Drug induced liver injury - a 2023 update.

Drug-Induced Liver Injury (DILI) constitutes hepatic damage attributed to drug exposure. DILI may be categorized as hepatocellular, cholestatic or mixed and might also involve immune responses. When DILI occurs in dose-dependent manner, it is referred to as intrinsic, while if the injury occurs spontaneously, it is termed as idiosyncratic. This review predominately focused on idiosyncratic liver injury. The established molecular mechanisms for DILI include (1) mitochondria dysfunction, (2) increased reactive oxygen species levels, (3) presence of elevated apoptosis and necrosis, (4) and bile duct injuries associated with immune mediated pathways. However, it should be emphasized that the underlying mechanisms responsible for DILI are still unknown. Prevention strategies are critical as incidences occur frequently, and treatment options are limited once the injury has developed. The aim of this review was to utilize retrospective cohort studies from across the globe to gain insight into epidemiological patterns. This review considers (1) what is currently known regarding the mechanisms underlying DILI, (2) discusses potential risk factors and (3) implications of the coronavirus pandemic on DILI presentation and research. Future perspectives are also considered and discussed and include potential new biomarkers, causality assessment and reporting methods.

AIMS: An Automatic Semantic Machine Learning Microservice Framework to Support Biomedical and Bioengineering Research.

The fusion of machine learning and biomedical research offers novel ways to understand, diagnose, and treat various health conditions. However, the complexities of biomedical data, coupled with the intricate process of developing and deploying machine learning solutions, often pose significant challenges to researchers in these fields. Our pivotal achievement in this research is the introduction of the Automatic Semantic Machine Learning Microservice (AIMS) framework. AIMS addresses these challenges by automating various stages of the machine learning pipeline, with a particular emphasis on the ontology of machine learning services tailored to the biomedical domain. This ontology encompasses everything from task representation, service modeling, and knowledge acquisition to knowledge reasoning and the establishment of a self-supervised learning policy. Our framework has been crafted to prioritize model interpretability, integrate domain knowledge effortlessly, and handle biomedical data with efficiency. Additionally, AIMS boasts a distinctive feature: it leverages self-supervised knowledge learning through reinforcement learning techniques, paired with an ontology-based policy recording schema. This enables it to autonomously generate, fine-tune, and continually adapt to machine learning models, especially when faced with new tasks and data. Our work has two standout contributions demonstrating that machine learning processes in the biomedical domain can be automated, while integrating a rich domain knowledge base and providing a way for machines to have self-learning ability, ensuring they handle new tasks effectively. To showcase AIMS in action, we have highlighted its prowess in three case studies of biomedical tasks. These examples emphasize how our framework can simplify research routines, uplift the caliber of scientific exploration, and set the stage for notable advances.

Nanomaterial-induced toxicity in pathophysiological models representative of individuals with pre-existing medical conditions.

The integration of nanomaterials (NMs) into an ever-expanding number of daily used products has proven to be highly desirable in numerous industries and applications. Unfortunately, the same "nano" specific physicochemical properties, which make these materials attractive, may also contribute to hazards for individuals exposed to these materials. In 2021, it was estimated that 7 out of 10 deaths globally were accredited to chronic diseases, such as chronic liver disease, asthma, and cardiovascular-related illnesses. Crucially, it is also understood that a significant proportion of global populace numbering in the billions are currently living with a range of chronic undiagnosed health conditions. Due to the significant number of individuals affected, it is important that people suffering from chronic disease also be considered and incorporated in NM hazard assessment strategies. This review examined and analyzed the literature that focused on NM-induced adverse health effects in models which are representative of individuals exhibiting pre-existing medical conditions with focus on the pulmonary, cardiovascular, hepatic, gastrointestinal, and central nervous systems. The overall objective of this review was to outline available data, highlighting the important role of pre-existing disease in NM-induced toxicity with the aim of establishing a weight of evidence approach to inform the public on the potential hazards posed by NMs in both healthy and compromised persons in general population.

Assessing the transferability and reproducibility of 3D in vitro liver models from primary human multi-cellular microtissues to cell-line based HepG2 spheroids.

