Dong-Ming Huang

Highly efficient cellular labeling of mesoporous nanoparticles in human mesenchymal stem cells: implication for stem cell tracking

Tracking the distribution of stem cells is crucial to their therapeutic use. However, the usage of current vectors in cellular labeling is restricted by their low internalizing efficiency. Here, we reported a cellular labeling approach with a novel vector composed of mesoporous silica nanoparticles (MSNs) conjugated with fluorescein isothiocyanate in human bone marrow mesenchymal stem cells and 3T3‐L1 cells, and the mechanism about fluorescein isothiocyanate‐conjugated MSNs (FITC‐MSNs) internalization was studied. FITC‐MSNs were efficiently internalized into mesenchymal stem cells and 3T3‐L1 cells even in short‐term incubation. The process displayed a time‐ and concentration‐dependent manner and was dependent on clathrin‐mediated endocytosis. In addition, clathrin‐dependent endocytosis seemed to play a decisive role on more internalization and longer stay of FITC‐MSNs in mesenchymal stem cells than in 3T3‐L1 cells. The internalization of FITC‐MSNs did not affect the cell viability, proliferation, immunophenotype, and differentiation potential of mesenchymal stem cells, and 3T3‐L1 cells. Finally, FITC‐MSNs could escape from endolysosomal vesicles and were retained the architectonic integrity after internalization. We conclude that the advantages of biocompatibility, durability, and higher efficiency in internalization suit MSNs to be a better vector for stem cell tracking than others currently used.

Mesoporous Silica Nanoparticles as a Delivery System of Gadolinium for Effective Human Stem Cell Tracking

The progress of using gadolinium (Gd)-based nanoparticles in cellular tracking lags behind that of superparamagnetic iron oxide (SPIO) nanoparticles in magnetic resonance imaging (MRI). Here, dual functional Gd-fluorescein isothiocyanate mesoporous silica nanoparticles (Gd-Dye@MSN) that possess green fluorescence and paramagnetism are developed in order to evaluate their potential as effective T1-enhancing trackers for human mesenchymal stem cells (hMSCs). hMSCs are labeled efficiently with Gd-Dye@MSN via endocytosis. Labeled hMSCs are unaffected in their viability, proliferation, and differentiation capacities into adipocytes, osteocytes, and chondrocytes, which can still be readily MRI detected. Imaging, with a clinical 1.5-T MRI system and a low incubation dosage of Gd, low detection cell numbers, and short incubation times is demonstrated on both loaded cells and hMSC-injected mouse brains. This study shows that the advantages of biocompatibility, durability, high internalizing efficiency, and pore architecture make MSNs an ideal vector of T1-agent for stem-cell tracking with MRI.

The promotion of human mesenchymal stem cell proliferation by superparamagnetic iron oxide nanoparticles

Superparamagnetic iron oxide (SPIO) nanoparticles are very useful in cell imaging; meanwhile, however, biosafety concerns associated with their use, especially on therapeutic stem cells, have arisen. Most studies of biosafety issues focus on whether the nanoparticles have deleterious effects. Here, we report that Ferucarbotran, an ionic SPIO, is not toxic to human mesenchymal stem cells (hMSCs) under the conditions of these experiments but instead increases cell growth. Ferucarbotran-promoted cell growth is due to its ability to diminish intracellular H2O2 through intrinsic peroxidase-like activity. Also, Ferucarbotran can accelerate cell cycle progression, which may be mediated by the free iron (Fe) released from lysosomal degradation and involves the alteration of Fe on the expression of the protein regulators of the cell cycle.

Aptamer-conjugated DNA icosahedral nanoparticles as a carrier of doxorubicin for cancer therapy.

DNA can be used to nanofabricate three-dimensional (3D) polyhedra. A variety of applications of 3D DNA assemblies have been proposed. Drug encasulation and intracellular delivery using DNA nanoparticles, however, have remained a challenge. Here, we create a distinct five-point-star motif and aptamer-conjugated six-point-star motif using well-used primer sequences to intermolecularly construct DNA icosahedra as a nanocarrier for doxorubicin. Aptamer-conjugated doxorubicin-intercalated DNA icosahedra (Doxo@Apt-DNA-icosa) show an efficient and specific internalization for killing epithelial cancer cells.

The inhibitory effect of superparamagnetic iron oxide nanoparticle (Ferucarbotran) on osteogenic differentiation and its signaling mechanism in human mesenchymal stem cells.

Superparamagnetic iron oxide (SPIO) nanoparticles are very useful for monitoring cell trafficking in vivo and distinguish whether cellular regeneration originated from an exogenous cell source, which is a key issue for developing successful stem cell therapies. However, the impact of SPIO labeling on stem cell behavior remains uncertain. Here, we show the inhibitory effect of Ferucarbotran, an ionic SPIO, on osteogenic differentiation and its signaling mechanism in human mesenchymal stem cells. Ferucarbotran caused a dose-dependent inhibition of osteogenic differentiation, abolished the differentiation at high concentration, promoted cell migration, and activated the signaling molecules, beta-catenin, a cancer/testis antigen, SSX, and matrix metalloproteinase 2 (MMP2). An iron chelator, desferrioxamine, suppressed all the above Ferucarbotran-induced actions, demonstrating an important role of free iron in the inhibition of osteogenic differentiation that is mediated by the promotion of cell mobilization, involving the activation of a specific signaling pathway.

