Bone marrow (BM) mesenchymal stem cells (MSC) support proliferation and differentiation
of hematopoietic progenitor cells (HPC) in vitro. Since they represent a rare subset of
BM cells, MSC preparations for clinical purposes involves a preparative step of ex vivo multiplication.
The aim of our study was to analyze the influence of culture duration on MSC
MSC were expanded for up to 10 passages. MSC and CD34+ cells were seeded in cytokinefree
co-cultures after which the phenotype, clonogenic capacity and in vivo repopulating
activity of harvested hematopoietic cells were assessed.
Early passage MSC supported HPC expansion and differentiation toward both B lymphoid
and myeloid lineages. Late passage MSC did not support HPC and myeloid cell outgrowth
but maintained B cell supportive ability. In vitro maintenance of NOD/SCID mouse repopulating
cells cultured for one week in contact with MSC was effective until the fourth MSC
passage and declined afterwards. CD34+ cells achieved higher levels of engraftment in
NOD/SCID mice when co-injected with early passage MSC; however MSC expanded
beyond 9 passages were ineffective in promoting CD34+ cell engraftment. Non-contact cultures
indicated that MSC supportive activity involved diffusible factors. Among these,
interleukin (IL)-6 and IL-8 contributed to the supportive activity of early passage MSC but
not of late passage MSC. MSC phenotype as well as fat, bone and cartilage differentiation
capacity did not change during MSC culture.
Extended MSC culture alters their supportive ability toward HPC without concomitant
changes in phenotype and differentiation capacity.