Copper oxide (CuO) nanoparticles (NPs) are abundant in manufacturing processes, but they are an airway irritant. In vitro pulmonary toxicity of CuO NPs has been modeled using cell lines such as human bronchial epithelial cell line BEAS‐2B. In 2D in vitro culture, BEAS‐2B undergoes squamous differentiation due to the presence of serum. Differentiation is part of the repair process of lung cells in vivo that helps to preserve the epithelial lining of the respiratory tract. Herein, the effects of serum on the hydrodynamic diameter, cellular viability, cellular differentiation, and cellular uptake of 5 and 35 nm CuO NPs are investigated, and the mean cell area is used as the differentiation marker for BEAS‐2B cells. The results demonstrate that the hydrodynamic diameter decreases with the addition of serum to the culture medium. Serum also increases the mean cell area, and only affects dose‐dependent cytotoxicity of 35 nm CuO NPs, while simultaneously having no effect on intracellular Cu2+. This study presents evidence that both NP size and the presence of serum in culture media influence the relative viability of BEAS‐2B cells following CuO NP exposure and highlights a critical need for carefully designed experiments and accurately reported conditions.

Sympathetic excitation contributes to clinical deterioration in systolic heart failure (HF). Significant inhibition of hypothalamic paraventricular nucleus (PVN) ERK1/2 signaling and a subsequent reduction of plasma norepinephrine (NE) levels in HF rats were achieved 2 weeks after a single subcutaneous injection of PD98059-loaded polymeric microparticles, without apparent adverse events, while blank microparticles had no effect. Similar reductions in plasma NE, a general indicator of sympathetic excitation, were previously achieved in HF rats by intracerebroventricular infusion of PD98059 or genetic knockdown of PVN ERK1/2 expression. This study presents a clinically feasible therapeutic approach to the central abnormalities contributing to HF progression.

There is a growing demand for biosensors that are reliable, accurate, rapid, and cost effective. Appropriately functionalized silicon nanowires (SiNWs) have offered an alternative to the gold standards of enzyme linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR)-based detection of proteins and DNA, respectively, by offering rapid label-free accurate detection of analytes. However, a major barrier limiting the progression of this technology is the cost of fabricating uniform functional nanowires (NWs) for biosensing and electrically contacting these NWs. Preliminary results are presented demonstrating the feasibility of a novel class of SiNW-based biosensor, which alleviates these issues. The SiNW-based biosensor presented here uses a SiNW-based solar cell (SiNW-SC) as a transducing element. It is demonstrated that SiNW-SCs can be modified by anchoring monoclonal antibodies specific for prostate-specific antigen (PSA) to the surface of SiNW-SCs. This surface modification allows for an increase in binding of PSA to the SiNW structures, which causes an increase in short-circuit density of SiNW-SCs when exposed to light and PSA. This paves the way for the development of a novel class of SiNW-based biosensors using surface modification of SiNW-SCs.

We tested whether inhibiting mechanically responsive articular chondrocyte mitochondria after severe traumatic injury and preventing oxidative damage represent a viable paradigm for posttraumatic osteoarthritis (PTOA) prevention. We used a porcine hock intra-articular fracture (IAF) model well suited to human-like surgical techniques and with excellent anatomic similarities to human ankles. After IAF, amobarbital or N -acetylcysteine (NAC) was injected to inhibit chondrocyte electron transport or downstream oxidative stress, respectively. Effects were confirmed via spectrophotometric enzyme assays or glutathione/glutathione disulfide assays and immunohistochemical measures of oxidative stress. Amobarbital or NAC delivered after IAF provided substantial protection against PTOA at 6 months, including maintenance of proteoglycan content, decreased histological disease scores, and normalized chondrocyte metabolic function. These data support the therapeutic potential of targeting chondrocyte metabolism after injury and suggest a strong role for mitochondria in mediating PTOA.

Asthma is a common lung disease affecting over 300 million people worldwide and is associated with increased reactive oxygen species, eosinophilic airway inflammation, bronchoconstriction, and mucus production. Targeting of novel therapeutic agents to the lungs of patients with asthma may improve efficacy of treatments and minimize side effects. We previously demonstrated that Ca2+/calmodulin-dependent protein kinase (CaMKII) is expressed and activated in the bronchial epithelium of asthmatic patients. CaMKII inhibition in murine models of allergic asthma reduces key disease phenotypes, providing the rationale for targeted CaMKII inhibition as a potential therapeutic approach for asthma. Herein we developed a novel cationic nanoparticle (NP)-based system for delivery of the potent and specific CaMKII inhibitor peptide, CaMKIIN, to airways.1 CaMKIIN-loaded NPs abrogated the severity of allergic asthma in a murine model. These findings provide the basis for development of innovative, site-specific drug delivery therapies, particularly for treatment of pulmonary diseases such as asthma.

