Recent research efforts about iron oxide nanoparticles has focused on the development of iron oxide-based T contrast agents for magnetic resonance imaging (MRI), such as ultrasmall iron oxide nanospheres (USNPs <4 nm) and ultrathin nanowires (NW, diameter <4 nm). In this paper, we report the cellular uptake behaviors of these two types of ultrasmall scale nanostructures on HepG2 cells. Both these two nanostructures were functionalized with tannic acid and their physical and chemical properties were carefully analyzed before cellular tests. Both USNPs and NWs exhibited strong paramagnetic signals, a property suitable for T MRI contrast agents. The distinct shapes also caused much difference in their cellular uptake behaviors. Specifically, the uptake of USNPs was five times higher than that of NWs after 72 hours incubation. The shape-dependent cellular uptake can potentially lead to different blood circulation times, and subsequently different applications of these two types of ultrasmall nanostructures.

We have studied the expression of 1l-myoinositol-1-phosphate synthase (MIPS; EC 5.5.1.4) in developing organs of Phaseolus vulgaris to define genetic controls that spatially regulate inositol phosphate biosynthesis. MIPS, the pivotal biosynthetic enzyme in inositol metabolism, is the only enzyme known to catalyze the conversion of glucose 6-phosphate to inositol phosphate. It is found in unicellular and multicellular eukaryotes and has been isolated as a soluble enzyme from both. Thus, it is widely accepted that inositol phosphate biosynthesis is largely restricted to the cytosol. Here, we report findings that suggest the enzyme is also expressed in membrane-bound organelles. Microscopic and biochemical analyses detected MIPS expression in plasma membranes, plastids, mitochondria, endoplasmic reticula, nuclei, and cell walls of bean. To address mechanisms by which the enzyme could be targeted to or through membranes, MIPS genes were analyzed for sorting signals within primary structures and upstream open reading frames that we discovered through our sequence analyses. Comprehensive computer analyses revealed putative transit peptides that are predicted to target the enzyme to different cellular compartments. Reverse transcriptase PCR experiments suggest that these putative targeting peptides are expressed in bean roots and leaves.Inositol metabolism is essential for the development of plants, animals, and some microorganisms. Metabolites of inositol (a six carbon cyclitol) such as phosphoinositides and phytate function as potent regulators of signal transduction for a wide variety of hormones, growth factors, and neurotransmitters (York et al., 1999;Loewus and Murthy, 2000; Irvine and Schell, 2001). Inositol phosphate, the immediate precursor of free inositol, is synthesized via the internal cyclization of Glc 6-phosphate. The overall reaction mechanism consists of a tightly coupled oxidation and reduction (Sherman et al., 1969;Loewus and Loewus, 1983). 1l-Myoinositol-1-phosphate synthase (MIPS; EC 5.5.1.4) is the only enzyme known to catalyze this reaction. MIPS is found in diverse organisms, both eukaryotic and prokaryotic, suggesting that the pathway for inositol 1-phosphate biosynthesis from Glc 6-phosphate arose early in the evolution of life (Majumder et al., 1997; Bachhawat and Mande, 2000). The properties and generally accepted catalytic mechanisms of the enzyme are similar in all organisms where such assessment has been undertaken (Loewus and Murthy, 2000). Alignment of MIPS amino acid sequences from diverse organisms including Arabidopsis, bean (Phaseolus vulgaris), Brewer's yeast (Saccharomyces cerevisiae), and Entamoeba histolytica revealed remarkable evolutionary conservation of the primary structure (Majumder et al., 1997). Genome sequencing projects have provided additional evidence for this striking conservation with deduced primary structures from organisms such as Caenorhabditis elegans, fruitfly (Drosophila melanogaster), Leishmania major, and Chlamydomonas reinhardtii.Although the essential...

