The inhibition of osteogenesis with human bone marrow mesenchymal stem cells by CdSe/ZnS quantum dot labels
Introduction
Quantum dots (QDs) have recently been explored as labeling agents of cells and tissues for biological imaging. As compared to conventional organic probes, these fluorescent agents can be excited by a wider range of wavelengths and exhibit narrower emission bandwidths [1], [2], [3]. Of particular interests are the uniquely strong luminance and high photostability exhibited by CdSe/ZnS [4]. Various techniques, such as microinjection, electroporation, scraping, transferrin-assisted transport and liposome-mediated transfection, have been employed to deliver QDs into cells [5]. Despite the increasing popularity of QDs as cell labeling agents, their cytotoxicity and the possibility of aberrant effects on gene expression remain a major concern.
A number of studies indicate that, as a fluorescent probe for cultured cells and animal models, the internalized QDs have no apparent deleterious effects on the morphology and the growth of cells. For example, it has been shown that when QDs are introduced into Xenopus [1] or HeLa cells [6], normal growth and differentiation of the cells are not affected. However, that conclusion, derived from morphological observations and analysis based on gross growth properties of cells, omitted any detailed characterization of cellular activities at the molecular level, such as gene expression. In addition, the cells employed in the above-mentioned studies are either non-mammalian or cancer cells. In the present study, we used human bone marrow mesenchymal stem cells (hBMSCs) as a cell model to explore the effect of QDs on the growth and induced osteogenesis of the cells. A number of criteria including enzyme activity and marker gene expression were examined to evaluate the process of osteogenesis. We found that despite a normal cell growth and cell cycle distribution displayed by the QD-containing cells, the expression of osteocyte specific marker genes, osteopontin and osteocalcin, and the activity of alkaline phosphatase (ALP) are significantly suppressed by the introduction of QDs into hBMSCs. This is the first report showing that the uptake of QDs may affect cellular responses to extracellular stimuli, suggesting that the effects of QDs on various aspects of cell activities should be carefully evaluated when used as a long-term cell tracer.
Section snippets
Cell culture and the delivery of QDs into hBMSCs
hBMSCs immortalized with HPV16 E6/E7 [7] were cultured in 10-cm dishes containing DMEM-LG (GIBCO) with 100 U/ml penicillin, 10 μg/ml streptomycin and 10% fetal bovine serum (FBS) (GIBCO), the cells were maintained in a humidified cell culture incubator at 37 °C with 5% CO2/95% air. Cells were subcultured at a 1:3 ratio every 5 days. For transfection, 7×105 cells were seeded in 10-cm dishes for 24 h and transfected with 1.625 μg of CdSe/ZnS (Ocean Optics, Carboxyl EviTag with emission of 520 nm) using
Delivery of CdSe/ZnS QDs into hBMSCs
We first explored the techniques for introducing CdSe/ZnS QDs into hBMSCs. It has been shown that QDs can be delivered into cells by various methods including microinjection, electroporation, scraping and liposomal transfection [1], [5], [8], [9]. Uptake of these particles into cells via endocytosis could also be facilitated by surface modification of the QDs. At high concentrations, QDs could significantly increase osmotic pressure, which is detrimental to cells. In the present study, a low
Discussion
Because of the characteristic strong luminance and high photostability exhibited by CdSe/ZnS QDs, extensive studies have been directed toward the potential application of these fluorophores in labeling cells. It is therefore important to explore methods of the QD delivery into cells and the effects on various aspects of cellular responses once internalized. We have successfully delivered CdSe/ZnS QDs into hBMSCs with high efficiency and demonstrated that the hydrophilic QDs remain detectable in
Conclusions
QDs have emerged as fluorescent agents for labeling intracellular targets and tracing live cells, it is therefore important to explore methods of QD delivery into cells and examine their effects on various aspects of cellular activities once internalized. We have compared different methods of transporting CdSe/ZnS QDs into hBMSCs, and shown that the liposome-mediated transportation attains the highest delivery efficiency. The fluorophores remain detectable for at least 3 weeks with or without
Acknowledgments
This work was supported by Grant NSC 92-2120-M-010-001 from the National Science Council, Taipei, Taiwan, Republic of China.
References (18)
- et al.
Applications of T-lymphoma labeled with fluorescent quantum dots to cell tracing markers in mouse body
Biochem Biophys Res Commun
(2004) - et al.
Optical coding of mammalian cells using semiconductor quantum dots
Anal Biochem
(2004) - et al.
Semiconductor quantum dot/albumin complex is a long-life and highly photostable endosome marker
Biochem Biophys Res Commun
(2003) - et al.
NGF signaling from clathrin-coated vesicles: evidence that signaling endosomes serve as a platform for the Ras-MAPK pathway
Neuron
(2001) - et al.
In vivo imaging of quantum dots encapsulated in phospholipid micelles
Science
(2002) - et al.
Diffusion dynamics of glycine receptors revealed by single-quantum dot tracking
Science
(2003) - et al.
Conjugation of luminescent quantum dots with antibodies using an engineered adaptor protein to provide new reagents for fluoroimmunoassays
Anal Chem
(2002) - et al.
Quantum dot bioconjugates for ultrasensitive nonisotopic detection
Science
(1998) - et al.
Long-term multiple color imaging of live cells using quantum dot bioconjugates
Nat Biotechnol
(2003)
Cited by (115)
Mesenchymal stem cell-based nanoparticles and scaffolds in regenerative medicine
2022, European Journal of PharmacologyNanotechnology shaping stem cell therapy: Recent advances, application, challenges, and future outlook
2021, Biomedicine and PharmacotherapyInteraction of L-cysteine functionalized CdSe quantum dots with metallic cations and selective binding of cobalt in water probed by fluorescence
2017, Sensors and Actuators, B: ChemicalInfluence of heavy nanocrystals on spermatozoa and fertility of mammals
2016, Materials Science and Engineering CCarbon Quantum Dots as Multi-Purpose Nanomaterial in Stem Cell Therapy
2023, Chemistry and Biodiversity