Xiyun Yan

Intrinsic peroxidase-like activity of ferromagnetic nanoparticles

Nanoparticles containing magnetic materials, such as magnetite (Fe3O4), are particularly useful for imaging and separation techniques. As these nanoparticles are generally considered to be biologically and chemically inert, they are typically coated with metal catalysts, antibodies or enzymes to increase their functionality as separation agents. Here, we report that magnetite nanoparticles in fact possess an intrinsic enzyme mimetic activity similar to that found in natural peroxidases, which are widely used to oxidize organic substrates in the treatment of wastewater or as detection tools. Based on this finding, we have developed a novel immunoassay in which antibody-modified magnetite nanoparticles provide three functions: capture, separation and detection. The stability, ease of production and versatility of these nanoparticles makes them a powerful tool for a wide range of potential applications in medicine, biotechnology and environmental chemistry.

Magnetoferritin nanoparticles for targeting and visualizing tumour tissues

Engineered nanoparticles have been used to provide diagnostic, therapeutic and prognostic information about the status of disease. Nanoparticles developed for these purposes are typically modified with targeting ligands (such as antibodies, peptides or small molecules) or contrast agents using complicated processes and expensive reagents. Moreover, this approach can lead to an excess of ligands on the nanoparticle surface, and this causes non-specific binding and aggregation of nanoparticles, which decreases detection sensitivity. Here, we show that magnetoferritin nanoparticles (M-HFn) can be used to target and visualize tumour tissues without the use of any targeting ligands or contrast agents. Iron oxide nanoparticles are encapsulated inside a recombinant human heavy-chain ferritin (HFn) protein shell, which binds to tumour cells that overexpress transferrin receptor 1 (TfR1). The iron oxide core catalyses the oxidation of peroxidase substrates in the presence of hydrogen peroxide to produce a colour reaction that is used to visualize tumour tissues. We examined 474 clinical specimens from patients with nine types of cancer and verified that these nanoparticles can distinguish cancerous cells from normal cells with a sensitivity of 98% and specificity of 95%.

Fe3O4 Magnetic Nanoparticle Peroxidase Mimetic-Based Colorimetric Assay for the Rapid Detection of Organophosphorus Pesticide and Nerve Agent

Rapid and sensitive detection methods are in urgent demand for the screening of extensively used organophosphorus pesticides and highly toxic nerve agents for their neurotoxicity. In this study, we developed a novel Fe(3)O(4) magnetic nanoparticle (MNP) peroxidase mimetic-based colorimetric method for the rapid detection of organophosphorus pesticides and nerve agents. The detection assay is composed of MNPs, acetylcholinesterase (AChE), and choline oxidase (CHO). The enzymes AChE and CHO catalyze the formation of H(2)O(2) in the presence of acetylcholine, which then activates MNPs to catalyze the oxidation of colorimetric substrates to produce a color reaction. After incubation with the organophosphorus neurotoxins, the enzymatic activity of AChE was inhibited and produced less H(2)O(2), resulting in a decreased catalytic oxidation of colorimetric substrates over MNP peroxidase mimetics, accompanied by a drop in color intensity. Three organophosphorus compounds were tested on the assay: acephate and methyl-paraoxon as representative organophosphorus pesticides and the nerve agent Sarin. The novel assay displayed substantial color change after incubation in organophosphorus neurotoxins in a concentration-dependent manner. As low as 1 nM Sarin, 10 nM methyl-paraoxon, and 5 μM acephate are easily detected by the novel assay. In conclusion, by employing the peroxidase-mimicking activity of MNPs, the developed colorimetric assay has the potential of becoming a screening tool for the rapid and sensitive assessment of the neurotoxicity of an overwhelming number of organophosphate compounds.

Carbon nanotube delivery of the GFP gene into mammalian cells.

