Cytokine preconditioning of engineered cartilage provides protection against interleukin-1 insult

Research reported in this publication was supported in part by the National Institute of Arthritis and Musculoskeletal and Skin Diseases and National Institute of Biomedical Imaging and Bioengineering of the National Institutes of Health under Award Number R01AR60361, R01AR061988, P41EB002520). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. ART was supported by a National Science Foundation Graduate Fellowship

Passage-dependent relationship between mesenchymal stem cell mobilization and chondrogenic potential

ObjectiveGalvanotaxis, the migratory response of cells in response to electrical stimulation, has been implicated in development and wound healing. The use of mesenchymal stem cells (MSCs) from the synovium (synovium-derived stem cells, SDSCs) has been investigated for repair strategies. Expansion of SDSCs is necessary to achieve clinically relevant cell numbers; however, the effects of culture passage on their subsequent cartilaginous extracellular matrix production are not well understood.MethodsOver four passages of SDSCs, we measured the expression of cell surface markers (CD31, CD34, CD49c, CD73) and assessed their migratory potential in response to applied direct current (DC) electric field. Cells from each passage were also used to form micropellets to assess the degree of cartilage-like tissue formation.ResultsExpression of CD31, CD34, and CD49c remained constant throughout cell expansion; CD73 showed a transient increase through the first two passages. Correspondingly, we observed that early passage SDSCs exhibit anodal migration when subjected to applied DC electric field strength of 6 V/cm. By passage 3, CD73 expression significantly decreased; these cells exhibited cell migration toward the cathode, as previously observed for terminally differentiated chondrocytes. Only late passage cells (P4) were capable of developing cartilage-like tissue in micropellet culture.ConclusionsOur results show cell priming protocols carried out for four passages selectively differentiate stem cells to behave like chondrocytes, both in their motility response to applied electric field and their production of cartilaginous tissue

Modulatory profiling identifies mechanisms of small molecule-induced cell death

Cell death is a complex process that plays a vital role in development, homeostasis, and disease. Our understanding of and ability to control cell death is impeded by an incomplete characterization of the full range of cell death processes that occur in mammalian systems, especially in response to exogenous perturbations. We present here a general approach to address this problem, which we call modulatory profiling. Modulatory profiles are composed of the changes in potency and efficacy of lethal compounds produced by a second cell death-modulating agent in human cell lines. We show that compounds with the same characterized mechanism of action have similar modulatory profiles. Furthermore, clustering of modulatory profiles revealed relationships not evident when clustering lethal compounds based on gene expression profiles alone. Finally, modulatory profiling of compounds correctly predicted three previously uncharacterized compounds to be microtubule-destabilizing agents, classified numerous compounds that act nonspecifically, and identified compounds that cause cell death through a mechanism that is morphologically and biochemically distinct from previously established ones

Passage-dependent relationship between mesenchymal stem cell mobilization and chondrogenic potential

ObjectiveGalvanotaxis, the migratory response of cells in response to electrical stimulation, has been implicated in development and wound healing. The use of mesenchymal stem cells (MSCs) from the synovium (synovium-derived stem cells, SDSCs) has been investigated for repair strategies. Expansion of SDSCs is necessary to achieve clinically relevant cell numbers; however, the effects of culture passage on their subsequent cartilaginous extracellular matrix production are not well understood.MethodsOver four passages of SDSCs, we measured the expression of cell surface markers (CD31, CD34, CD49c, CD73) and assessed their migratory potential in response to applied direct current (DC) electric field. Cells from each passage were also used to form micropellets to assess the degree of cartilage-like tissue formation.ResultsExpression of CD31, CD34, and CD49c remained constant throughout cell expansion; CD73 showed a transient increase through the first two passages. Correspondingly, we observed that early passage SDSCs exhibit anodal migration when subjected to applied DC electric field strength of 6 V/cm. By passage 3, CD73 expression significantly decreased; these cells exhibited cell migration toward the cathode, as previously observed for terminally differentiated chondrocytes. Only late passage cells (P4) were capable of developing cartilage-like tissue in micropellet culture.ConclusionsOur results show cell priming protocols carried out for four passages selectively differentiate stem cells to behave like chondrocytes, both in their motility response to applied electric field and their production of cartilaginous tissue

Human chondrocyte migration behaviour to guide the development of engineered cartilage

Tissue-engineering techniques have been successful in developing cartilage-like tissues in vitro using cells from animal sources. The successful translation of these strategies to the clinic will likely require cell expansion to achieve sufficient cell numbers. Using a two-dimensional (2D) cell migration assay to first identify the passage at which chondrocytes exhibited their greatest chondrogenic potential, the objective of this study was to determine a more optimal culture medium for developing three-dimensional (3D) cartilage-like tissues using human cells. We evaluated combinations of commonly used growth factors that have been shown to promote chondrogenic growth and development. Human articular chondrocytes (AC) from osteoarthritic (OA) joints were cultured in 3D environments, either in pellets or encapsulated in agarose. The effect of growth factor supplementation was dependent on the environment, such that matrix deposition differed between the two culture systems. ACs in pellet culture were more responsive to bone morphogenetic protein (BMP2) alone or combinations containing BMP2 (i.e. BMP2 with PDGF or FGF). However, engineered cartilage development within agarose was better for constructs cultured with TGFβ3. These results with agarose and pellet culture studies set the stage for the development of conditions appropriate for culturing 3D functional engineered cartilage for eventual use in human therapies. Copyright © 2015 John Wiley & Sons, Ltd