Exposure to fine airborne particulate matter (PM2.5) is associated with cardiovascular disease and increased atherogenesis. While some prior studies provided mechanistic insights linking exposure to lesion progression, instability, and rupture, a complete definition of cellular events contributing to early vascular inflammation is lacking. Here we show that C57BL/6J mice exposed to concentrated ambient PM2.5 (CAP), as opposed to control mice inhaling filtered air, displayed endothelial activation, as evidenced by the increased number of leukocytes rolling on, and adhering to, the intact vasculature in vivo. As CAP exposure also induced the senescence of peripheral blood mononuclear cells (MNCs) and endothelial progenitor cells (EPCs), we further assessed the role of this outcome in PM2.5-induced vascular dysfunction. We observed that treatment with the senolytics Dasatinib and Quercetin (DQ), which eliminate senescent cells, was effective in reversing the CAP-induced senescence of both of MNCs and EPCs as demonstrated by reductions in β-galactosidase activity and the expression of genes characteristic of the senescence activated secretory phenotype (SASP). Furthermore, we found that DQ treatment was effective in reversing CAP-induced defects in in vitro EPC function (tube forming capacity) and limited in vivo endothelial activation as well. These results provide strong evidence for a role of PM2.5 in the early stages of atherogenesis by inducing endothelial activation leading to leukocyte recruitment. Our results further suggest that senolytic treatment may be efficacious in combatting adverse cardiovascular outcomes in those chronically exposed to high levels of PM2.5.

Ambient particulate matter (PM2.5) exposure is a major environmental risk factor for cardiopulmonary disease, but its effects on erythroid homeostasis remain incompletely understood. Although prior in vitro work indicates that PM2.5 can damage circulating red blood cells (RBCs), whether exposure alters erythropoiesis and the coordinated clearance of senescent RBCs has not been fully explored. Using a mouse model of whole-body exposure to concentrated ambient PM2.5 (CAP), we investigated associations between inhaled PM on erythroid output, splenic macrophage function, and lipid mediator signaling. CAP exposure was associated with suppressed erythropoietin levels, reduced circulating reticulocytes, and decreased erythroid precursor populations in the bone marrow, consistent with impaired erythropoiesis. Despite preserved splenic architecture, CAP-exposed mice exhibited reduced splenic iron and heme content, consistent with diminished erythrocyte turnover and processing. Targeted lipidomic profiling revealed broad suppression of proresolving lipid mediators in the spleen, with lipoxin A4 (LXA4) among the most consistently reduced species. Expression of the LXA4 receptor, ALX/FPR2, was also downregulated with prolonged exposure. Importantly, removal of CAP and return to filtered air resulted in normalization of splenic lipid mediator profiles, restoration of LXA4 levels, and recovery of erythroid parameters, including reticulocyte abundance and RBC stress markers. Together, these findings suggest that altered resolution signaling contributes to PM2.5-induced disruption of erythroid homeostasis and implicate macrophage-lipid mediator pathways in the hematologic response to environmental stress.

Background:Benzene is a ubiquitous environmental pollutant generated by a variety of natural and anthropological sources. It is a known carcinogen and hematopoietic toxin; however, little is known about benzene's potential atherogenicity. Hypothesis:Inhaled benzene induces atherogenesis by increasing vascular inflammation in LDL receptor Knockout (LDLR-KO) mice. Methods:Male LDLR-KO mice were exposed to HEPA-filtered air or benzene (1 ppm, 6h/day, 5days/week) for 24 weeks. For the last 12 weeks of exposure, the mice were maintained on a western diet. The single nuclei RNA sequencing (snRNAseq) of aortae was performed at Novogene. For in vitro experiments, splenic naïve T cells were exposed to 1 μM of hydroquinone (HQ) for 24 hours, and intracellular ROR-gamma levels were measured by flow cytometry. Results:Benzene inhalation increased the aortic valve lesion area by more than 25% (P<0.05) in LDLR-KO mice. Using snRNAseq, eleven major cell types were detected, including T cells and vascular smooth muscle cells (VSMC). Benzene increased the number of T cells by 2.5-fold, proliferating T-cells by 5.8-fold, and VSMC by 1.6-fold, suggesting increased cellularity and reduced plaque stability. In addition, benzene upregulated Th17 polarization marker Rorc and negative regulators of apoptosis Rag1 and Bcl11b while significantly attenuating the expression of proliferation inhibitor Ms4a4b in T cells. In VSMC, benzene downregulated extracellular matrix organization genes and upregulated platelet degranulation pathways. Polarization of T cells into Th17 was confirmed by HQ-dependent upregulation of ROR-gamma in vitro. Conclusion:Our data suggest that inhaled benzene exposure compromises plaque cellularity and stability by facilitating T-cell proliferation and polarization, which coincides with the degradation of smooth muscle extracellular matrix and platelet activation.