Exogenous-gene expression and manipulation in mammalian cells has become a mainstay of biomedical research. Consequently, improving methods for efficient gene transfer to a broad range of cell types is of great interest and remains a high priority. Several classes of transfection methods have been developed, which include traditional cationic moleculemediated agents, such as Lipofectamine 20000 and FuGENE 6, viral-vector systems, and the “gene gun” approach. With the rapid development of nanobiotechnology, a variety of new materials, such as gold nanoparticles, silica nanoparticles, polymers, nanogels, and dendrimers have been investigated as biocompatible transporters. Recently, carbon nanotube—a well-studied nanomaterial— have been investigated for their ability to interact with and affect living systems. For instance, carbon nanotubes have been found to enhance DNA amplification in PCR and affect the growth pattern of neurons. Pantarotto et al. have reported the internalization of fluorescein isothiocynate (FITC) labeled nanotubes and nanotube delivery of the gene that encodes b-galactosidase into cells, with no apparent toxic effects. Kam et al. have studied the mechanism of protein-conjugated carbon nanotube uptake into cells via the endocytic pathway. Here we present our finding that amino-functionalized multiwalled carbon nanotubes (NH2-MWCNTs) are able to interact with plasmid DNA and deliver the green fluorescence protein (GFP) gene into cultured human cells. Our data strongly suggest that carbon nanotubes can be considered as a new carrier for the delivery of biomolecules, such as DNA, proteins, and peptides into mammalian cells. Therefore, this novel system might have potential applications in biology and therapy, including vaccine and gene delivery. In order to increase their biocompatibility, we introduced amino-, carboxyl-, hydroxyl-, and alkyl groups onto the surface of MWCNTs. COOH-MWCNTs were first prepared by nitric / sulfuric acid oxidation, and then NH2and CH3CH2CH2-groups were added. Finally, we obtained four types of MWCNTs with different chemical groups on their surface. Functionalized MWCNTs were observed under an electron microscope and were found to be 60–70 nm in diameter and 1–2 mm in length. Although we did not find a significant difference in size between the NH2-MWCNTs and NH2-MWCNT–DNAs, the latter appeared to have the tendency to aggregate (Figure 1B). In order to test the DNA-binding ability of amino-, carboxyl-, hydroxyl-, and alkyl-group-modified MWCNTs, we incubated them with pEGFPN1-plasmid DNA, and MWCNT–DNA mixtures were analyzed by agarose-gel electrophoresis. The results show that only NH2-MWCNT bound to DNA (Figure 2); since the NH2-MWCNT–DNA complex was too big to run into the

CD146, a multi-functional molecule beyond adhesion.

CD146 is a cell adhesion molecule (CAM) that is primarily expressed at the intercellular junction of endothelial cells. CD146 was originally identified as a tumor marker for melanoma (MCAM) due to its existence only in melanoma but not in the corresponding normal counterpart. However CD146 is not just a CAM for the inter-cellular and cell-matrix adhesion. Recent evidence indicates that CD146 is actively involved in miscellaneous processes, such as development, signaling transduction, cell migration, mesenchymal stem cells differentiation, angiogenesis and immune response. CD146 has increasingly become an important molecule, especially identified as a novel bio-marker for angiogenesis and for cancer. Here we have reviewed the dynamic research of CD146, particularly newly identified functions and the underlying mechanisms of CD146.

CD146, an epithelial-mesenchymal transition inducer, is associated with triple-negative breast cancer

The epithelial-mesenchymal transition (EMT) plays an important role in breast cancer metastasis, especially in the most aggressive and lethal subtype, “triple-negative breast cancer” (TNBC). Here, we report that CD146 is a unique activator of EMTs and significantly correlates with TNBC. In epithelial breast cancer cells, overexpression of CD146 down-regulated epithelial markers and up-regulated mesenchymal markers, significantly promoted cell migration and invasion, and induced cancer stem cell-like properties. We further found that RhoA pathways positively regulated CD146-induced EMTs via the key EMT transcriptional factor Slug. An orthotopic breast tumor model demonstrated that CD146-overexpressing breast tumors showed a poorly differentiated phenotype and displayed increased tumor invasion and metastasis. We confirmed these findings by conducting an immunohistochemical analysis of 505 human primary breast tumor tissues and found that CD146 expression was significantly associated with high tumor stage, poor prognosis, and TNBC. CD146 was expressed at abnormally high levels (68.9%), and was strongly associated with E-cadherin down-regulation in TNBC samples. Taken together, these findings provide unique evidence that CD146 promotes breast cancer progression by induction of EMTs via the activation of RhoA and up-regulation of Slug. Thus, CD146 could be a therapeutic target for breast cancer, especially for TNBC.