To reduce, replace, and refine in vivo testing, there is increasing emphasis on the development of more physiologically relevant in vitro test systems to improve the reliability of non-animal-based methods for hazard assessment. When developing new approach methodologies, it is important to standardize the protocols and demonstrate the methods can be reproduced by multiple laboratories. The aim of this study was to assess the transferability and reproducibility of two advanced in vitro liver models, the Primary Human multicellular microtissue liver model (PHH) and the 3D HepG2 Spheroid Model, for nanomaterial (NM) and chemical hazard assessment purposes. The PHH model inter-laboratory trial showed strong consistency across the testing sites. All laboratories evaluated cytokine release and cytotoxicity following exposure to titanium dioxide (TiO2) and zinc oxide (ZnO) nanoparticles. No significant difference was observed in cytotoxicity or IL-8 release for the test materials. The data were reproducible with all three laboratories with control readouts within a similar range. The PHH model ZnO induced the greatest cytotoxicity response at 50.0 µg/mL and a dose-dependent increase in IL-8 release. For the 3D HepG2 spheroid model, all test sites were able to construct the model and demonstrated good concordance in IL-8 cytokine release and genotoxicity data. This trial demonstrates the successful transfer of new approach methodologies across multiple laboratories, with good reproducibility for several hazard endpoints.

The application of existing genotoxicity methodologies for grouping of nanomaterials: towards an integrated approach to testing and assessment.

The incorporation of nanomaterials (NMs) in consumer products has proven to be highly valuable in many sectors. Unfortunately, however, the same nano specific physicochemical properties, which make these material attractive, might also contribute to hazards for people exposed to these materials. The physicochemical properties of NMs will impact their interaction with biological surroundings and influence their fate and their potential adverse effects such as genotoxicity. Due to the large and expanding number of NMs produced, their availability in different nanoforms (NFs) and their utilization in various formats, it is impossible for risk assessment to be conducted on an individual NF basis. Alternative methods, such as grouping are needed for streamlining hazard assessment. The GRACIOUS Framework provides a logical and science evidenced approach to group similar NFs, allowing read-across of hazard information from source NFs (or non-NFs) with adequate hazard data to target NFs that lack such data. Here, we propose a simple three-tiered testing strategy to gather evidence to determine whether different NFs are sufficiently similar with respect to their potential to induce genotoxicity, in order to be grouped. The tiered testing strategy includes simple in vitro models as well as a number of alternative more complex multi-cellular in vitro models to allow for a better understanding of secondary NM-induced DNA damage, something that has been more appropriate in vivo until recently.

Nanomaterial-Induced Extra-Pulmonary Health Effects - the Importance of Next Generation Physiologically Relevant In Vitro Test Systems for the Future of Nanotoxicology.

Manufactured nanomaterials (NMs) offer incredible scientific and societal benefits but their potential hazard to human health is not yet fully comprehended. In the last decade, a significant body of evidence indicates that certain NMs are capable of translocating from the primary exposure site (skin, lungs and gastrointestinal tract) to a number of secondary organs which includes the liver. Moreover, recent advances in the field of nanomedicine has resulted in increasing direct intravenous injection of NMs with the liver being a particularly important organ with regards to potential toxic effects and accumulation of said materials. It is generally acknowledged that it is not always possible to make direct or meaningful comparisons between in vitro and in vivo xenobiotic-induced toxicological responses. One of the main reasons for the lack of comparability between the testing strategies is that biological responses are not often alike which can in part be attributed to the numerous limitations of traditional mono-cellular in vitro test systems which are acting as a surrogate for a whole organ. In an attempt to address and highlight this important issue, this chapter will discuss the progress made in the production and validation oof next generation more physiologically relevant multi-cellular in vitro models of skin, GIT and the liver utilised for the assessment of the NM-induced toxicological effects.

Particulate and drug-induced toxicity assessed in novel quadruple cell human primary hepatic disease models of steatosis and pre-fibrotic NASH.

