Biological characterization of long-term cultured human mesenchymal stem cells

Abstract

Human mesenchymal stem cells (hMSCs) have generated a great deal of interest in clinical applications. The reason is that they may have the plasticity needed to differentiate into multiple lineages and the ability to expand ex vivo. For the therapeutic applications of hMSCs to be of practical use, it is crucial to assess the efficacy and safety of hMSCs in long-term ex vivo expansion. In this study, we cultured hMSCs by population doubling (PD) 60, and investigated their growth, osteogenic and adipogenic differential abilities, change of surface markers, telomerase activity, telomere length, and gene expression related to tumorigenesis. An in vivo tumorigenesis assay was also carried out. In long-term expanded hMSCs, the cells became aged above PD 30 and their adipogenic and osteogenic differentiation potential decreased. Telomerase activity unchanged whereas telomere length decreased and karyotypes were not changed. Gene expressions related to tumorigenesis decreased in proportion as the PD of hMSCs increased. In vivo transplantation of long-term cultured hMSCs to nude mice did not result in tumor formation. These findings suggest that diverse tests for cellular therapy should be considered during the ex vivo culture of hMSCs, particularly when a prolonged and extended propagation period is required.

Human Embryonic Mesodermal Progenitors Highly Resemble Human Mesenchymal Stem Cells and Display High Potential for Tissue Engineering Applications

“…Decreased proliferative potential of hMSCs during long-term in vitro culture has earlier been demonstrated and has to some extent been explained by the decreasing telomere length. 48 The high proliferative potential of the hES-MPs is further in accordance with the presence of longer telomeres compared with the hMSCs, while no differences in telomerase activity was detected. The differences in telomere length observed between hES-MPs and hMSCs may be associated with the intrinsic different source of the two cell types.…”

Stromal-Cell-Derived Extracellular Matrix Promotes the Proliferation and Retains the Osteogenic Differentiation Capacity of Mesenchymal Stem Cells on Three-Dimensional Scaffolds

“…Expansion of MSCs on 2D substrates is problematic as the cells spontaneously differentiate into more committed cell lineages and gradually ''lose'' their stem cell properties (''stemness''). 1,2 Further, with extensive passaging the cells senesce due to DNA damage and can transform to become cancerous. A recent study reported spontaneous malignant transformation of nearly 50% of MSCs in long-term cultures.…”

Reconstitution of Marrow-Derived Extracellular Matrix Ex Vivo: A Robust Culture System for Expanding Large-Scale Highly Functional Human Mesenchymal Stem Cells

“…However, a major bottleneck in clinical application of MSCs has been their limited number, because they are rare in the primary tissue (∼0.001%) [ 7 ]. With extensive passaging, MSCs often lose multilineage differentiation potential [8][9][10][11]. Moreover, it has been reported that expansion of human and mouse MSCs is accompanied by cellular senescence and outgrowth of transformed cells, though transformation is less frequent in cultured human MSCs [12][13][14][15][16].…”