Activation of rat nucleus pulposus cells by coculture with whole bone marrow cells collected by the perfusion method.

Cell proliferation and matrix synthesis were compared for rat nucleus pulposus cells cocultured with mesenchymal stem cells (MSCs) or fresh whole bone marrow cells (BMCs), harvested by the perfusion or aspiration methods. Nucleus pulposus cells were isolated from tail intervertebral discs of F344/slc rats, and BMCs were obtained from femora. Proteoglycan synthesis, DNA synthesis, and aggrecan mRNA expression were measured. The level of transforming growth factor-beta in supernatants from the culture system was also measured. Cell number, aggrecan mRNA expression, and uptake of [(35)S]-sulfate and [(3)H]-thymidine by nucleus pulposus cells cocultured with fresh whole BMCs all increased significantly compared with nucleus pulposus cells cocultured with MSCs. TGF-beta secreted by nucleus pulposus cells cocultured with fresh whole BMCs also significantly increased when compared with cocultures with MSCs. The perfusion method was superior to the aspiration method for preventing contamination of BMCs with peripheral red blood cells and lymphocytes, which may cause an autoimmune response in the disc. In conclusion, we suggest that fresh whole BMCs harvested by the perfusion method are more effective for increasing the proliferative and matrix synthesis capacity of nucleus pulposus cells.

Regenerative effects of transplanting autologous mesenchymal stem cells on corticosteroid-induced osteonecrosis in rabbits.

Osteonecrosis (ON) of the femoral head is one of the most serious complications associated with steroid administration. Here, we treated corticosteroid-induced ON in the rabbit by transplanting mesenchymal cells (MCs). Rabbits were injected once with 20 mg/kg of methylprednisolone (MPSL) and divided into three groups as follows: (1) MPSL alone (no further treatment); (2) MPSL+MCs (7 days after MPSL, MCs [1 x 10(7)/2 ml] were injected into the bone marrow cavity of the femurs); (3) MPSL+saline (7 days after MPSL, saline [2 ml] was injected into the bone marrow cavity of the femurs). Subsequently, the incidence of ON in the femurs 4 weeks after MPSL alone and MPSL+saline was 80 and 68.4%, respectively. In contrast, no ON was recorded in rabbits treated with MPSL+MCs. Vascular endothelial growth factor (VEGF) staining and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate (dUTP) nick end-labeling (TUNEL) staining was more marked in the MPSL alone and MPSL+saline groups than in the MPSL+MCs rabbits. The percentages of cells in the G1 phase in the MPSL+MCs group were significantly lower than in the other two groups. These findings suggest that the injection of autologous MCs into the femur could prevent corticosteroid-induced ON in patients treated with high-dose short-term steroid medication.