The emergence of synthetic nicotine analogs in "tobacco-free" products, such as 6-methylnicotine (6MN; aka Metatine) in SPREE BAR, presents new regulatory and public health challenges. Alarmingly, little is known about the metabolism of 6MN, its potential biomarkers of exposure, or its toxicity. In this study, we systematically characterized oxidized metabolites of 6MN in the urine of mice exposed to 6MN (via intraperitoneal or inhalation route) using liquid chromatography-high resolution mass spectrometry. Similarly, human urine samples were analyzed for 6MN metabolites after use of the SPREE BAR (Blue Razz Ice) product. Nine 6MN metabolites were identified in mouse urine, and each metabolite corresponded with a known nicotine metabolite, albeit with increased mass (i.e. m/z + 14 Da). Although 6MN and nicotine share oxidative routes, the metabolism of 6MN was dominated via N-oxidation (likely FMO3-mediated) rather than C-oxidation (likely CYP2A6-dependent) pathways, whereas nicotine metabolism is vice versa. Six 6MN metabolites were detected in human urine after SPREE BAR use, demonstrating strong cross-species metabolic concordance. Among these 6MN human metabolites, 6-methylcotinine, 6-methyl-3'-hydroxycotinine, and 6-methylcotinine-N-oxide emerged as potential urinary biomarkers of exposure due to their prevalence. Importantly, 6MN, yet not an equimolar dose of nicotine, induced acute neurotoxic effects in mice, highlighting distinct toxicological risks of 6MN compared with nicotine. This research revealed a distinct metabolic profile of 6MN and established a framework for biomonitoring of 6MN exposure. Together, these findings advanced our understanding of the metabolism of synthetic nicotine analogs and emphasized the importance of compound-specific profiling to support regulatory oversight of emerging nicotine-like products.

Exposure to green spaces has been linked to numerous health benefits, including a reduction in the risk of cardiovascular disease and lower mortality rates. However, to better understand how green spaces influence human health, it is essential to have valid, quantitative measures of greenness exposure. Building on our previous identification of urinary limonene metabolites as potential biomarkers of exposure, we now investigate α-pinene, another abundant plant-emitted monoterpene, to identify and quantify its urinary metabolites and evaluate their suitability as biomarkers. We used liquid chromatography-high resolution mass spectrometry (LC-HRMS) to analyze samples of human urine collected following either controlled α-pinene inhalation or real-world greenness exposure. Through a combination of pseudo-targeted and untargeted analyses, we discovered 22 α-pinene metabolites post-inhalation including nine novel structures, with two confirmed against synthetic standards. Relative quantitation of the urinary levels of the metabolites was used to estimate their kinetic parameters. A 4 h exposure to a forest environment resulted in significant increases in the most abundant metabolites. This suggests that α-pinene metabolites, specifically myrtenic acid glucuronide and dihydromyrtenic acid glucuronide, may serve as valid biomarkers when assessing individual exposure to green environments. When combined with other subjective and objective measures, these novel urinary biomarkers promote a more comprehensive assessment of exposure to greenness.

