Effect of Cryopreservation on Canine and Human Activated Nucleus Pulposus Cells: A Feasibility Study for Cell Therapy of the Intervertebral Disc

摘要

It has been shown that coculture of bone marrow–derived stromal cells (BMSCs) with intervertebral disc (IVD) nucleus pulposus (NP) cells significantly activates the biological characteristics of NP cells in animal models and in humans. We therefore predicted that activated NP cells would be a useful graft source for cellular transplantation therapy in the treatment of degenerative IVDs. However, the activation protocol is based on fresh isolation and activation of NP cells, which limits the timing of clinical application. Cell transplantation therapy could be offered to more patients than is now possible if activated NP cells could be transplanted as and when required by the condition of the patient. No study has investigated the effect of cryopreservation on NP cells after enzymatic isolation. We investigated the effects of cryopreservation of canine and human NP cells in both cell and tissue form before coculture with autologous BMSCs. Cell viability, proliferation, glycosaminoglycan production, aggrecan transcriptional activity, colony generation, and gene expression profile of the cells after cryopreservation and subsequent coculture were analyzed. The influence of cryopreservation on cell chromosomal abnormalities and tumorigenesis was also studied. The results showed that there were no clear differences between the noncryopreserved and cryopreserved cells in terms of cell viability, proliferation capacity, and capacity to synthesize extracellular matrix. Furthermore, the cells showed no apparent chromosomal abnormalities or tumorigenic ability and exhibited similar patterns of gene expression. These findings suggest that by using cryopreservation, it may be possible to transplant activated NP cells upon request for patients' needs.