H-ferritin–nanocaged doxorubicin nanoparticles specifically target and kill tumors with a single-dose injection

Significance Nanoparticles capable of specifically binding to target cells and delivering high doses of therapeutic drugs with optimized safety profiles are much sought after in the nanomedical field. Here, we developed a natural H-ferritin (HFn) nanocarrier that specifically delivered a high concentration of the therapeutic drug doxorubicin (Dox) to tumor cells and significantly inhibited tumor growth with a single-dose treatment while also showing excellent biocompatibility and safety profiles in murine cancer models. Compared with the clinically approved liposomal Dox (Doxil), HFn-Dox exhibited longer median survival times and lower toxicity when administered at the same dose in all tumor models studied. An ideal nanocarrier for efficient drug delivery must be able to target specific cells and carry high doses of therapeutic drugs and should also exhibit optimized physicochemical properties and biocompatibility. However, it is a tremendous challenge to engineer all of the above characteristics into a single carrier particle. Here, we show that natural H-ferritin (HFn) nanocages can carry high doses of doxorubicin (Dox) for tumor-specific targeting and killing without any targeting ligand functionalization or property modulation. Dox-loaded HFn (HFn-Dox) specifically bound and subsequently internalized into tumor cells via interaction with overexpressed transferrin receptor 1 and released Dox in the lysosomes. In vivo in the mouse, HFn-Dox exhibited more than 10-fold higher intratumoral drug concentration than free Dox and significantly inhibited tumor growth after a single-dose injection. Importantly, HFn-Dox displayed an excellent safety profile that significantly reduced healthy organ drug exposure and improved the maximum tolerated dose by fourfold compared with free Dox. Moreover, because the HFn nanocarrier has well-defined morphology and does not need any ligand modification or property modulation it can be easily produced with high purity and yield, which are requirements for drugs used in clinical trials. Thus, these unique properties make the HFn nanocage an ideal vehicle for efficient anticancer drug delivery.

CD146 is a coreceptor for VEGFR-2 in tumor angiogenesis

CD146 is a novel endothelial biomarker and plays an essential role in angiogenesis; however, its role in the molecular mechanism underlying angiogenesis remains poorly understood. In the present study, we show that CD146 interacts directly with VEGFR-2 on endothelial cells and at the molecular level and identify the structural basis of CD146 binding to VEGFR-2. In addition, we show that CD146 is required in VEGF-induced VEGFR-2 phosphorylation, AKT/p38 MAPKs/NF-κB activation, and thus promotion of endothelial cell migration and microvascular formation. Furthermore, we show that anti-CD146 AA98 or CD146 siRNA abrogates all VEGFR-2 activation induced by VEGF. An in vivo angiogenesis assay showed that VEGF-promoted microvascular formation was impaired in the endothelial conditional knockout of CD146 (CD146(EC-KO)). Our animal experiments demonstrated that anti-CD146 (AA98) and anti-VEGF (bevacizumab) have an additive inhibitory effect on xenografted human pancreatic and melanoma tumors. The results of the present study suggest that CD146 is a new coreceptor for VEGFR-2 and is therefore a promising target for blocking tumor-related angiogenesis.

A magnetic protein biocompass

The notion that animals can detect the Earths magnetic field was once ridiculed, but is now well established. Yet the biological nature of such magnetosensing phenomenon remains unknown. Here, we report a putative magnetic receptor (Drosophila CG8198, here named MagR) and a multimeric magnetosensing rod-like protein complex, identified by theoretical postulation and genome-wide screening, and validated with cellular, biochemical, structural and biophysical methods. The magnetosensing complex consists of the identified putative magnetoreceptor and known magnetoreception-related photoreceptor cryptochromes (Cry), has the attributes of both Cry- and iron-based systems, and exhibits spontaneous alignment in magnetic fields, including that of the Earth. Such a protein complex may form the basis of magnetoreception in animals, and may lead to applications across multiple fields.

The promotion of human malignant melanoma growth by mesoporous silica nanoparticles through decreased reactive oxygen species

The concept that mesoporous silica nanoparticles (MSNs) are regarded as ideal novel drug delivery carriers in tumor therapy has been introduced extensively, but the effects of MSNs on tumor growth have received little attention. Here a model of nude mice xenografted with human malignant melanoma cells (A375) was used to investigate the effect of MSNs on tumor growth. Surprisingly, we found that MSNs have no toxicity to human malignant melanoma but increasing tumor growth in vivo. It was also confirmed that MSNs significantly promoted A375 cell proliferation and accelerated cell cycle progression in vitro. Cellular uptake mechanism showed that MSNs may affect molecular behavior of A375 cells when they entered into cytoplasm. Then, a detailed mechanism indicated that the promotion effect induced by MSNs was due to the decreasing of endogenous reactive oxygen species (ROS) in cells. Further results demonstrated that the upregulation of anti-apoptotic molecules Bcl-2 expression and the inhibition of NF-kappaB activation by MSNs may promote cell proliferation in a redox-sensitive signal pathway. These results show that tumor growth can be regulated by nanocarriers themselves in a ROS-dependent manner and imply that nanocarriers are not necessarily suitable for all kinds of tumor therapy in development drug delivery system.