The cardiovascular and pulmonary disease risks of the use of electronic nicotine delivery systems (ENDS) are uncertain. We recently showed that ENDS solvent-derived aerosol (propylene glycol and vegetable glycerin, PG:VG) exposure induced a transient receptor potential ankyrin-1 (TRPA1)-dependent endothelial dysfunction (ED) in healthy female mice. As thermal degradation of PG:VG generates aldehydes, we hypothesized that acrolein (AC), a constituent of ENDS-derived aerosol and a known TRPA1 agonist, was responsible, in part, for the observed TRPA1-dependent pulmonary and vascular effects of PG:VG. To test this, female wild-type (WT) and TRPA1 null mice were exposed by inhalation to either filtered air or AC alone, and biomarkers of exposure and of harm were measured. Compared with their genotype-matched air control group, JUUL Virginia Tobacco (VT), PG:VG, and AC alone exposures (6 h) significantly increased urinary levels of the AC metabolite, 3-hydroxypropyl mercapturic acid (3HPMA), in both female WT and TRPA1 null mice. AC exposures at 1 and 3 ppm led to the rapid onset and reversal (upon cessation) of ‘respiratory braking’ in female WT but not in TRPA1 null mice indicating a TRPA1 dependence. As AC stimulated TRPA1-dependent respiratory braking, we measured urinary monoamines and their metabolites after exposure as a proxy of nervous system activation. In WT mice, AC exposure suppressed levels of dopamine, metanephrine, serotonin (5HT), and 5HT metabolite (5HIAA), whereas in TRPA1 null mice only 5HT was equally suppressed by AC. To assess vascular effects, mice were exposed for 4 days to Air or AC (6 h/day, 1 ppm), and aortic function was measured ex vivo. Although endothelial-dependent relaxation was similar in air control and AC-exposed mice, aortic sensitivity to an NO donor was enhanced significantly and equally by AC in both WT and TRPA1 null mice reflective of a TRPA1-independent and compensatory effect. Collectively, AC exposure at a level present in ENDS aerosols stimulated both TRPA1-dependent and -independent pulmonary, vascular, and systemic effects. These data suggest that ENDS use may increase cardiovascular and pulmonary disease risk, in part, via AC present in ENDS-derived aerosols yet independent of either nicotine or flavorants. The level of AC present in ENDS aerosols should be lowered to an amount where it does not induce biomarkers of vascular, pulmonary, and systemic harm to mitigate potential long-term disease risk. Vascular dysfunction in mid-thoracic aorta ex vivo following short-term exposure to acrolein (AC, 1 ppm, 6 h/day × 4 days) in female WT mice was similarly exposed female TRPA1 null mice indicating a TRPA1-independent effect. TRPA1-dependent nervous system-mediated respiratory braking response in wild-type mice (WT; represented by lightning bolt) was absent in female TRPA1 null mice exposed acutely to AC. Short-term AC exposure induced a ‘compensatory relaxation’ in thoracic aorta of both WT and TRPA1 null mice indicating a TRPA1-independent effect. Irritant compounds such as AC in ENDS-derived aerosols simultaneously promote both ED and ‘vascular compensation’ through TRPA1-dependent and -independent mechanisms that are perhaps, in part, dependent on changes in circulating monoamines. These findings suggest exposures to ENDS-derived aerosols have complex effects on the vasculature and thus, long-term ENDS use likely increases CVD risk. Created in BioRender.

Although some prior studies have identified an association between exposure to fine air-borne particulate matter (PM2.5) and indices of aging, the extent of these associations and their underlying mechanisms are uncertain. In this study, we exposed male C57BL/6J mice to filtered air and concentrated ambient PM2.5 (CAP) and assessed 2 common hallmarks of aging, telomere shortening and a senescent phenotype. Of the cell types examined, peripheral blood mononuclear cells (PBMNCs), endothelial progenitor cells (EPCs), and bone marrow-derived c-kit+ cells, all 3 demonstrated shortened telomeres when isolated from CAP-exposed mice as compared with cells derived from filtered air controls. We found that telomere attrition in PBMNCs and EPCs was mitigated in those CAP-exposed mice receiving water supplemented with the antioxidant, carnosine, and was reversible in PBMNCs, but not EPCs, when CAP-exposed mice were allowed to recover in normal air conditions. Telomere attrition in these cell types appeared to result from the attenuated catalytic activity of telomerase reverse transcriptase (Tert). PBMNCs and EPCs obtained from CAP-exposed mice also displayed increased β-galactosidase activity and expression of genes characteristic of the senescence-activated secretory phenotype. Of PBMNC subtypes, the increase of β-galactosidase activity was greatest in CD8+ T-cells. Our results suggest that the pro-aging effects of PM2.5 impact multiple cell types, including bone marrow stem cells, and that telomere attrition resulted from attenuated Tert activity. The aging and senescence of multiple cell types, including bone marrow stem cells, may underlie the diverse pathological outcomes of PM2.5 exposure.