Iron oxide nanoparticle-induced epidermal growth factor receptor expression in human stem cells for tumor therapy.

Superparamagnetic iron oxide (SPIO) nanoparticles show promise as labels for cellular magnetic resonance imaging (MRI) in the application of stem cell-based therapy. However, the unaddressed concerns about the impact of SPIO nanoparticles on stem cell attributes make the feasibility of SPIO labeling uncertain. Here, we show that the labeling of human mesenchymal stem cells (hMSCs) with ferucarbotran can induce epidermal growth factor receptor (EGFR) overexpression. Labeled hMSCs with their overexpressed EGFR were attracted by tumorous EGF and more effectively migrated toward tumor than unlabeled cells, resulting in more potent intrinsic antitumor activity. Moreover, the captured binding of tumorous EGF by overexpressed EGFR of labeled hMSCs blocked EGF/EGFR signaling-derived tumor growth, tumorous angiogenesis, and tumorous VEGF expression also responsible for tumor progression and development. Our results show that the impact of SPIO nanoparticles on stem cell attributes is not necessarily harmful but can be cleverly used to be beneficial to stem cell-based therapy.

Homologous RBC-derived vesicles as ultrasmall carriers of iron oxide for magnetic resonance imaging of stem cells.

Ultrasmall superparamagnetic iron oxide (USPIO) particles are very useful for cellular magnetic resonance imaging (MRI), which plays a key role in developing successful stem cell therapies. However, their low intracellular labeling efficiency, and biosafety concerns associated with their use, have limited their potential usage. In this study we develop a novel system composed of RBC-derived vesicles (RDVs) for efficient delivery of USPIO particles into human bone marrow mesenchymal stem cells (MSCs) for cellular MRI in vitro and in vivo. RDVs are highly biosafe to their autologous MSCs as manifested by cell viability, differentiation, and gene microarray assays. The data demonstrate the potential of RDVs as intracellular delivery vehicles for biomedical applications.

Dextran-coated iron oxide nanoparticles turn protumor mesenchymal stem cells (MSCs) into antitumor MSCs

Mesenchymal stem cells or multipotent mesenchymal stromal cells (MSCs) with tumor tropism have shown promise as a cellular deliverer of anticancer agents for cancer therapy. However, despite the fact that engineered MSCs have demonstrated profound anticancer effects, the exact intrinsic impact of MSCs themselves on tumor progression is still being debated. Therefore, in order to ensure pertinent MSC-based cancer therapy, it is essential to further explore the intrinsic antitumor and protumor effects of MSCs on tumor progression. Here, we show that dextran-coated iron oxide nanoparticles can not only increase the antitumor effect of antitumor MSCs but also reverse the protumor effect of protumor MSCs and hence turn protumor MSCs into antitumor MSCs in vivo, which can be attributed to the in vitro finding that dextran-coated iron oxide nanoparticles can not only inhibit the colony formation of cancer cells likely through induction of cytokine receptor expression to competitively capture the tumorous cytokine but also diminish cancer-mediated fibroblast differentiation and angiogenesis of protumor MSCs. This is the first report to demonstrate the induction of antitumor MSCs from protumor MSCs.

Ferucarbotran, a carboxydextran-coated superparamagnetic iron oxide nanoparticle, induces endosomal recycling, contributing to cellular and exosomal EGFR overexpression for cancer therapy

Superparamagnetic iron oxide (SPIO) nanoparticles have shown many impacts on stem cell attributes when they are used as labels for cellular magnetic resonance imaging (MRI) in the application of stem cell-based therapy. Although it is plausible that iron ions from the lysosomal degradation of SPIO nanoparticles are one of the possible candidates, the mechanisms underlying SPIO-induced cellular responses remain unclear. Herein, the mechanism of ferucarbotran, an ionic SPIO, for the regulation of epidermal growth factor receptor (EGFR) expression in human mesenchymal stem cells (hMSCs) is explored. Ferucarbotran can be internalized into EGFR-localized endosomes, and the endosomal EGFRs in ferucarbotran-labeled hMSCs, compared to unlabeled cells, are mainly localized on the early endosomes and recycling endosomes, but not on late endosomes/lysosomes, and thus escape from lysosomal degradation. Afterward, the recycling endosomal EGFRs are transferred to the cellular membrane and extracellular exosomal vesicles (exosomes) through back fusion and a secretory pathway, respectively, resulting in EGFR-overexpressed hMSCs and EGFR-overexpressed exosomes. Moreover, as EGFR-overexpressed hMSCs, EGFR-overexpressed exosomes can more effectively capture tumorous EGF than native exosomes, which may contribute to the inhibition of tumor growth. This is the first report to find that the SPIO nanoparticles have an impact on stem cell attributes through inducing endosomal recycling instead of them undergoing lysosomal degradation.