In an effort to replace, reduce and refine animal experimentation, there is an unmet need to advance current in vitro models that offer features with physiological relevance and enhanced predictivity of in vivo toxicological output. Hepatic toxicology is key following chemical, drug and nanomaterials (NMs) exposure, as the liver is vital in metabolic detoxification of chemicals as well as being a major site of xenobiotic accumulation (i.e., low solubility particulates). With the ever-increasing production of NMs, there is a necessity to evaluate the probability of consequential adverse effects, not only in health but also in clinically asymptomatic liver, as part of risk stratification strategies. In this study, two unique disease initiation and maintenance protocols were developed and utilised to mimic steatosis and pre-fibrotic NASH in scaffold-free 3D liver microtissues (MT) composed of primary human hepatocytes, hepatic stellate cells, Kupffer cells and sinusoidal endothelial cells. The characterized diseased MT were utilized for the toxicological assessment of a panel of xenobiotics. Highlights from the study included: 1. Clear experimental evidence for the pre-existing liver disease is important in the augmentation of xenobiotic-induced hepatotoxicity and 2. NMs are able to activate stellate cells. The data demonstrated that pre-existing disease is vital in the intensification of xenobiotic-induced liver damage. Therefore, it is imperative that all stages of the wide spectrum of liver disease are incorporated in risk assessment strategies. This is of significant consequence, as a substantial number of the general population suffer from sub-clinical liver injury without any apparent or diagnosed manifestations.

Acute hazard assessment of silver nanoparticles following intratracheal instillation, oral and intravenous injection exposures.

With ever-increasing production and use of nanoparticles (NPs), there is a necessity to evaluate the probability of consequential adverse effects in individuals exposed to these particles. It is now understood that a proportion of NPs can translocate from primary sites of exposure to a range of secondary organs, with the liver, kidneys and spleen being some of the most important. In this study, we carried out a comprehensive toxicological profiling (inflammation, changes in serum biochemistry, oxidative stress, acute phase response and histopathology) of Ag NP induced adverse effects in the three organs of interest following acute exposure of the materials at identical doses via intravenous (IV), intratracheal (IT) instillation and oral administration. The data clearly demonstrated that bioaccumulation and toxicity of the particles were most significant following the IV route of exposure, followed by IT. However, oral exposure to the NPs did not result in any changes that could be interpreted as toxicity in any of the organs of interest within the confines of this investigation. The finding of this study clearly indicates the importance of the route of exposure in secondary organ hazard assessment for NPs. Finally, we identify Connexin 32 (Cx32) as a novel biomarker of NP-mediated hepatic damage which is quantifiable both (in vitro) and in vivo following exposure of physiologically relevant doses.

A review of hepatic nanotoxicology - summation of recent findings and considerations for the next generation of study designs.

The liver is one of the most important multi-functional organs in the human body. Amongst various crucial functions, it is the main detoxification center and predominantly implicated in the clearance of xenobiotics potentially including particulates that reach this organ. It is now well established that a significant quantity of injected, ingested or inhaled nanomaterials (NMs) translocate from primary exposure sites and accumulate in liver. This review aimed to summarize and discuss the progress made in the field of hepatic nanotoxicology, and crucially highlight knowledge gaps that still exist.Key considerations include In vivo studies clearly demonstrate that low-solubility NMs predominantly accumulate in the liver macrophages the Kupffer cells (KC), rather than hepatocytes.KCs lining the liver sinusoids are the first cell type that comes in contact with NMs in vivo. Further, these macrophages govern overall inflammatory responses in a healthy liver. Therefore, interaction with of NM with KCs in vitro appears to be very important.Many acute in vivo studies demonstrated signs of toxicity induced by a variety of NMs. However, acute studies may not be that meaningful due to liver's unique and unparalleled ability to regenerate. In almost all investigations where a recovery period was included, the healthy liver was able to recover from NM challenge. This organ's ability to regenerate cannot be reproduced in vitro. However, recommendations and evidence is offered for the design of more physiologically relevant in vitro models.Models of hepatic disease enhance the NM-induced hepatotoxicity.The review offers a number of important suggestions for the future of hepatic nanotoxicology study design. This is of great significance as its findings are highly relevant due to the development of more advanced in vitro, and in silico models aiming to improve physiologically relevant toxicological testing strategies and bridging the gap between in vitro and in vivo experimentation.

Polylactic is a Sustainable, Low Absorption, Low Autofluorescence Alternative to Other Plastics for Microfluidic and Organ-on-Chip Applications.