Microplastics (MP) derived from the weathering of polymers, or synthesized in this size range, have become widespread environmental contaminants and have found their way into water supplies and the food chain. Despite this awareness, little is known about the health consequences of MP ingestion. We have previously shown that the consumption of polystyrene (PS) beads was associated with intestinal dysbiosis and diabetes and obesity in mice. To further evaluate the systemic metabolic effects of PS on the gut-liver-adipose tissue axis, we supplied C57BL/6J mice with normal water or that containing 2 sizes of PS beads (0.5 and 5 µm) at a concentration of 1 µg/ml. After 13 weeks, we evaluated indices of metabolism and liver function. As observed previously, mice drinking the PS-containing water had a potentiated weight gain and adipose expansion. Here we found that this was associated with an increased abundance of adipose F4/80+ macrophages. These exposures did not cause nonalcoholic fatty liver disease but were associated with decreased liver:body weight ratios and an enrichment in hepatic farnesoid X receptor and liver X receptor signaling. PS also increased hepatic cholesterol and altered both hepatic and cecal bile acids. Mice consuming PS beads and treated with the berry anthocyanin, delphinidin, demonstrated an attenuated weight gain compared with those mice receiving a control intervention and also exhibited a downregulation of cyclic adenosine monophosphate (cAMP) and peroxisome proliferator-activated receptor (PPAR) signaling pathways. This study highlights the obesogenic role of PS in perturbing the gut-liver-adipose axis and altering nuclear receptor signaling and intermediary metabolism. Dietary interventions may limit the adverse metabolic effects of PS consumption.

Cigarette smoking is positively and robustly associated with cardiovascular disease (CVD), including hypertension, atherosclerosis, cardiac arrhythmias, stroke, thromboembolism, myocardial infarctions, and heart failure. However, after more than a decade of ENDS presence in the U.S. marketplace, uncertainty persists regarding the long-term health consequences of ENDS use for CVD. New approach methods (NAMs) in the field of toxicology are being developed to enhance rapid prediction of human health hazards. Recent technical advances can now consider impact of biological factors such as sex and race/ethnicity, permitting application of NAMs findings to health equity and environmental justice issues. This has been the case for hazard assessments of drugs and environmental chemicals in areas such as cardiovascular, respiratory, and developmental toxicity. Despite these advances, a shortage of widely accepted methodologies to predict the impact of ENDS use on human health slows the application of regulatory oversight and the protection of public health. Minimizing the time between the emergence of risk (e.g., ENDS use) and the administration of well-founded regulatory policy requires thoughtful consideration of the currently available sources of data, their applicability to the prediction of health outcomes, and whether these available data streams are enough to support an actionable decision. This challenge forms the basis of this white paper on how best to reveal potential toxicities of ENDS use in the human cardiovascular system—a primary target of conventional tobacco smoking. We identify current approaches used to evaluate the impacts of tobacco on cardiovascular health, in particular emerging techniques that replace, reduce, and refine slower and more costly animal models with NAMs platforms that can be applied to tobacco regulatory science. The limitations of these emerging platforms are addressed, and systems biology approaches to close the knowledge gap between traditional models and NAMs are proposed. It is hoped that these suggestions and their adoption within the greater scientific community will result in fresh data streams that will support and enhance the scientific evaluation and subsequent decision-making of tobacco regulatory agencies worldwide. Models for Cardiovascular Toxicity Testing E-cigarettes and nicotine delivery systems can be examined using multiple model systems. In silico models might predict adverse cardiovascular effects that can be screened for using 2-D and 3-D models using induced pluripotent stem cell (iPSC) derived cardiac tissue and “omics” profiling such as single-cell RNA sequencing (scRNA-seq) or high-throughput functional analysis with a multiple electrode array (MEA). Biologic plausibility of detected effects can be corroborated using ex vivo or in vivo models, which may also lead to the discovery of new biomarkers or treatments for CVD.

