Cell processing system for bone regeneration

Objectives: Transplantation of mesenchymal stem cells (MSC) from human bone marrow (hMSC) is a possible approach for bone tissue engineering. According to Japanese law, cell culture using autologous hMSC should be performed at a Cell Processing Center (CPC). The aim of present study was to establish a method for utilization of a CPC, including a practical work schedule and standard operating procedure. Methods: Bone marrow samples were aspirated from the posterior iliac crest and transported from a blood collection facility to a CPC at 22°C. Mononuclear cells were isolated and re-suspended in alpha-modified minimal essential medium containing 15% autologous serum and adherent cells were cultured for further expansion. Cultured hMCS were characterized and cell phenotypes were compared with bone marrow samples. After cultivation of hMSC for 3 weeks according to Good Manufacturing Practices, hMSC were transported to a blood collection facility and were used as a cell source with a beta-tricalcium phosphate (TCP) scaffold and a source of autogenous growth factors, platelet-rich plasma. Results: Human MSC grew to approximately 1×107 cells within 3 weeks. More than 90% viable cells were seen after transportation between the blood collection facility and CPC. On flow cytometric analysis, bone marrow samples contained approximately 0.2% of CD271+ and 2% of CD44+/CD105+ cells. After cultivation with growth medium for 3 weeks, most cultured hMSC expressed CD271. Co-culture of hMSC with beta-TCP or implant material did not influence cell growth and differentiation to the osteoblastic phenotype. Mycoplasmal and bacterial infection was not observed during overall autologous MSC cultivation or transportation between facilities. Conclusion: Autologous hMSC culture at a CPC in collaboration with a blood collection facility is useful for tissue engineering. This system is safe and could be applied to regeneration medicine for the head and neck region.