Organ-on-chip (OOC) devices are miniaturized devices replacing animal models in drug discovery and toxicology studies. The majority of OOC devices are made from polydimethylsiloxane (PDMS), an elastomer widely used in microfluidic prototyping, but posing a number of challenges to experimentalists, including leaching of uncured oligomers and uncontrolled absorption of small compounds. Here we assess the suitability of polylactic acid (PLA) as a replacement material to PDMS for microfluidic cell culture and OOC applications. We changed the wettability of PLA substrates and demonstrated the functionalization method to be stable over a time period of at least 9 months. We successfully cultured human cells on PLA substrates and devices, without coating. We demonstrated that PLA does not absorb small molecules, is transparent (92% transparency), and has low autofluorescence. As a proof of concept of its manufacturability, biocompatibility, and transparency, we performed a cell tracking experiment of prostate cancer cells in a PLA device for advanced cell culture.

Assessment of nanomaterial-induced hepatotoxicity using a 3D human primary multi-cellular microtissue exposed repeatedly over 21 days - the suitability of the in vitro system as an in vivo surrogate.

BACKGROUND: With ever-increasing exposure to engineered nanomaterials (NMs), there is an urgent need to evaluate the probability of consequential adverse effects. The potential for NM translocation to distal organs is a realistic prospect, with the liver being one of the most important target organs. Traditional in vitro or ex vivo hepatic toxicology models are often limiting (i.e. short life-span, reduced metabolic activity, lacking important cell populations, etc.). In this study, we scrutinize a 3D human liver microtissue (MT) model (composed of primary hepatocytes and non-parenchymal cells). This unique experiment benefits from long-term (3 weeks) repeated very low exposure concentrations, as well as incorporation of recovery periods (up to 2 weeks), in an attempt to account for the liver's recovery capacity in vivo. As a means of assessing the toxicological potential of NMs, cell cytotoxicity (cell membrane integrity and aspartate aminotransferase (AST) activity), pro/anti-inflammatory response and hepatic function were investigated. RESULTS: The data showed that 2 weeks of cell culture might be close to limits before subtle ageing effects start to overshadow low sub-lethal NM-induced cellular responses in this test system (adenylate kinase (AK) cytotoxicity assay). We showed that in vitro AST measurement are not suitable in a nanotoxicological context. Moreover, the cytokine analysis (IL6, IL8, IL10 and TNF-α) proved useful in highlighting recovery periods as being sufficient for allowing a reduction in the pro-inflammatory response. Next, low soluble NM-treated MT showed a concentration-dependent penetration of materials deep into the tissue. CONCLUSION: In this study the advantages and pitfalls of the multi-cellular primary liver MT are discussed. Furthermore, we explore a number of important considerations for allowing more meaningful in vitro vs. in vivo comparisons in the field of hepatic nanotoxicology.

The variances in cytokine production profiles from non- or activated THP-1, Kupffer cell and human blood derived primary macrophages following exposure to either alcohol or a panel of engineered nanomaterials.

The portfolio of cytokines is key to the function of macrophages as sentries of the innate immune system as well as being critical for the transition from innate to adaptive immunity. Cytokine bias is critical in the fate of macrophages into a continuum of inflammatory to anti-inflammatory macrophages. Due to advances in the field of toxicology, increasingly advanced multi-cellular in vitro safety assessment models are being developed in order to allow for a better predication of potential adverse effects in humans with many of these models include a macrophage population. The selection of the correct macrophage cells in these advanced in vitro models is critical for a physiologically relevant and realistic immune response. In this study we investigated cytokine response profile (IL1-ß, IL6, IL10 and TNF-α) of activated and non-activated THP-1 (immortalized monocyte-like cell line), primary human Kupffer cells (liver resident macrophages) and human primary peripheral blood mononuclear cells following exposure of a panel of nanomaterials or ethanol. The data demonstrated that the THP-1 cell line are not great cytokine producers. The PBMC appear to be a good in vitro surrogate for circulating/pro-inflammatory macrophages but are not a suitable replacement for Kupffer cells. The findings from this study highlight the necessity for the selection of appropriate macrophages populations to meet the specific physiological requirements of in vitro experiment.

The mechanism-based toxicity screening of particles with use in the food and nutrition sector via the ToxTracker reporter system.

