Search

Abstract: Agglomeration dramatically affects many aspects of nanoparticle–cell interactions. Here we show that hydroxyapatite nanoparticles formed large agglomerates in biological medium resulting in extensive particle uptake and dose-dependent cytotoxicity in human macrophages. Particle citration and/or the addition of the dispersant Darvan 7 dramatically reduced mean agglomerate sizes, the amount of particle uptake and concomitantly cytotoxicity. More surprisingly, agglomeration governed the mode of particle uptake. Agglomerates were sequestered within an extensive, interconnected membrane labyrinth open to the extracellular space. In spite of not being truly intracellular, imaging studies suggest particle degradation occurred within this surface-connected compartment (SCC). Agglomerate dispersion prevented the SCC from forming, but did not completely inhibit nanoparticle uptake by other mechanisms. The results of this study could be relevant to understanding particle–cell interactions during developmental mineral deposition, in ectopic calcification in disease, and during application of hydroxyapatite nanoparticle vectors in biomedicine. [Copyright &y& Elsevier]

Abstract: Calcium phosphate and hydroxyapatite nanoparticles are extensively researched for medical applications, including bone implant materials, DNA and SiRNA delivery vectors and slow release vaccines. Elucidating the mechanisms by which cells internalize nanoparticles is fundamental for their long-term exploitation. In this study, we demonstrate that hydrophilic hydroxyapatite nanoparticles are sequestered within a specialized compartment called SCC (surface-connected compartment). This membrane-bound compartment is an elaborate labyrinth-like structure directly connected to the extracellular space. This continuity is demonstrated by in vivo 2-photon microscopy of ionic calcium using both cell-permeable and cell-impermeable dyes and by 3-D reconstructions from serial block-face SEM of fixed cells. Previously, this compartment was thought to be initiated specifically by exposure of macrophages to hydrophobic nanoparticles. However, we show that the SCC can be triggered by a much wider range of nanoparticles. Furthermore, we demonstrate its formation in A549 human lung epithelial cells, which are considerably less phagocytic than macrophages. EDX shows that extensive amounts of hydroxyapatite nanoparticles can be sequestered in this manner. We propose that SCC formation may be a means to remove large amounts of foreign material from the extracellular space, followed by slow degradation, may be to avoid excessive damage to surrounding cells or tissues. [Copyright &y& Elsevier]

Abstract: Multi-walled carbon nanotubes (MWNTs) have been proposed for use in many applications and concerns about their potential effect on human health have led to the interest in understanding the interactions between MWNTs and human cells. One important technique is the visualisation of the intracellular distribution of MWNTs. We exposed human macrophage cells to unpurified MWNTs and found that a decrease in cell viability was correlated with uptake of MWNTs due to mainly necrosis. Cells treated with purified MWNTs and the main contaminant Fe2O3 itself yielded toxicity only from the nanotubes and not from the Fe2O3. We used 3-D dark-field scanning transmission electron microscopy (DF-STEM) tomography of freeze-dried whole cells as well as confocal and scanning electron microscopy (SEM) to image the cellular uptake and distribution of unpurified MWNTs. We observed that unpurified MWNTs entered the cell both actively and passively frequently inserting through the plasma membrane into the cytoplasm and the nucleus. These suggest that MWNTs may cause incomplete phagocytosis or mechanically pierce through the plasma membrane and result in oxidative stress and cell death. [Copyright &y& Elsevier]

Abstract: Ferumoxtran-10 is an ultrasmall superparamagnetic iron oxide nanoparticle potentially useful as a contrast material in magnetic resonance imaging for the diagnosis of inflammatory and degenerative disorders associated with high macrophage activity. In clinical trials, it is currently applied to monitor the effect of atorvastatin therapy on macrophage activity in human carotid plaques. A recent study reported the inhibition of iron oxide nanoparticle uptake in macrophages by lovastatin, an effect which could compromise the suitability of Ferumoxtran-10 as an MRI contrast material in patients on statin therapy. Therefore, we examined the effect of atorvastatin on human monocyte–macrophage uptake of Ferumoxtran-10 in vitro using biochemical assays, magnetic resonance imaging and transmission electron microscopy. Our study showed that non-toxic concentrations of atorvastatin did not affect the amount of Ferumoxtran-10 taken up by HMMs. Furthermore, the intracellular distribution of iron oxide nanoparticles and the resulting MRI signal intensities remained unchanged by statin treatment. These results were obtained using atorvastatin concentrations probably vastly exceeding those reached in patient plasma in vivo. Atorvastatin therapy itself is therefore unlikely to affect Ferumoxtran-10 based macrophage detection by MRI, a prerequisite for the use of this contrast material to monitor lesion macrophage burden during lipid-lowering therapy. [Copyright &y& Elsevier]

Abstract: Ferumoxtran-10, a dextran-coated ultrasmall superparamagnetic iron oxide particle, has the potential to reveal macrophages in vivo using magnetic resonance imaging potentially acting as a marker of inflammatory status. Pending clinical trials, we examined the interactions of Ferumoxtran-10 with human monocyte-macrophages (HMMs) in vitro to assess its safety and lack of pro-inflammatory activity. After 72h, Ferumoxtran-10 was not toxic at 1mg/ml and may be only mildly toxic at 10mg/ml. Viability in cells with a high intracellular Ferumoxtran-10 load was not affected over 14 days. Ferumoxtran-10 did not interfere with baseline or stimulated cytokine (interleukin-12, interleukin-6, tumour necrosis factor-α or interleukin-1β) or superoxide anion production or with Fc-receptor-mediated phagocytosis. Similarly, Ferumoxtran-10 did not induce cytokine production and was not chemotactic. High-resolution electron microscopy and selected-area electron diffraction confirmed the core of Ferumoxtran-10 is composed of crystalline magnetite. Bright field transmission electron microscopy of thin sections demonstrated that Ferumoxtran-10 was retained in lysosomes of HMM for several days. Ferumoxtran-10 is not toxic to HMMs in vitro, does not activate them to produce pro-inflammatory cytokines or superoxide anions, is not chemotactic and does not interfere with Fc-receptor-mediated phagocytosis. Furthermore, extremely high intracellular Ferumoxtran-10 concentrations had only slight or no effects on these key activities. [Copyright &y& Elsevier]