Data on the histological and immune cell response in the popliteal lymph node in mice following exposure to metal particles and ions

AbstractHip implants containing cobalt–chromium (CoCr) have been used for over 80 years. In patients with metal-on-metal (MoM) hip implants, it has been suggested that wear debris particles may contribute to metal sensitization in some individuals, leading to adverse reactions. This article presents data from a study in which the popliteal lymph node assay (PLNA) was used to assess immune responses in mice treated with chromium-oxide (Cr2O3) particles, metal salts (CoCl2, CrCl3, and NiCl2) or Cr2O3 particles with metal salts (“A preliminary evaluation of immune stimulation following exposure to metal particles and ions using the mouse popliteal lymph node assay” (B.E. Tvermoes, K.M. Unice, B. Winans, M. Kovochich, E.S. Fung, W.V. Christian, E. Donovan, B.L. Finley, B.L. Kimber, I. Kimber, D.J. Paustenbach, 2016) [1]). Data are presented on (1) the chemical characterization of TiO2 particles (used as a particle control), (2) clinical observations in mice treated with Cr2O3 particles, metal salts or Cr2O3 particles with metal salts, (3) PLN weight and weight index (WI) in mice treated with Cr2O3 particles, metal salts or Cr2O3 particles with metal salts, (4) histological changes in PLNs of mice treated with Cr2O3 particles, metal salts or Cr2O3 particles with metal salts, (5) percentages of immune cells in the PLNs of mice treated with Cr2O3 particles, metal salts or Cr2O3 particles with metal salts, and (6) percentages of proliferating cells in the PLNs of mice treated with Cr2O3 particles, metal salts or Cr2O3 particles with metal salts

Activation of Latent HIV Using Drug-Loaded Nanoparticles

Antiretroviral therapy is currently only capable of controlling HIV replication rather than completely eradicating virus from patients. This is due in part to the establishment of a latent virus reservoir in resting CD4+ T cells, which persists even in the presence of HAART. It is thought that forced activation of latently infected cells could induce virus production, allowing targeting of the cell by the immune response. A variety of molecules are able to stimulate HIV from latency. However no tested purging strategy has proven capable of eliminating the infection completely or preventing viral rebound if therapy is stopped. Hence novel latency activation approaches are required. Nanoparticles can offer several advantages over more traditional drug delivery methods, including improved drug solubility, stability, and the ability to simultaneously target multiple different molecules to particular cell or tissue types. Here we describe the development of a novel lipid nanoparticle with the protein kinase C activator bryostatin-2 incorporated (LNP-Bry). These particles can target and activate primary human CD4+ T-cells and stimulate latent virus production from human T-cell lines in vitro and from latently infected cells in a humanized mouse model ex vivo. This activation was synergistically enhanced by the HDAC inhibitor sodium butyrate. Furthermore, LNP-Bry can also be loaded with the protease inhibitor nelfinavir (LNP-Bry-Nel), producing a particle capable of both activating latent virus and inhibiting viral spread. Taken together these data demonstrate the ability of nanotechnological approaches to provide improved methods for activating latent HIV and provide key proof-of-principle experiments showing how novel delivery systems may enhance future HIV therapy

Lipid nanoparticle (LNP) characterization and uptake in various cell types.

<p>(A) LNP were synthesized and characterized for their size by dynamic light scattering. (B) The membrane stain wheat germ agglutinin (WGA) visualized in blue was used with fluorescent microscopy in order to visualize the uptake of LNP-FITC (green) after a 16 hr incubation with HeLa cells. (C) WGA visualized in red was used to observe LNP-FITC (green) uptake in primary macrophages. (D) CEM cells were visualized by phase contrast images with LNP (green) using fluorescent microscopy. (E) LNP uptake in CEM cells was dose and energy dependent as detected by fold increase in FITC mean fluorescent intensity (MFI) using flow cytometry. (F) LNP uptake in CEM cells also increased over time as detected by flow cytometry.</p

Simultaneous incorporation of the protease inhibitor nelfinavir (Nel) and bryostatin-2 (Bry) into the lipid nanoparticles (LNP-Bry-Nel) can both activate latent virus expression and inhibit viral spread.

<p>(A) CEM cells were infected with HIV_NL4-3 and the cells were incubated for 3 days in the presence of various drug combinations including LNP-Bry-Nel. Viral p24 protein in the culture supernatant was measured by ELISA. (B) LNP-Bry-Nel was further tested for its ability to activate latent virus in J-Lat 10.6 cells as measured by induction of GFP expression. Error bars indicate the standard deviation of triplicate data points and are representative of at least 2 experiments. Media represents untreated infected cultures and LNP-con represents non-drug loaded nanoparticles.</p

Effects of LNP-Bry on latent virus expression and induction of CD69.

