Bioreactor Expansion of Human Adult Bone Marrow-Derived Mesenchymal Stem Cells

Supplementation of mesenchymal stem cells (MSCs) during hematopoietic stem cell transplantation (HSCT) alleviates complications such as graft-versus-host disease, leading to a speedy recovery of hematopoiesis. To meet such clinical demand, a fast MSCs expansion method is required. In the present study, we examined the feasibility of expanding MSCs from the isolated bone marrow mononuclear cells using a rotary bioreactor system. The cells were cultured in a rotary bioreactor with Myelocult� medium containing a combination of supplementary factors, including stem cell factor (SCF), interleukin 3 and 6 (IL-3, IL-6). After 8 days of culture, total cell numbers, Stro-1+CD44+CD34- MSCs and CD34+CD44+Stro-1- HSCs were increased 9, 29, and 8 folds respectively. Colony forming efficiency-fibroblast per day (CFE-F/day) of the bioreactor-treated cells was 1.44-fold higher than that of the cells without bioreactor treatment. The bioreactor-expanded MSCs showed expression of primitive MSCs markers endoglin (SH2) and vimentin, whereas markers associated with lineage differentiation including osteocalcin (osteogenesis), Type II collagen (chondrogenesis) and C/EBPα (adipogenesis) were not detected. Upon induction, the bioreactor-expanded MSCs were able to differentiate into osteoblasts, chondrocytes and adipocytes. Taken together, we conclude that the rotary bioreactor with the modified Myelocult� medium reported in this study may be used to rapidly expand MSCs.

The influence of cyclosporin alone, or cyclosporin and methotrexate, on the incidence of mixed haematopoietic chimaerism following allogeneic sibling bone marrow transplantation for severe aplastic anaemia

We previously reported a randomized trial comparing Cyclosporin-A (CsA) and short-term methotrexate versus CsA alone for graft-versus-host disease (GvHD) prophylaxis in 71 patients undergoing allogeneic haematopoietic stem cell transplantation (HSCT) from a human leucocyte antigen-identical sibling for severe aplastic anaemia (SAA). We found a better survival in the group receiving the two-drug prophylaxis regimen with no significant difference in the probability of developing GvHD between the two groups. The present study details chimaeric analysis and its influence on survival and GvHD occurrence in 45 of the original 71 patients in whom serial samples were available. Analysis was carried out in a blinded prospective manner. Seventy-two per cent achieved complete donor chimaerism (DC), 11% stable mixed chimaerism (SMC) and 17% progressive mixed chimaerism (PMC). The overall 5-year survival probability was 82% (+/-11%) with a significant survival advantage (P = 0.0009) in DC or SMC compared to those with PMC. Chronic GvHD was more frequent in DC patients, whereas no patient with SMC developed chronic GvHD. Graft failure occurred in 50% of the PMC group. This study demonstrates the relevance of chimaerism analysis in patients receiving HSCT for SAA and confirms the occurrence of mixed chimaerism in a significant proportion of recipients.