Background: Benzene is a ubiquitous environmental pollutant abundant in automobile exhaust, cigarette smoke, forest fires, and present in several household products. It is ranked # 6 on Agency for Toxic Substances and Disease Registry (ATSDR) priority list, however, little is known about its effect on cardiovascular disease, especially atherosclerosis. Hypothesis: Inhaled benzene induces atherogenesis by increasing vascular inflammation in LDL receptor Knockout (LDLR-KO) mice. Methods: Male LDLR-KO or apoE-KO mice were exposed to HEPA-filtered air or benzene (1.0 or 10.0 ppm, 6h/day, 5 days/week) for 24 weeks and maintained on Western diet for the last 12 weeks of exposure. Atherosclerotic lesion composition and nature were determined by liquid chromatography-mass spectrometry (LC-MS), single nuclei RNA sequencing (snRNAseq), and Cytometry by time of flight (CyTOF). Results: Inhaled benzene exposure significantly increased lesions in the aortic valves (>25%, P<0.05). Single cell RNA sequencing of the aortae of benzene-exposed mice revealed 11 major cell types (e.g., vascular smooth muscle cells (VSMCs), endothelial cells, macrophages, T cells, etc.) categorized into 23 sub-populations. snRNAseq and CyTOF analysis showed that benzene exposure robustly increased (P<0.05) CD4 + and CD8 + T-cells, and B-cells in the lesions. Clustering of T cells showed significantly higher expression (P<0.05) of CD4, CD8, and Th17 marker RAR Related Orphan Receptor C (RORc). snRNAseq data also suggested increased cell proliferation and >1.5-fold increase in VSMC number following benzene exposure. LC-MS analysis revealed that benzene exposure differentially regulated the abundance of 930 proteins (at least 2 two unique peptides) in innominate plaque. Importantly, several of the proteins involved in an extracellular matrix organization and elastic fibre formation (e.g. Elastin, Fibronectin , and Latent transforming growth factor beta binding protein 1 ) were downregulated in benzene-exposed mice. These data agree with the snRNAseq data. In vitro , benzene metabolite hydroquinone (HQ, 1 μM) induced ROR-gamma, the transcription factor that regulates Th17, in spleen-derived CD4 + T cells (P<0.05). HQ (0.1 μM) also increased protein kinase C activity and proliferation of human aortic smooth muscle cells (P<0.001). Conclusion: Benzene exposure exacerbates atherosclerosis and compromises plaque cellularity and stability.

Electronic nicotine delivery systems (ENDS) aerosol exposures can induce endothelial dysfunction (ED) in healthy young humans and animals. Thermal degradation of ENDS solvents, propylene glycol, and vegetable glycerin (PG: VG), generates abundant formaldehyde (FA) and other carbonyls. Because FA can activate the transient receptor potential ankyrin-1 (TRPA1) sensor, we hypothesized that FA in ENDS aerosols provokes TRPA1-mediated changes that include ED and "respiratory braking"-biomarkers of harm. To test this, wild-type (WT) and TRPA1-null mice were exposed by inhalation to either filtered air, PG: VG-derived aerosol, or FA (5 ppm). Short-term exposures to PG: VG and FA-induced ED in female WT but not in female TRPA1-null mice. Moreover, acute exposures to PG: VG and FA stimulated respiratory braking in WT but not in TRPA1-null female mice. Urinary metabolites of FA (ie, N-1,3-thiazolidine-4-carboxylic acid, TCA; N-1,3-thiazolidine-4-carbonyl glycine, TCG) and monoamines were measured by LC-MS/MS. PG: VG and FA exposures significantly increased urinary excretion of both TCA and TCG in both WT and TRPA1-null mice. To confirm that inhaled FA directly contributed to urinary TCA, mice were exposed to isotopic 13C-FA gas (1 ppm, 6 h). 13C-FA exposure significantly increased the urine level of 13C-TCA in the early collection (0 to 3 h) supporting a direct relationship between inhaled FA and TCA. Collectively, these data suggest that ENDS use may increase CVD risk dependent on FA, TRPA1, and catecholamines, yet independently of either nicotine or flavorants. This study supports that levels of FA in ENDS-derived aerosols should be lowered to mitigate CVD risk in people who use ENDS.