Specific targeting is a key step to realize the full potential of iron oxide nanoparticles in biomedical applications, especially tumor-associated diagnosis and therapy. Here, we developed anti-GD2 antibody conjugated iron oxide nanoparticles for highly efficient neuroblastoma cell targeting. The antibody conjugation was achieved through an easy, linker-free method based on catechol reactions. The targeting efficiency and specificity of the antibody-conjugated nanoparticles to GD2-positive neuroblastoma cells were confirmed by flow cytometry, fluorescence microscopy, Prussian blue staining and transmission electron microscopy. These detailed studies indicated that the receptor-recognition capability of the antibody was fully retained after conjugation and the conjugated nanoparticles quickly attached to GD2-positive cells within four hours. Interestingly, longer treatment (12 h) led the cell membrane-bound nanoparticles to be internalized into cytosol, either by directly penetrating the cell membrane or escaping from the endosomes. Last but importantly, the uniquely designed functional surfaces of the nanoparticles allow easy conjugation of other bioactive molecules.

Immune cells play an important role in recognizing and removing foreign objects, such as nanoparticles. Among various parameters, surface coatings of nanoparticles are the first contact with biological system, which critically affect nanoparticle interactions. Here, surface coating effects on nanoparticle cellular uptake, toxicity and ability to trigger immune response were evaluated on a human monocyte cell line using iron oxide nanoparticles. The cells were treated with nanoparticles of three types of coatings (negatively charged polyacrylic acid, positively charged polyethylenimine and neutral polyethylene glycol). The cells were treated at various nanoparticle concentrations (5, 10, 20, 30, 50 μg ml À1 or 2, 4, 8, 12, 20 μg cm À2 ) with 6 h incubation or treated at a nanoparticle concentration of 50 μg ml À1 (20 μg cm À2 ) at different incubation times (6, 12, 24, 48 or 72 h). Cell viability over 80% was observed for all nanoparticle treatment experiments, regardless of surface coatings, nanoparticle concentrations and incubation times. The much lower cell viability for cells treated with free ligands (e.g.~10% for polyethylenimine) suggested that the surface coatings were tightly attached to the nanoparticle surfaces. The immune responses of cells to nanoparticles were evaluated by quantifying the expression of toll-like receptor 2 and tumor necrosis factor-α. The expression of tumor necrosis factor-α and toll-like receptor 2 were not significant in any case of the surface coatings, nanoparticle concentrations and incubation times. These results provide useful information to select nanoparticle surface coatings for biological and biomedical applications.

B-cell lymphoma-extra large (Bcl-xL) is an anti-apoptotic member of the Bcl2 family of proteins, which supports neurite outgrowth and neurotransmission by improving mitochondrial function. During excitotoxic stimulation, however, Bcl-xL undergoes post-translational cleavage to ∆N-Bcl-xL, and accumulation of ∆N-Bcl-xL causes mitochondrial dysfunction and neuronal death. In this study, we hypothesized that the generation of reactive oxygen species (ROS) during excitotoxicity leads to formation of ∆N-Bcl-xL. We further proposed that the application of an antioxidant with neuroprotective properties such as α-tocotrienol (TCT) will prevent ∆N-Bcl-xL-induced mitochondrial dysfunction via its antioxidant properties. Primary hippocampal neurons were treated with α-TCT, glutamate, or a combination of both. Glutamate challenge significantly increased cytosolic and mitochondrial ROS and ∆N-Bcl-xL levels. ∆N-Bcl-xL accumulation was accompanied by intracellular ATP depletion, loss of mitochondrial membrane potential, and cell death. α-TCT prevented loss of mitochondrial membrane potential in hippocampal neurons overexpressing ∆N-Bcl-xL, suggesting that ∆N-Bcl-xL caused the loss of mitochondrial function under excitotoxic conditions. Our data suggest that production of ROS is an important cause of ∆N-Bcl-xL formation and that preventing ROS production may be an effective strategy to prevent ∆N-Bcl-xL-mediated mitochondrial dysfunction and thus promote neuronal survival.