Immunomodulation and T Helper TH1/TH2 Response Polarization by CeO2 and TiO2 Nanoparticles

Immunomodulation by nanoparticles, especially as related to the biochemical properties of these unique materials, has scarcely been explored. In an in vitro model of human immunity, we demonstrate two catalytic nanoparticles, TiO2 (oxidant) and CeO2 (antioxidant), have nearly opposite effects on human dendritic cells and T helper (T-H) cells. For example, whereas TiO2 nanoparticles potentiated DC maturation that led towards T(H)1-biased responses, treatment with antioxidant CeO2 nanoparticles induced APCs to secrete the anti-inflammatory cytokine, IL-10, and induce a T(H)2-dominated T cell profile. In subsequent studies, we demonstrate these results are likely explained by the disparate capacities of the nanoparticles to modulate ROS, since TiO2, but not CeO2 NPs, induced inflammatory responses through an ROS/inflammasome/IL-1 beta pathway. This novel capacity of metallic NPs to regulate innate and adaptive immunity in profoundly different directions via their ability to modulate dendritic cell function has strong implications for human health since unintentional exposure to these materials is common in modern societies

Immunomodulation and T Helper TH1/TH2 Response Polarization by CeO2 and TiO2 Nanoparticles

Immunomodulation by nanoparticles, especially as related to the biochemical properties of these unique materials, has scarcely been explored. In an in vitro model of human immunity, we demonstrate two catalytic nanoparticles, TiO2 (oxidant) and CeO2 (antioxidant), have nearly opposite effects on human dendritic cells and T helper (T-H) cells. For example, whereas TiO2 nanoparticles potentiated DC maturation that led towards T(H)1-biased responses, treatment with antioxidant CeO2 nanoparticles induced APCs to secrete the anti-inflammatory cytokine, IL-10, and induce a T(H)2-dominated T cell profile. In subsequent studies, we demonstrate these results are likely explained by the disparate capacities of the nanoparticles to modulate ROS, since TiO2, but not CeO2 NPs, induced inflammatory responses through an ROS/inflammasome/IL-1 beta pathway. This novel capacity of metallic NPs to regulate innate and adaptive immunity in profoundly different directions via their ability to modulate dendritic cell function has strong implications for human health since unintentional exposure to these materials is common in modern societies

Coupling sensitive \u3cem\u3ein vitro\u3c/em\u3e and in silico techniques to assess cross-reactive CD4\u3csup\u3e+\u3c/sup\u3e T cells against the swine-origin H1N1 influenza virus

The outbreak of the novel swine-origin H1N1 influenza in the spring of 2009 took epidemiologists, immunologists, and vaccinologists by surprise and galvanized a massive worldwide effort to produce millions of vaccine doses to protect against this single virus strain. Of particular concern was the apparent lack of pre-existing antibody capable of eliciting cross-protective immunity against this novel virus, which fueled fears this strain would trigger a particularly far-reaching and lethal pandemic. Given that disease caused by the swine-origin virus was far less severe than expected, we hypothesized cellular immunity to cross-conserved T cell epitopes might have played a significant role in protecting against the pandemic H1N1 in the absence of cross-reactive humoral immunity. In a published study, we used an immunoinformatics approach to predict a number of CD4+ T cell epitopes are conserved between the 2008–2009 seasonal H1N1 vaccine strain and pandemic H1N1 (A/California/04/2009) hemagglutinin proteins. Here, we provide results from biological studies using PBMCs from human donors not exposed to the pandemic virus to demonstrate that pre-existing CD4+ T cells can elicit cross-reactive effector responses against the pandemic H1N1 virus. As well, we show our computational tools were 80–90% accurate in predicting CD4+ T cell epitopes and their HLA-DRB1-dependent response profiles in donors that were chosen at random for HLA haplotype. Combined, these results confirm the power of coupling immunoinformatics to define broadly reactive CD4+ T cell epitopes with highly sensitive in vitro biological assays to verify these in silico predictions as a means to understand human cellular immunity, including cross-protective responses, and to define CD4+ T cell epitopes for potential vaccination efforts against future influenza viruses and other pathogens

Data from: Evaluation of the innate immunostimulatory potential of originator and non-originator copies of insulin glargine in an in vitro human immune model

Background: The manufacture of insulin analogs requires sophisticated production procedures which can lead to differences in the structure, purity, and/or other physiochemical properties of resultant products that can affect their biologic activity. Here, we sought to compare originator and non-originator copies of insulin glargine for innate immune activity and mechanisms leading to differences in these response profiles in an in vitro model of human immunity. Methods: An endothelial/dendritic cell-based innate immune model was used to study antigen-presenting cell activation, cytokine secretion, and insulin receptor signalling pathways induced by originator and non-originator insulin glargine products. Mechanistic studies included signalling pathway blockade with specific inhibitors, analysis of the products in a Toll-like receptor reporter cell line assay, and insulin removal from the products by immunopurification. Findings: All insulin glargine products elicited at least a minor innate immune response comparable to human insulin, but some lots of a non-originator copy product induced the elevated secretion of the cytokines, IL-8 and IL-6. In studies aimed at addressing the mechanisms leading to differential cytokine production by these products, we found (1) the inflammatory response was not mediated by bacterial contaminants, (2) the innate response was driven by the insulin receptor through the MAPK pathway, and (3) the removal of insulin significantly reduced their capacity to induce innate activity. No evidence of product aggregates was detected, though the presence of some high molecular weight proteins argues for the presence of insulin dimers or others contaminants in these products. Conclusion: The data presented here suggests some non-originator insulin glargine product lots drive heightened in vitro human innate activity and provides preliminary evidence that changes in their biochemical composition (dimers, impurities) might be responsible for their greater immunostimulatory potential

TiO₂ and CeO₂ NPs induce differential T cell responses.