Background: The WHO estimates that air pollution causes 7 million premature deaths or about 1 in 8 global deaths. Epidemiological studies indicate that 60-70% of the premature mortality attributed to air pollution are cardiovascular deaths especially in those with pre-existing conditions such as hypertension and heart failure. The underlying pathophysiological mechanisms by which exposures to air pollution worsen cardiovascular disease are unclear. Hypothesis: We hypothesized that the cardiovascular toxicity of particulate air pollution (PM 2.5 ) exposure would be enhanced in the setting of hypertension. Methods: To test this, we combined air pollution exposure with a hypertension model (angiotensin II, 2.5 mg/kg bwt/day: ANGII osmotic pump) where normotensive and hypertensive male wildtype (WT, C57BL/6J) mice were exposed to filtered air or concentrated ambient PM 2.5 (CAP) for 3 weeks. To understand how combined hypertension and CAP exposure may alter cardiac remodeling, fibrosis and gene transcription (bulk RNAseq) were quantified. Results: Mice with ANGII-infusion developed hypertension (non-invasive tail cuff) that was significantly elevated by CAP exposure. Hypertensive mice also developed cardiac hypertrophy (heart weight/tibia length ratio, mg/mm) independent of exposure [hypertensive: WT+Air, 9.6±0.4; WT+CAP, 10.3±0.4; normotensive groups: WT+Air, 8.3±0.3; WT+CAP, 7.4±0.1). CAP exposure had no effect on differential gene transcription in normotensive mice, yet CAP exposure significantly induced 996 differentially expressed genes (DEG) in hypertensive mice (332 up, 664 down). Gene Ontogeny (GO) analysis found dysregulated gene clusters (>40 genes) primarily for cardiac and striated muscle development and differentiation. Increased genes included caspase 12 ( Casp12 ) and catechol- O -methyltransferase ( Comt) genes that likely reflect enhanced apoptosis and sympathetic input. Downregulated genes included 2 collagen genes ( Col5a3 and Col6a3 ) and death inducer-obliterator 1 ( Dido1 ) -- reflecting dysregulated cardiac remodeling. Conclusions: Hypertension enhanced the susceptibility of short-term air pollution exposure to worsen cardiovascular effects especially cardiac remodeling. This study reveals potential genetic mechanisms by which air pollution hastens cardiac dysregulation and promotes heart failure – a serious, globally relevant cardiovascular health risk of particulate air pollution.

Environmental pollution causes cardiovascular disease, heart failure, and arrythmias [1–4]. Wildfire emissions are a complex mixture composed of particulate matter (PM), carbon monoxide, methane, nitrous oxide, and polyaromatic hydrocarbons, among others [5], which are linked to arrhythmias [6]. Benzo[a]pyrene is a polyaromatic hydrocarbon and known carcinogen in animal models and is implicated in breast cancer, lung cancer, liver cancer, and skin cancer [7]. Therefore, the further impact of BaP on the cardiovascular system merits further investigation.

Background: Benzene is a ubiquitous environmental pollutant generated by a variety of natural and anthropological sources. It is a known carcinogen and hematopoietic toxin; however, little is known about benzene's potential atherogenicity. Hypothesis: Inhaled benzene induces atherogenesis by increasing vascular inflammation in LDL receptor Knockout (LDLR-KO) mice. Methods: Male LDLR-KO mice were exposed to HEPA-filtered air or benzene (1 ppm, 6h/day, 5days/week) for 24 weeks. For the last 12 weeks of exposure, the mice were maintained on a western diet. The single nuclei RNA sequencing (snRNAseq) of aortae was performed at Novogene. In an independent experiment, male ApoE-KO mice were maintained on a western diet for 12 weeks and then exposed to benzene for 2 weeks. Cytometry by time of flight (CyTOF) analysis was used for the plaque immunophenotyping. For in vitro experiments, splenic naïve T cells were exposed to 1 μM of hydroquinone (HQ) for 24 hours, and intracellular ROR-gamma levels were measured by flow cytometry. Results: Benzene inhalation increased the aortic valve lesion area by more than 25% (P<0.05) in LDLR-KO mice. Using snRNAseq, eleven major cell types were detected, including T cells and vascular smooth muscle cells (VSMC). Benzene increased the number of T cells by 2.5-fold, proliferating T-cells by 5.8-fold, and VSMC by 1.6-fold, suggesting increased cellularity and reduced plaque stability. In addition, benzene upregulated Th17 polarization marker Rorc and negative regulators of apoptosis Rag1 and Bcl11b while significantly attenuating the expression of proliferation inhibitor Ms4a4b in T cells. In VSMC, benzene downregulated extracellular matrix organization genes and upregulated platelet degranulation pathways. CyTOF analysis of the plaques of ApoE-KO mice showed that even short-term benzene exposure increased T-cell abundance in pre-existing lesions. Polarization of T cells into Th17 was confirmed by HQ-dependent upregulation of ROR-gamma in vitro . Conclusion: Our data suggest that inhaled benzene exposure compromises plaque cellularity and stability by facilitating T-cell proliferation and polarization, which coincides with the degradation of smooth muscle extracellular matrix and platelet activation.

