Abstract
BackgroundMesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into bone, cartilage, fat and muscle cells and are being investigated for their utility in cell-based transplantation therapy. Yet, adequate methods to track transplanted MSCs in vivo are limited, precluding functional studies. Quantum Dots (QDs) offer an alternative to organic dyes and fluorescent proteins to label and track cells in vitro and in vivo. These nanoparticles are resistant to chemical and metabolic degradation, demonstrating long term photostability. Here, we investigate the cytotoxic effects of in vitro QD labeling on MSC proliferation and differentiation and use as a cell label in a cardiomyocyte co-culture.ResultsA dose-response to QDs in rat bone marrow MSCs was assessed in Control (no-QDs), Low concentration (LC, 5 nmol/L) and High concentration (HC, 20 nmol/L) groups. QD yield and retention, MSC survival, proinflammatory cytokines, proliferation and DNA damage were evaluated in MSCs, 24 -120 hrs post QD labeling. In addition, functional integration of QD labeled MSCs in an in vitro cardiomyocyte co-culture was assessed. A dose-dependent effect was measured with increased yield in HC vs. LC labeled MSCs (93 ± 3% vs. 50% ± 15%, p < 0.05), with a larger number of QD aggregates per cell in HC vs. LC MSCs at each time point (p < 0.05). At 24 hrs >90% of QD labeled cells were viable in all groups, however, at 120 hrs increased apoptosis was measured in HC vs. Control MSCs (7.2% ± 2.7% vs. 0.5% ± 0.4%, p < 0.05). MCP-1 and IL-6 levels doubled in HC MSCs when measured 24 hrs after QD labeling. No change in MSC proliferation or DNA damage was observed in QD labeled MSCs at 24, 72 and 120 hrs post labeling. Finally, in a cardiomyocyte co-culture QD labeled MSCs were easy to locate and formed functional cell-to-cell couplings, assessed by dye diffusion.ConclusionFluorescent QDs label MSC effectively in an in vitro co-culture model. QDs are easy to use, show a high yield and survival rate with minimal cytotoxic effects. Dose-dependent effects suggest limiting MSC QD exposure.
Highlights
Mesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into bone, cartilage, fat and muscle cells and are being investigated for their utility in cell-based transplantation therapy
As the mesenchymal stem cells (MSCs) proliferated in culture the number of Quantum Dots (QDs) labeled MSCs detected with flow cytometry decreased to 64% ± 12% vs. 25% ± 9% at 72 hrs and 48% ± 10% vs. 19% ± 10% at 120 hrs in the half (LC) and twice (HC) vs. LC MSCs (p < 0.05)
Our results suggest that labeled MSCs should be used within the first 24 hrs after QD labeling when evaluated in a co-culture system, as detection of QD labeled MSCs decreased as cells proliferated in culture
Summary
Mesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into bone, cartilage, fat and muscle cells and are being investigated for their utility in cell-based transplantation therapy. Quantum Dots (QDs) offer an alternative to organic dyes and fluorescent proteins to label and track cells in vitro and in vivo. These nanoparticles are resistant to chemical and metabolic degradation, demonstrating long term photostability. Cell transplantation therapy using adult derived bone marrow mesenchymal stem cells (MSCs) is currently being investigated as a potential therapy to treat injured heart tissue [1,2]. The mechanisms underlying MSC engraftment and electrical and mechanical integration with host cardiac tissue are not understood. In part, this is due to limited methods to track MSCs in vivo, precluding long-term functional studies of transplanted cells. Chemical and metabolic degradation, reduced photostability and signal quality [7] compromise in vitro and in vivo cell labeling and tracking
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