<p>CD4⁺ T cells were labeled with the division-sensitive dye, CFSE, and cultured in the presence or absence of the indicated stimuli (NPs: 10 µM, PHA: 1 µg/mL, PMA: 50 ng/mL) for 5 days. Thereafter, the cells were harvested and examined for proliferating (CFSE-low) cells by flow cytometry. Histograms are representative plots from one of the five donors investigated, CFSE plotted on x-axis as a percent of maximum (y-axis).</p

Redox activities of nanomaterials modulate ROS production and NLRP3 inflammasome activation in DCs.

<p>(A) Human DCs were cultured in the absence or presence of the indicated doses of TiO₂ or CeO₂ NPs for 24 hr prior to being examined for their production of ROS. (B) DCs were cultured in the presence of cerium oxide at various concentrations for 8 hours and then H₂O₂, an inducer of ROS, was added for the remainder of the 24 hour incubation period. Oxidative stress was measured by DCF-DA staining of ROS. Six donors where examined in total. (C) DCs were stimulated for 24 hours with Alhydrogel (AlHy, 150 µg/ml) as a positive control for NLRP3 activation. Alternatively, TiO₂ NPs or CeO₂ NPs were delivered at 1 µM to the cultures for 24 hours prior to being measured for the presence of IL-1β in the presence or absence of NLRP3 inhibitor, glybenclamide (50 µM). Each data point is representative of an individual donor, n = 10. A paired t-test was performed: **p<0.005, ***p<0.0005 versus mock group.</p

CeO₂ NPs and TiO₂ NPs appear as soft agglomerates when diluted in X-VIVO 15 serum free media.

<p>High resolution transmission electron microscopy of (A) CeO₂ NPs indicates a composition of individual 3–5 nm nanocrystallites and (B) 7–10 nm TiO₂(anatase) NPs. The average size distribution of (C) CeO₂ and (D) TiO₂ NPs were measured using dynamic light scattering following a 24 hour incubation of the prepared NP solutions (each at 500 µM) in X-VIVO 15. Selected area electron diffraction patterns (SAEDP) of the CeO₂ (E) and TiO₂ NPs (F) were carried out using a high-resolution transmission electron microscope (HRTEM) equipped with a FEI Tecnai F30 having an energy-dispersive X-ray (EDX) analyzer. The SAED pattern of CeO₂ NPs, where A(111), B(200), C(220) and D(311) correspond to the different lattice planes of CeO₂ and confirms the crystalline structure of this material. Similarly, the SAED pattern of TiO₂ also confirms the crystalline nature of the material since the A(101), B(004), C(200) and D(211) rings correspond to the different lattice planes of the NPs. Surface oxidation state of CeO<b>₂</b> and TiO<b>₂</b> NPs were calculated from the XPS spectrum of Ce3d (G) and Ti 2p (H). (G) Deconvoluted peaks at 882.36 eV, 898.20 eV, 901.23 eV, 907.03 eV, and 916.64 eV are attributed to a Ce⁴⁺ oxidation state (light gray solid line) while 880.22 eV, 885.24 eV, 899.16 eV and 903.68 eV are the characteristic peaks of a Ce³⁺ oxidation state (dark gray solid line). Intensity of the peaks for Ce3+ and Ce4+ were estimated, and Ce³⁺/Ce⁴⁺ ratio on the surface of the nanoparticles were calculated and found to be 1.66. (H) In the case of TiO₂ NPs, the binding energies of Ti 2p_3/2 and Ti 2p_1/2 are at approximately 458.84eV and 464.62 eV, respectively. The difference of ∼5.8 eV in both peaks indicates a valence state of +4 for Ti on the surface of the NPs.</p

Immunological and biochemical effect of nanomaterials investigated.

<p>Immunological and biochemical effect of nanomaterials investigated.</p

Human DCs have the capacity to internalize CeO₂ and TiO₂ NPs.

<p>Cytokine-derived human DCs were pulsed for 24 hours with the listed dosing range of either NP. The DCs were harvested and washed several times before examination by inductively coupled plasma-mass spectroscopy (ICP-MS) for metal analysis and detection (ppb). Each sample was examined for the presence of both cerium (bottom) and titanium (top) as an assay detection control. Ten donors were analyzed in total. The paired t-test was used for statistical analyses. n = 10; **p<0.005, ***p<0.0005 versus mock group.</p