Aims Gene therapies to induce cardiomyocyte (CM) cell cycle re-entry have shown a potential to treat subacute ischaemic heart failure (IHF) but have not been tested in the more relevant setting of chronic IHF. Our group recently showed that polycistronic non-integrating lentivirus encoding Cdk1/CyclinB1 and Cdk4/CyclinD1 (TNNT2-4Fpolycistronic-NIL) is effective in inducing CM cell cycle re-entry and ameliorating subacute IHF models and preventing the subsequent IHF-induced congestions in the liver, kidneys, and lungs in rats and pigs. Here, we aim to test the long-term efficacy of TNNT2-4Fpolycistronic-NIL in a rat model of chronic IHF, a setting that differs pathophysiologically from subacute IHF and has greater clinical relevance.Methods and results Rats were subjected to a 2-h coronary occlusion followed by reperfusion; 4 weeks later, rats were injected intramyocardially with either TNNT2-4Fpolycistronic-NIL or LacZ-NIL. Four months post-viral injection, TNNT2-4Fpolycistronic-NIL-treated rats showed a significant reduction in scar size and a significant improvement in left ventricular (LV) systolic cardiac function but not in the LV dilatation associated with chronic IHF. A mitosis reporter system developed in our lab showed significant induction of CM mitotic activity in TNNT2-4Fpolycistronic-NIL-treated rats.Conclusion This study demonstrates, for the first time, that TNNT2-4Fpolycistronic-NIL gene therapy induces CM cell cycle re-entry in chronic IHF and improves LV function, and that this salubrious effect is sustained for at least 4 months. Given the high prevalence of chronic IHF, these results have significant clinical implications for developing a novel treatment for this deadly disease.

Exposure to plants is known to improve physical and mental health and living in areas of high vegetation is associated with better health. The addition of quantitative measures of greenness exposure at individual-level to other objective and subjective study measures will help establish cause-and-effect relationships between greenspaces and human health. Because limonene is one of the most abundant biogenic volatile organic compounds emitted by plants, we hypothesized that urinary metabolites of inhaled limonene can serve as biomarkers of exposure to greenness. To test our hypothesis, we analyzed urine samples collected from eight human volunteers after limonene inhalation or after greenness exposure using liquid chromatography-high resolution mass spectrometry-based profiling. Eighteen isomers of nine metabolites were detected in urine after limonene inhalation, and their kinetic parameters were estimated using nonlinear mixed effect models. Urinary levels of most abundant limonene metabolites were elevated after brief exposure to a forested area, and the ratio of urinary limonene metabolites provided evidence of recent exposure. The identities and structures of these metabolites were validated using stable isotope tracing and tandem mass spectral comparison. Together, these data suggest that urinary metabolites of limonene, especially uroterpenol glucuronide and dihydroperillic acid glucuronide, could be used as individualized biomarkers of greenness exposure.