The development of reproducible methods for the fabrication of nanoparticles in the 21st century is a major scientific achievement. Currently, there are many well-established methods for the production of nanoparticles of different shapes, sizes, and compositions. Along with these advancements in nanotechnology, nanoparticles have emerged as excellent tools for a diverse range of applications and have become a focus of research globally. With fundamental research being established, efforts are now being directed toward intelligently designed nanoparticles in which the properties of the nanomaterial are finely tuned depending on the application of interest. Usually, this involves the functionalization of the nanoparticle surface with a ligand. In this review, the impact of nanoparticle surface chemistry is discussed as it applies to biological systems.

An excess of calcium (Ca2+) influx into mitochondria during mitochondrial re-energization is one of the causes of myocardial cell death during ischemic/reperfusion injury. This overload of Ca2+ triggers the mitochondrial permeability transition pore (mPTP) opening which leads to programmed cell death. During the ischemic/reperfusion stage, the activated Ca2+/calmodulin-dependent protein kinase II (CaMKII) enzyme is responsible for Ca2+ influx. To reduce CaMKII-related cell death, sub-micron particles composed of poly(lactic-co-glycolic acid) (PLGA), loaded with a CaMKII inhibitor peptide were fabricated. The CaMKII inhibitor peptide-loaded (CIP) particles were coated with a mitochondria targeting moiety, triphenylphosphonium cation (TPP), which allowed the particles to accumulate and release the peptide inside mitochondria to inhibit CaMKII activity. The fluorescently labeled TPP-CIP was taken up by mitochondria and successfully reduced reactive oxygen species (ROS) caused by Isoprenaline (ISO) in a differentiated rat cardiomyocyte-like cell line. When cells were treated with TPP-CIP prior to ISO exposure, they maintained mitochondrial membrane potential. The TPP-CIP protected cells from ISO-induced ROS production and decreased mitochondrial membrane potential. Thus, TPP-CIP has the potential to be used in protection against ischemia/reperfusion injury.

Uterine serous carcinoma, one of the most aggressive types of endometrial cancer, is characterized by poor outcomes and mutations in the tumour suppressor p53. Our objective was to engender synthetic lethality to paclitaxel (PTX), the frontline treatment for endometrial cancer, in tumours with mutant p53 and enhance the therapeutic efficacy using polymeric nanoparticles (NPs). First, we identified the optimal NP formulation through comprehensive analyses of release profiles and cellular-uptake and cell viability studies. Not only were PTX-loaded NPs superior to PTX in solution, but the combination of PTX-loaded NPs with the antiangiogenic molecular inhibitor BIBF 1120 (BIBF) promoted synthetic lethality specifically in cells with the loss-of-function (LOF) p53 mutation. In a xenograft model of endometrial cancer, this combinatorial therapy resulted in a marked inhibition of tumour progression and extended survival. Together, our data provide compelling evidence for future studies of BIBF- and PTX-loaded NPs as a therapeutic opportunity for LOF p53 cancers.

