Ex vivo expansion of human umbilical cord blood hematopoietic stem/progenitor cells with support of microencapsulated rabbit mesenchymal stem cells in a rotating bioreactor

Expansion of human umbilical cord blood mononuclear cells (MNCs) was carried out with/without the support of alginate-chitosan-alginate (ACA) microcapsules containing rabbit bone marrow (rBM) mesenchymal stem cells (MSCs). Cells were cultured in a rotating wall vessel (RWV) bioreactor and also in tissue-culture plates using serum-free media supplemented with conventional doses of purified human recombinant cytokines for 7 days. The total nucleated cell density, pH and osmolality of the culture media in both co-culture systems were measured every 24 hours. Flow cytometry analysis of the CD34(+) population and methylcellulose colony assays for assessing the pluripotency of the population were carried out after Oh, 72h and 168h of culture. The RWV bioreactor co-culture, combined with a cell-dilution feeding protocol, was observed to be efficient in expanding UCB MNCs. By the end of 168h of culture using this system, the total nucleated cell number had grown around 107-fold, whilst the CD34(+) cells 26-fold and colony-forming units in culture 19-fold. Within RWV alone control and static co-culture control groups, however, expansions of total nucleated cell number were 52-fold and 10-fold, respectively, while CD34(+) cells and CFU-Cs numbers both changed mildly (p < 0.01, compared with RWV co-culture group). It was thus demonstrated that the expansion of HSCs can be achieved at a large-scale with the support of microencapsulated stromal cells using this bioreactor.