Stem Cell Technology: mesenchymal stem cells, clonal heterogeneity, cell signaling, aging and regenerative therapies
My research is in the area of stem cell technology with the goal of improving human health through advances in regenerative medicine. Currently, my research focuses on human mesenchymal stem cells (MSCs). These are highly robust cells with broad differentiation potential that regulate the immune response and migrate to injured tissue, among other therapeutic properties. As such, these adult stem cells have potential application to treat a wide range of disorders including arthritis, heart attack and cancer. The scope of my research projects ranges from fundamental discovery at the cellular and molecular levels to computational analysis that resolves complex interactions among cells and signaling pathways. With both approaches, our objective is to gain unique insight into the mechanisms by which stem cells interact with their surroundings and to employ this knowledge to develop novel regenerative therapies. This research is inherently interdisciplinary and provides opportunities to collaborate with stem cell biologists and clinicians.
Below, I summarized recent accomplishments that my research group has made in the area of stem cell technology. We have made several important contributions to cytokine regulation of migration, clonal heterogeneity in potency and proliferation potential of MSCs. References are provided for representative articles on these topics.
Fig. 1. Ribbon structure of MIF. L. Leng
and R. Bucala, Cell Research 16,
The efficacy of stem cell therapies is dependent on the ability of stem cells to migrate to injured tissue. Our study was innovative in providing the first report of a protein inhibitor of MSC migration: macrophage migration inhibitory factor (MIF, Fig. 1). Moreover, this was the first report of the effects of MIF and its antagonist on the function of any type of stem cell or progenitor. Stem cells may encounter MIF during times of injury and stress and, thus, MIF may be relevant to a variety of MSC therapies. The anti-inflammatory agent ISO-1, a small-molecule MIF antagonist, restored MSC migration for all donors to levels found in the absence of MIF in serum-free medium and increased migration to conditioned medium by ≥3-fold in all cases. An activating CD74 antibody was employed to examine the effect of the CD74 MIF receptor on MSC motility. CD74 activation inhibits in a dose-dependent manner up to 90% of in vitro MSC migration, indicating that MIF may act on MSCs, at least in part, through CD74. MIF and ISO-1 may have utility to control MSC migration to and retention in injured tissue.
Representative article: Barrilleaux, Phinney, Fischer-Valuck, Russell, Wang, Prockop and O’Connor. Small-molecule antagonist of macrophage migration inhibitory factor enhances migratory response of mesenchymal stem cells to bronchial epithelial cells. Tissue Eng Part A, 15: 2335-46 (2009).
Heterogeneity in Potency
MSCs are a heterogeneous mixture of cell populations at different stages of lineage commitment. Donor variation in the composition of MSC populations in culture has a significant impact on the efficacy of MSC therapies. An obstacle to quantify this heterogeneity is the absence of an immunophenotype of cell-surface markers that specifies the differentiation potential (or potency) of MSCs. We circumvented this obstacle by developing a unique high-capacity assay based on functional differentiation (Fig. 2A). The assay utilizes a 96-well format to (1) generate clones by limiting dilution, (2) differentiate matched clonal colonies to quantify trilineage potential to exhibit adipo-, chondro- and osteogenesis as a measure potency, and (3) cryopreserve clones of known potency. When the assay was applied to MSCs from two donors with comparable colony-forming efficiencies (55-62%), all 8 possible categories of trilineage potential were observed (Fig. 2B). This assay has enabled us to elucidate regenerative properties of MSCs as a function of their potency and to identify novel cell-surface markers of MSC potency.
Representative article: Russell, Phinney, Lacey, Barrilleaux, Meyertholen and O'Connor. In vitro high-capacity assay to quantify the clonal heterogeneity in trilineage potential of mesenchymal stem cells reveals a complex hierarchy of lineage commitment. Stem Cells, 28: 788-98 (2010).[Among Top 50 Downloaded Articles in Stem Cells for March 2010]
Fig. 2. High-capacity assay to quantify MSC potency: (A) overview of the assay, and
(B) heterogeneity in potency of 96 MSC clones from two donors. MSCs exhibit
osteo (O), adipo (A) and/or chondrogenesis (C).
Fig. 3.Colony-forming efficiency of MSC clones
as a function of potency. Nomenclature:
See legend to Figure 2. Median (line, n = 5 clones).
*P<0.01 vs. OAC clones.
Ex vivo proliferation to produce sufficient quantities of MSCs is essential for most therapeutic applications of these stem cells. Clonal analysis of distinct MSC populations with the high-capacity assay described in the previous section resolved the relationship between proliferation potential of MSCs and their potency. Tripotent MSC clones are highly proliferative in terms of their colony-forming efficiency and capacity for ex vivo expansion. Bipotent osteo-adipogenic and osteo-chondrogenic clones have similar proliferation potentials. Lineage-committed, unipotent clones are slow-growing, senescent and, to a lesser extent, apoptotic. Our results are germane to the ex vivo expansion of MSCs for clinical applications; use of colony-forming efficiency to monitor the content of multipotent cells in MSC therapies and predict their efficacy; enrichment of multipotent MSCs; and preparation of MSC therapies to treat the elderly and non-union bone fractures, when accumulation of senescent MSCs in bone is likely.
Representative article: Russell, Lacey, Gilliam, Tucker, Phinney and O'Connor. Clonal analysis of proliferation potential of human bone marrow mesenchymal stem cells as a function of potency. Biotechnol Bioeng, 108: 2716-26 (2011). [FEATURED ARTICLE]
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