Engineered nanomaterials (ENs) comprise a diverse suite of structures manufactured from a range of distinct components with an ever-increasing number of applications in industry, commerce, and medicine (Fig 1). ENs reported to be produced in the largest quantities include titanium dioxide, silicon dioxide, zinc oxide (ZnO), aluminum oxide, iron oxide, silver, carbon nanotubes, fullerenes, and quantum dots at levels ranging from 0.5 to 5500 tons/y worldwide.1Piccinno F. Gottschalk F. Seeger S. Nowack B. Industrial production quantities and uses of ten engineered nanomaterials in Europe and the world.J Nanopart Res. 2012; 14: 1109Crossref Scopus (980) Google Scholar The negative effect of ENs on the environment and human health because of off-target effects is of major concern and requires ongoing investigation. A recent database (NanoE-Tox) compiled from 224 articles published over the preceding decade was created to obtain an overview of the environmental effects of 8 chemically distinct ENs in which toxicity to crustaceans (Daphnia magna), fish, algae, or bacteria was the primary readout.2Juganson K. Ivask A. Blinova I. Mortimer M. Kahru A. NanoE-Tox: new and in-depth database concerning ecotoxicity of nanomaterials.Beilstein J Nanotechnol. 2015; 6: 1788-1804Crossref PubMed Scopus (103) Google Scholar The purposes of such databases are to (1) gain an understanding of the relative toxicities of various ENs and how these relate to their physicochemical properties, (2) decipher the mechanism or mechanisms by which their toxicities occur, and (3) provide an alternative to ethically questionable, time-consuming, and costly animal studies through the development of predictive models that derive from these databases. NanoE-Tox reported the most ecotoxic ENs to be made from silver, followed by ZnO and copper oxide (CuO), with the primary mechanism of toxicity of these ENs being dissolution of ions. However, direct cell membrane damage, oxidative stress, and genotoxicity were also potentially contributing factors for silver and ZnO ENs. A lack of sufficient information regarding the various physicochemical parameters (primary size, shape, surface charge, and surface area) meant that their contributions to cytotoxicity could not be conclusively defined. This lack of characterization of these and other physicochemical properties, such as aspect ratios, crystallinity, state of agglomeration, hydrophobicity, and solubility, are a definite drawback of many past studies with respect to ultimately understanding the relationship between such properties and cytotoxic potential. In the context of both the environment and human health, more expansive characterizations of ENs are required if predictive models, which might ultimately obviate the need for laborious wet laboratory work, are to become of significant value in terms of hazard screening. Another limitation of studies assessing the effect of ENs on the environment is the paucity of attention paid to the terrestrial versus aquatic environment.3McKee M.S. Filser J. Impacts of metal-based engineered nanomaterials on soil communities.Environ Sci: Nano. 2016; 3: 506-533Crossref Google Scholar A recently published detailed review covering the effect of metal-based ENs on soil communities describes examples of low concentrations of these ENs having a negative effect on invertebrates and microbial organisms, which in turn might have detrimental ecological consequences.3McKee M.S. Filser J. Impacts of metal-based engineered nanomaterials on soil communities.Environ Sci: Nano. 2016; 3: 506-533Crossref Google Scholar These authors also highlight the vast knowledge gap that exists concerning the effect of these ENs on soil communities and recommend studies not simply focused on individual species and short-term effects, especially considering that metal-based ENs are generally insoluble and gradually accumulating in soil over time. There is currently a dearth of empiric data published concerning the effect of ENs and the influence of their various physicochemical parameters on human health. Nevertheless, because the lungs are one of the primary routes of inadvertent EN entry, there has been an increasing number of studies using human lung cells in vitro to determine the potential of ENs to cause inflammation, genotoxicity, oxidative stress, or cytotoxicity. Although in vitro data often cannot substitute for results obtained in vivo, it has been shown to be feasible in specific circumstances. For example, an in vitro model assessing the proinflammatory potential of ultrafine particles on the bronchoalveolar adenocarcinoma–derived lung cell line A549 generated findings strongly correlative with in vivo results.4Monteiller C. Tran L. MacNee W. Faux S. Jones A. Miller B. et al.The pro-inflammatory effects of low-toxicity low-solubility particles, nanoparticles and fine particles, on epithelial cells in vitro: the role of surface area.Occup Environ Med. 2007; 64: 609-615Crossref PubMed Scopus (393) Google Scholar Additionally, in vitro studies assessing the cytotoxicity of CuO ENs (and ZnO ENs) on A549 cells were shown to correlate well with in vivo effects on lungs.5Cho W.S. Duffin R. Bradley M. Megson I.L. MacNee W. Lee J.K. et al.Predictive value of in vitro assays depends on the mechanism of toxicity of metal oxide nanoparticles.Part Fibre Toxicol. 