Pod-based electronic (e-) cigarettes more efficiently deliver nicotine using a protonated formulation. The cardiovascular effects associated with these devices are poorly understood. We evaluated whether pod-based e-liquids and their individual components impair endothelial cell function. We isolated endothelial cells from people who are pod users (n = 10), tobacco never users (n = 7), and combustible cigarette users (n = 6). After a structured use, pod users had lower acetylcholine-mediated endothelial nitric oxide synthase (eNOS) activation compared with never users and was similar to levels from combustible cigarette users (overall P = 0.008, P = 0.01 pod vs never; P = 0.96 pod vs combustible cigarette). The effects of pod-based e-cigarettes and their constituents on vascular cell function were further studied in commercially available human aortic endothelial cells (HAECs) incubated with flavored JUUL e-liquids or propylene glycol (PG):vegetable glycerol (VG) at 30:70 ratio with or without 60 mg/mL nicotine salt for 90 min. A progressive increase in cell death with JUUL e-liquid exposure was observed across 0.0001–1% dilutions; PG:VG vehicle with and without nicotine salt induced cell death. A23187-stimulated nitric oxide production was decreased with all JUUL e-liquid flavors, PG:VG and nicotine salt exposures. Aerosols generated by JUUL e-liquid heating similarly decreased stimulated nitric oxide production. Only mint flavored e-liquids increased inflammation and menthol flavored e-liquids enhanced oxidative stress in HAECs. In conclusion, pod e-liquids and their individual components appear to impair endothelial cell function. These findings indicate the potential harm of pod-based devices on endothelial cell function and thus may be relevant to cardiovascular injury in pod type e-cigarette users.

Anthracycline-induced cardiotoxicity and potential interventions. (A) The mechanisms of anthracycline toxicity. Anthracycline causes dilated cardiomyopathy that can result in heart failure and arrhythmia. The mechanism of anthracycline-induced cardiotoxicity is dose dependent. Anthracycline-induced cardiotoxicity is related to cardiomyocyte mitochondrial fission, necrosis, and autophagy. (B) Therapies that attenuate the effects of anthracycline on the heart include: afzelin, carvedilol, and GSK2795039 that mitigate the adverse effects of anthracyclines in vitro and in vivo.Unlabelled Image

Although Electronic Nicotine Delivery Systems (ENDS)-derived aerosols induce endothelial dysfunction (ED), the mechanisms and constituents responsible for ED are unknown. Because ENDS use propylene glycol and vegetable glycerin (PG:VG) that thermally decompose into aldehydes, we hypothesized that acrolein and formaldehyde stimulate ED via the sensory transient receptor potential ankyrin-1 (TRPA1) receptor and the autonomic nervous system (ANS). To test this, female wild-type (WT) and TRPA1-null mice were exposed to either filtered air, PG:VG-derived aerosol, acrolein (AC; 1 ppm), or formaldehyde (FA, 5 ppm). Biomarkers of exposure (AC: 3HPMA; FA: 4TCA) and of harm (aortic ED; urinary catecholamines) were measured. The PG:VG and AC exposures significantly increased urinary 3HPMA levels; and the PG:VG and FA exposures significantly increased urinary 4TCA levels vs air controls. Both PG:VG and FA exposures (6h/d x 4d) in WT mice led to aortic ED (-64.6±3.8; –66.7±5.5 % ACh, respectively) vs air control (-81.5±1.5 % ACh); yet AC exposure did not induce ED (-76±2.9 % ACh). Exposures in TRPA1-null mice to PG:VG, AC, or FA did not lead to ED (-93.0±3.6; –82.8±3.1; -86.6±2.8 % ACh; respectively). As an index of ANS activation and a potential mediator of ED, urinary catecholamines (dopamine, DA; norepinephrine, NE; epinephrine, EPI; serotonin, 5HT) and metabolites were measured by UPLC-MS/MS in early (0-3h) and late (3-18h) post-exposure (PE) samples. In WT mice, PG:VG significantly increased early PE urine levels of NE (and two NE metabolites; +1.5-2.5x), DA metabolite (+1.5x), but not EPI nor 5HT; FA exposure increased DA (+3x), EPI (+2.5x), NE and NE metabolites (+1.2-2.5x), 5HT and 5HT metabolite (+2.5x) only in late PE; while AC exposure decreased early PE DA (-73%) and 5HT (-42%) levels. In TRPA1-null mice, PG:VG and AC exposures increased early PE levels of NE (+2x) and depressed EPI and 5HT levels (-40%); whereas FA exposure increased NE (+2x), EPI (+4x), and 5HT (+1.5) levels in late PE. Aldehydes induce ED through time-dependent mechanisms mediated in part by TRPA1 activation and ANS stimulation. These data indicate that ENDS use may increase cardiovascular disease risk via aldehydes generated by PG:VG independent of nicotine and flavorants.