The rapid expansion of the incorporation of nano-sized materials in consumer products overlaps with the necessity for high-throughput reliable screening tools for the identification of the potential hazardous properties of the nanomaterials. The ToxTracker assay (mechanism-based reporter assay based on embryonic stem cells that uses GFP-tagged biomarkers for detection of DNA damage, oxidative stress and general cellular stress) is one such tool, which could prove useful in the field of particle toxicology allowing for high throughput screening. Here, ToxTracker was utilised to evaluate the potential hazardous properties of two particulates currently used in the food industry (vegetable carbon (E153) and food-grade TiO2 (E171)). Due to the fact that ToxTracker is based on a stem cell format, it is crucial that the data generated is assessed for its suitability and comparability to more conventionally used relevant source of cells - in this case cells from the gastrointestinal tract and the liver. Therefore, the cell reporter findings were compared to data from traditional assays (cytotoxicity, anti-oxidant depletion and DNA damage) and tissue relevant cell types. The data showed E171 to be the most cytotoxic, decreased intracellular glutathione and the most significant with regards to genotoxic effects. The ToxTracker data showed comparability to conventional toxicity and oxidative stress assays; however, some discrepancies were evident between the findings from ToxTracker and the comet assay.

The importance of inter-individual Kupffer cell variability in the governance of hepatic toxicity in a 3D primary human liver microtissue model.

The potential for nanomaterial (NM) translocation to secondary organs is a realistic prospect, with the liver one of the most important target organs. Traditional in vitro or ex vivo hepatic toxicology models are often limiting and/or troublesome (i.e. short life-span reduced metabolic activity, lacking important cell populations, high inter-individual variability, etc.). Building on previous work, this study utilises a 3D human liver microtissue (MT) model (MT composed of mono-culture of hepatocytes or two different co-culture MT systems with non-parenchymal cell (NPC) fraction sourced from different donors) to investigate the importance of inter-donor variability of the non-parenchymal cell population in the overall governance of toxicological response following exposure to a panel of NMs. To the best of our knowledge, this is the first study of its kind to investigate inter-donor variability in hepatic NPC population. The data showed that the Kupffer cells were crucial in dictating the overall hepatic toxicity following exposure to the materials. Furthermore, a statistically significant difference was noted between the two co-culture MT models. However, the trend for particle-induced biological responses was similar between the co-cultures (cytotoxicity, cytokine production and caspase activity). Therefore, despite the recognition of some discrepancies in the absolute values between the co-culture models, the fact that the trends and patterns of biological responses were comparable between the multi-cellular models we propose the 3D liver MT to be a valuable tool in particle toxicology.

Preparation and Utilization of a 3D Human Liver Microtissue Model for Nanotoxicological Assessment.

The liver is the principal detoxification center of the body, removing xenobiotics and waste products which could potentially include some nanomaterials (NM). With the ever increasing public and occupational exposure associated with accumulative production of nanomaterials, there is an urgent need to consider the possibility of harmful health consequences of engineered NM exposure. It is understood that following exposure via inhalation, ingestion, or direct intravenous injection a fraction of NMs reach the liver. Traditional in vitro or ex vivo hepatic nanotoxicology models are often limiting and/or troublesome (i.e., reduced metabolism enzymes, lacking important cell populations, unstable with very high variability, etc.). This chapter highlights a methodology for the preparation of a physiologically relevant 3D human liver microtissue model which addresses most of the negative issues associated with the models used in traditional in vitro hepatic toxicological investigations. The spheroids are a very promising model for the assessment of the toxicological effects associated with engineered NM exposure.

Nanodelivery systems and stabilized solid-drug nanoparticles for orally administered medicine: current landscape.

The use of nanoparticles as a means of targeted delivery of therapeutics and imaging agents could greatly enhance the transport of biologically active contents to specific target tissues, while avoiding or reducing potentially undesired side effects. Generally speaking, the oral route of administration is associated with good patient compliance, as it is convenient, economical, noninvasive, and does not require special training. Here, we review the progress of the utilization of nanodelivery-system carriers or stabilized solid-drug nanoparticles following oral administration, with particular attention on toxicological data. Mechanisms of cytotoxicity are discussed and the problem of extrapolating knowledge to human scenarios highlighted. Additionally, issues associated with administration of drugs via the oral route are underlined, while strategies utilized to overcome these are highlighted. This review aims to offer a balanced overview of strategies currently being used in the application of nanosize constructs for oral medical applications.

Substantial modification of the gene expression profile following exposure of macrophages to welding-related nanoparticles.