2013; 10: 55Crossref PubMed Scopus (96) Google Scholar At this stage, the paucity of in vitro data assessing the effect of ENs on lung cells places limitations on the understanding of how ENs mediate their cytotoxic effects. Although a picture is developing of these mechanisms, it is predominantly derived from studies of ENs using one cell line, A549 (Fig 2), and therefore more studies using other lung cell lines, preferably in parallel, are required (Fig 3).Fig 3Histograms highlighting the paucity of in vitro studies that assess the effect of ENs on lung cell health and inflammatory potential. A, Number of published articles (2006-2016) documenting the potentially detrimental effects (in vitro) of copper oxide ENs, titanium dioxide ENs, single-walled carbon nanotubes (SWCNTs), or multiwalled carbon nanotubes (MWCNTs) on the human adenocarcinoma cell line A549 (representative of alveolar basal epithelia). B, Comparison of the number of published articles documenting the potentially detrimental effects of titanium dioxide ENs on the A549 cell line, the BEAS-2B cell line (a cell line representative of normal human bronchial epithelia), or both studied in parallel (indicated as both). The findings shown here for titanium dioxide ENs represent a general trend for ENs in which a greater number of articles have been published involving A549 compared with BEAS-2B cells. Data were generated by using the Thomson Reuters Web of Science citation indexing service and based on a search for articles in all databases using the following search terms: Nano* AND “copper oxide” or CuO for copper oxide ENs; Nano* AND “titanium dioxide” OR TiO2 for titanium dioxide ENs; “carbon nanotubes” NOT multi-walled for SWCNTs; and “carbon nanotubes” AND multi-walled OR multiwalled for MWCNTs (Fig 3, A). The search for each EN was then followed by a refined search with A549 cells, followed by another refined search for cytotox*, genotox*, inflamm* or “ROS.” After a search for titanium dioxide ENs (described above), a refined search was performed by using A549 or BEAS-2B or A549 followed by BEAS-2B (both) cells (Fig 3, B). Results from each search were modified further by scrutinizing each citation and eliminating any irrelevant publications, thereby increasing the accuracy of the resulting data.View Large Image Figure ViewerDownload Hi-res image Download (PPT) In lieu of waiting for exhaustive data on their effects on human health, certain ENs are surface modifiable, such that their effect on human health could be significantly reduced. For instance, we have recently demonstrated that surface modification of amorphous silica nanoparticles (approximately 50 nm in diameter) with amine groups reduced their ability to promote inflammatory responses in mice lungs.6Morris A.S. Adamcakova-Dodd A. Lehman S.E. Wongrakpanich A. Thorne P.S. Larsen S.C. et al.Amine modification of nonporous silica nanoparticles reduces inflammatory response following intratracheal instillation in murine lungs.Toxicol Lett. 2016; 241: 207-215Crossref PubMed Scopus (39) Google Scholar This modification made no significant change in the size of these ENs and is believed to reduce their oxidative potential by decreasing the amount of exposed surface silanol groups. It is becoming increasingly apparent that each EN formulation requires testing for potential harm to health as opposed to invoking generalized assumptions because the alteration of one physicochemical parameter within the formulation can have a significant effect on downstream biological processes. As an example, we have recently shown that CuO ENs (nanoparticles), which are physically and chemically similar in all respects apart from size (4 vs 24 nm), had significantly different effects on the viability of and reactive oxygen species production in lung cells.7Wongrapanich A. Mudunkotuwa I.A. Geary S.M. Morris A.S. Mapuskar K.A. Spitz D.R. et al.Size-dependent cytotoxicity of copper oxide nanoparticles in lung epithelial cells.Environ Sci: Nano. 2016; 3: 365-374Crossref PubMed Google Scholar The finding that 4-nm CuO nanoparticles were less toxic than 24-nm CuO nanoparticles coincided with lower intracellular levels of Cu for the former, suggesting that the smaller nanoparticles (on a per-weight basis) were less readily taken up by lung cells in vitro. These results are in contrast with expectations based on findings with ENs made from other metals, such as titanium dioxide (size range, 14-196 nm), gold (1.4 vs 15 nm; range, 10-50 vs 100-200 nm), and silver (range, 15-55 nm), in which smaller ENs correlated with higher cytotoxicity. This highlights that each EN requires intense analysis in regard to its potentially deleterious biological effects.8Schrand A.M. Rahman M.F. Hussain S.M. Schlager J.J. Smith D.A. Syed A.F. Metal-based nanoparticles and their toxicity assessment.Wiley Interdiscip Rev Nanomed Nanobiotechnol. 2010; 2: 544-568Crossref PubMed Scopus (532) Google Scholar Understanding the effect of ENs on lung tissue function and health would also pave the way for potentially useful nanomedicines, particularly those aimed at local delivery of drugs to patients with lung cancer. Currently, however, there is understandable reluctance in the medical community to use inhalation delivery of EN-based nanomedicines, particularly for less life-threatening lung pathologies, such as asthma, and this is primarily due to the knowledge gap with respect to the harmful effects of these potential nanomedicines on patients, as well as occupational health concerns. Recently, a promising EN-based vaccine was developed in our laboratory to prevent dust mite–induced asthma.9Joshi V.B. Adamcakova-Dodd A. Jing X. Wongrakpanich A. Gibson-Corley K.N. Thorne P.S. et al.Development of a poly (lactic-co-glycolic acid) particle vaccine to protect against house dust mite induced allergy.AAPS J. 2014; 16: 975-985Crossref PubMed Scopus (39) Google Scholar The vaccine, comprising the house dust mite antigen Der p 2 and the Toll-like receptor 9 agonist CpG oligodeoxynucleotide coloaded into nanoparticles (300 nm) made from the biodegradable, nontoxic, and US Food and Drug Administration–approved polymer poly(lactic-co-glycolic acid), was administered subcutaneously rather than through the lungs and was shown to be effective in a mouse model of dust mite–induced asthma, demonstrating protection against Der p 2–induced airway hyperresponsiveness and allergic airway inflammation. However, it is evident that certain lung pathologies, such as lung cancer, will benefit from direct delivery of ENs to the lungs through, for example, local delivery by means of inhalation.10Kuzmov A. Minko T. Nanotechnology approaches for inhalation treatment of lung diseases.J Control Release. 2015; 219: 500-518Crossref PubMed Scopus (206) Google Scholar In summary, with the inevitable role that nanomedicines are to play combined with the increased production of ENs for consumer and industrial applications, there is an onus on the scientific community to vigilantly interrogate, in a considered and multidisciplinary fashion, the potential hazards of ENs to both human health and the environment.