<p>(A) Representative flow cytometry plots of GFP expressing J-Lat 10.6 cells incubated with 10 nM bryostatin-2 or brostain-2 loaded LNP (LNP-Bry). (B) Percentage GFP+ cells was measured after incubation of LNP-Bry or bryostatin-2 with J-Lat 8.4 cells for 16 hr. (C) Percentage GFP+ cells was measured after incubation of LNP-Bry or bryostatin with J-Lat 10.6 cell lines for 16 hr. (D) Representative flow cytometry plots displaying the synergistic effect of 2 mM sodium butyrate (NaBut) in conjunction with LNP-Bry in J-Lat 8.4 cells. (E) J-Lat 8.4 cells were incubated with a fixed dose of NaBut (2 mM) and increasing dose of bryostatin-2 or LNP-Bry. (F) Representative flow cytometry plots displaying CD69 induction in primary resting CD4+ cells after incubation with LNP-Bry or bryostatin-2 for 16 hr. (G) Percentage CD69+ cells as detected by flow cytometric analysis. (H) Fold increase in CD69 mean fluorescence intensity was measured after incubation with increasing amounts of LNP-Bry or bryostatin-2 in primary CD4+ cells. (I) Flow cytometry plots displaying the CD4+/CD8+ cell profiles from mock or NL4-3 infected SCID-hu (Thy/Liv) implants. (J) In the presence of 1 µM raltegravir, CD4 single-positive thymocytes were stimulated <i>ex-vivo</i> with anti-CD3/CD28 beads, 10 nM bryostatin-2 or 10 nM LNP-Bry for 48 hr and assayed for intracellular Gag protein by flow cytometry. Error bars indicate the standard deviation of triplicate data points and are representative of at least 2 experiments. * p<0.01 as compared with media only (infected non-stimulated cultures) or LNP-con (non-drug loaded nanoparticles) in a paired t-test.</p

Review and Evaluation of the Potential Health Effects of Oxidic Nickel Nanoparticles

The exceptional physical and chemical properties of nickel nanomaterials have been exploited in a range of applications such as electrical conductors, batteries, and biomaterials. However, it has been suggested that these unique properties may allow for increased bioavailability, bio-reactivity, and potential adverse health effects. Thus, the purpose of this review was to critically evaluate data regarding the toxicity of oxidic nickel nanoparticles (nickel oxide (NiO) and nickel hydroxide (Ni(OH)2) nanoparticles) with respect to: (1) physico-chemistry properties; (2) nanomaterial characterization in the defined delivery media; (3) appropriateness of model system and translation to potential human effects; (4) biodistribution, retention, and clearance; (5) routes and relevance of exposure; and (6) current research data gaps and likely directions of future research. Inhalation studies were prioritized for review as this represents a potential exposure route in humans. Oxidic nickel particle size ranged from 5 to 100 nm in the 60 studies that were identified. Inflammatory responses induced by exposure of oxidic nickel nanoparticles via inhalation in rodent studies was characterized as acute in nature and only displayed chronic effects after relatively large (high concentration and long duration) exposures. Furthermore, there is no evidence, thus far, to suggest that the effects induced by oxidic nickel nanoparticles are related to preneoplastic events. There are some data to suggest that nano- and micron-sized NiO particles follow a similar dose response when normalized to surface area. However, future experiments need to be conducted to better characterize the exposure–dose–response relationship according to specific surface area and reactivity as a dose metric, which drives particle dissolution and potential biological responses

Targeting LNP to primary CD4+ cells and activation of latent virus expression.

<p>(A) Schematic representation showing construction of the LNP-CD4 (anti-CD4 coated nanoparticles) and LNP-Iso (isotype control antibody coated nanoparticles) with bryostatin-2 incorporation. (B) Fluorescent microscopy image displaying the targeting of LNP-CD4 (green) to CD4+ cells (red) while minimizing cellular association with CD8+ cells (violet) in the same culture of total peripheral blood mononuclear cells (PBMCs) after a 30 min exposure. (C) Fluorescent microscopy image displaying the lack of targeting by LNP-Iso (green) to CD4+ cells (red) or CD8+ cells (violet). (D) Quantification of LNP-CD4 uptake in PBMCs using flow cytometry (% cells CD4+ with increase in FITC-LNP uptake) after a 30 min exposure. (E) Representative flow cytometry plots of CD4+ and CD8+ cells analyzed for the induction of the early activation marker CD69 after incubation with CD4 targeting nanoparticles for 16 hr. (F) Bar graphs illustrating the fold increase in CD69 MFI in CD4+ and CD8+ cells after incubation with LNP-CD4 Bry. (G) LNP-CD4 Bry was further tested for its ability to stimulate latent virus expression in J-Lat 8.4 cells. (H) <i>Ex vivo</i> CD4 single-positive thymocytes isolated from infected SCID-hu (Thy/Liv) implants were incubated with LNP-CD4 Bry and viral p24 protein present in the culture supernatants was analyzed after 48 hr. Error bars indicate the standard deviation of triplicate data points and are representative of at least 2 experiments. * p<0.01 in a paired t-test as indicted in (F) and as compared with media only or LNP-con in (G) and (H).</p

Comparative Toxicity of C60 Aggregates toward Mammalian Cells: Role of Tetrahydrofuran (THF) Decomposition

International audienceC 60 fullerene is a promising material because of its unique physiochemical properties. However, previous studies have reported that colloidal aggregates of C 60 (nC 60) produce toxicity in fish and human cell cultures. The preparation method of nC 60 raises questions as to whether the observed effects stem from fullerenes or from the organic solvents used during the preparation of the suspensions. In this paper, we set out to elucidate the mechanism by which tetrahydrofuran (THF) treatment to enhance the preparation of nC 60 leads to cytotoxicity in a mouse macrophage cell line. Our results demonstrate that THF/nC 60 but not fullerol or aqueous nC 60 generates cellular toxicity through a pathway that involves increased intracellular flux and mitochondrial perturbation in RAW 264.7 cells. Interestingly, the supernatant of the THF/nC 60 suspension rather than the colloidal fullerene aggregates mimics the cytotoxic effects due to the presence of γ-butyrolactone and formic acid. Thus, the role of nC 60 in the cellular responses is likely not due to the direct effect of the nC 60 material surface on the cells but is related to the conversion of THF into a toxic byproduct during preparation of the suspension