Anthropic nanoparticles (NP) are increasingly produced and emitted, with accompanying concerns for human health. Currently there is no global understanding as to the exact mechanistics of NP toxicity, as the traditional nanotoxicological approaches only provide a restricted overview. To address this issue, we performed an in-depth transcriptomic analysis of human macrophages exposed to a panel of welding-related metal oxide NP that we previously identified in welders lungs (Fe2O3, Fe3O4, MnFe2O4 and CrOOH NP). Utilizing the specified analysis criteria (|fold change| ≥1.5, p ≤ 0.001), a total of 2164 genes were identified to be differentially expressed after THP-1 macrophage exposure to the different NP. Performing Gene Ontology enrichment analysis, for cellular content, biological processes and Swiss-Prot/Protein Information Resource keywords the data show for the first time a profound modification of gene differential expression in response to the different NP, among which MnFe2O4 NP were the most potent to induce THP-1 macrophage activation. The transcriptomic analysis utilized in the study, provides novel insights into mechanisms that could contribute to NP-induced adverse effects and support the need for widened approaches to supplement existing knowledge of the processes underlying NP toxicity which would have not been possible using traditional nanotoxicological studies.

Assessment of polycyclic aromatic hydrocarbon exposure, lung function, systemic inflammation, and genotoxicity in peripheral blood mononuclear cells from firefighters before and after a work shift.

Firefighting is regarded as possibly carcinogenic, although there are few mechanistic studies on genotoxicity in humans. We investigated exposure to polycyclic aromatic hydrocarbons (PAH), lung function, systemic inflammation and genotoxicity in peripheral blood mononuclear cells (PBMC) of 22 professional firefighters before and after a 24-h work shift. Exposure was assessed by measurements of particulate matter (PM), PAH levels on skin, urinary 1-hydroxypyrene (1-OHP) and self-reported participation in fire extinguishing activities. PM measurements indicated that use of personal protective equipment (PPE) effectively prevented inhalation exposure, but exposure to PM occurred when the environment was perceived as safe and the self-contained breathing apparatuses were removed. The level of PAH on skin and urinary 1-OHP concentration were similar before and after the work shift, irrespective of self-reported participation in fire extinction activities. Post-shift, the subjects had reduced levels of oxidatively damaged DNA in PBMC, and increased plasma concentration of vascular cell adhesion molecule 1 (VCAM-1). The subjects reporting participation in fire extinction activities during the work shift had a slightly decreased lung function, increased plasma concentration of VCAM-1, and reduced levels of oxidatively damaged DNA in PBMC. Our results suggest that the firefighters were not exposed to PM while using PPE, but exposure occurred when PPE was not used. The work shift was not associated with increased levels of genotoxicity. Increased levels of VCAM-1 in plasma were observed. Environ. Mol. Mutagen. 59:539-548, 2018. © 2018 Wiley Periodicals, Inc.

Inhalation of House Dust and Ozone Alters Systemic Levels of Endothelial Progenitor Cells, Oxidative Stress, and Inflammation in Elderly Subjects.

Ambient air pollution including ozone and especially particulate matter represents important causes of cardiovascular disease. However, there is limited knowledge on indoor air dust with respect to this risk and the potential interactions between dust and ozone. Here, we exposed 23 healthy elderly subjects for 5.5 h, to either clean air, house dust at 275 µg/m3 (diameter < 2.5 µm), ozone at 100 ppb or combined house dust and ozone in a double-blinded randomized cross-over study. The combined house dust and ozone exposure was associated with a 48% (95% CI 24%-65%) decrease as compared with the clean air exposure, in CD34+KDR+ late endothelial progenitor cells (EPCs) per leukocyte in the blood shortly after exposure, whereas none of the single exposures resulted in a significant effect. The combined exposure also increased reactive oxygen species production capacity in granulocytes and monocytes as well as an up-regulation of interleukin-8 mRNA levels in leukocytes. Ozone alone reduced the gene expression of tumor necrosis factor and C-C motif chemokine ligand 2, while dust alone showed no effects. The combined exposure to house dust and ozone also reduced levels of oxidized purines in DNA consistent with concomitant up-regulation of mRNA of the repair enzyme 8-oxoguanine DNA glycosylase. The reduction in late EPCs can be an indicator of cardiovascular risk caused by the combination of pulmonary oxidative stress induced by ozone and the inflammatory potential of the house dust. These data were corroborated with in vitro findings from exposed human macrophages and endothelial cells.