Amorphous silica nanoparticles (NPs) possess unique material properties that make them ideal for many different applications. However, the impact of these materials on human and environmental health needs to be established. We investigated nonporous silica NPs both bare and modified with amine functional groups (3-aminopropyltriethoxysilane (APTES)) in order to evaluate the effect of surface chemistry on biocompatibility. In vitro data showed there to be little to no cytotoxicity in a human lung cancer epithelial cell line (A549) for bare silica NPs and amine-functionalized NPs using doses based on both mass concentration (below 200μg/mL) and exposed total surface area (below 14m(2)/L). To assess lung inflammation, C57BL/6 mice were administered bare or amine-functionalized silica NPs via intra-tracheal instillation. Two doses (0.1 and 0.5mg NPs/mouse) were tested using the in vivo model. At the higher dose used, bare silica NPs elicited a significantly higher inflammatory response, as evidence by increased neutrophils and total protein in bronchoalveolar lavage (BAL) fluid compared to amine-functionalized NPs. From this study, we conclude that functionalization of nonporous silica NPs with APTES molecules reduces murine lung inflammation and improves the overall biocompatibility of the nanomaterial.

The increasing use of copper oxide (CuO) nanoparticles (NPs) in medicine and industry demands an understanding of their potential toxicities. In this study, we compared the in vitro cytotoxicity of CuO NPs of two distinct sizes (4 and 24 nm) using the A549 human lung cell line. Despite possessing similar surface and core oxide compositions, 24 nm CuO NPs were significantly more cytotoxic than 4 nm CuO NPs. The difference in size may have affected the rate of entry of NPs into the cell, potentially influencing the amount of intracellular dissolution of Cu2+ and causing a differential impact on cytotoxicity.

Evaluating toxicological responses of engineered nanomaterials such as silica nanoparticles is critical in assessing health risks and exposure limits. Biological assays can be used to evaluate cytotoxicity of individual materials, but specific nano-bio interactions-which govern its physiological response-cannot currently be predicted from materials characterization and physicochemical properties. Understanding the role of free radical generation from nanomaterial surfaces facilitates understanding of a potential toxicity mechanism and provides insight into how toxic effects can be assessed. Size-matched mesoporous and nonporous silica nanoparticles in aminopropyl-functionalized and native forms were investigated to analyze the effects of porosity and surface functionalization on the observed cytotoxicity. In vitro cell viability data in a murine macrophage cell line (RAW 264.7) provides a model for what might be observed in terms of cellular toxicity upon an environmental or industrial exposure to silica nanoparticles. Electron paramagnetic resonance spectroscopy was implemented to study free radical species generated from the surface of these nanomaterials and the signal intensity was correlated with cellular toxicity. In addition, in vitro assay of intracellular reactive oxygen species (ROS) matched well with both the EPR and cell viability data. Overall, spectroscopic and in vitro studies correlate well and implicate production of ROS from a surface-catalyzed reaction as a predictor of cellular toxicity. The data demonstrate that mesoporous materials are intrinsically less toxic than nonporous materials, and that surface functionalization can mitigate toxicity in nonporous materials by reducing free radical production. The broader implications are in terms of safety by design of nanomaterials, which can only be extracted by mechanistic studies such as the ones reported here.