferation and osteogenic differentiation of cBMSCs in TEB after vitreous cryopreservation were examined with parallel comparisons being made with those cryopreserved in VS55 vitreous solution. Compared with that cryopreserved in VS55, cell viability and subsequent proliferative ability of TEB in VS442 after being rewarmed were significantly higher as detected by live/dead staining and DNA assay. The level of alkaline phosphatase (ALP) expression and osteocalcin (OCN) deposition in VS442 preserved TEB was also higher than those in the VS55 group since 3days post-rewarm. Both cell viability and osteogenic capability of the VS55 group were found to be declined to a negligible level within 15 days post-rewarm. Furthermore, it was observed that extending the preservation of TEB in VS442 to 3 months did not render any significant effect on its survival and osteogenic potential. Thus, the newly developed VS442 vitreous solution was demonstrated to be more efficient in maintaining cellular viability and osteogenic function for vitreous cryopreservation of TEB over VS55.
Yin
Hongyu
H
Plastic Surgery Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China.
Cui
Lei
L
Liu
Guangpeng
G
Cen
Lian
L
Cao
Yilin
Y
D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D001857:Bone Matrix / Q000187:drug effects*; D001842:Bone and Bones / Q000601:surgery; D002454:Cell Differentiation; D015925:Cryopreservation; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004285:Dogs; D059630:Mesenchymal Stem Cells / Q000166:cytology; D010012:Osteogenesis / Q000187:drug effects; D013552:Swine; D023822:Tissue Engineering
D004121:Dimethyl Sulfoxide