The Adverse Outcome Pathway (AOP) framework provides a means to outline a knowledge-driven sequence of events from exposure to adverse outcome. As a concept, AOPs have been readily accepted by the toxicology community as a means to organize available mechanistic information linking exposures to toxic effects in a standardized way encompassing all organizational levels of a given biological system. However, there is also an inherent benefit in applying AOPs to health outcomes that are not necessarily linked to standard toxicological end points, for example, in the context of predicting disease risk or in drug development. In this study, we propose an AOP for decreased lung function that originates in oxidative stress-mediated epidermal growth factor receptor activation in the airway epithelium. This article provides an overview of the supporting evidence for key events (KEs) on the molecular, cellular, tissue/organ, and organism level and how they relate to each other through key event relationships (KERs). The essentiality of the identified KEs, as well as the biological plausibility of the KERs, is also assessed. Moreover, the envisioned application of the proposed AOP is discussed, highlighting the utility of the AOP framework for developing preclinical tests that could prove useful in public health risk assessment for long-term e-cigarette use.

The use of electronic cigarettes (e-cigarettes) potentially offers a safer alternative to conventional tobacco products. The advance in molecular biology and computational sciences offers new perspective to assess adverse biological responses for product risk assessment by combining omics screens with knowledge-based biological pathways. Our aim was to compare transcriptomic perturbations in MucilAirÔ, a commercially available lung epithelial tissue, after short repeated exposure to cigarette smoke (3R4F) and e-cigarette (Vype ePen) aerosols. We performed deep RNA sequencing and secreted inflammatory cytokine profiling postexposure. One hundred twenty-three genes were differentially expressed at fold change (FC) >1.5 and p-false discovery rate (pFDR) <0.1 for 3R4F exposure and 0 genes for Vype ePen aerosol exposure. When a relaxed filter pFDR <0.5 and FC >1.5 was applied, 29 genes were identified with e-cigarette aerosol exposure and used for validation of potential candidates by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Gene enrichment analysis was conducted and predicted a response to 3R4F smoke exposure in biological processes involving inflammation and oxidative stress pathways. No enrichment could be performed for Vype ePen aerosol exposure due to the lack of regulated gene candidates at those exposure conditions even after qRT-PCR validation. Of a panel of 33 cytokines screened, 8 were upregulated (FC >1.5 p < 0.05) following 3R4F smoke exposure, which was in agreement with our enrichment analysis. In conclusion, aerosol from the tested e-cigarette caused limited perturbations in gene and inflammatory cytokine expression compared to conventional cigarette smoke, as assessed using next-generation sequencing-based systems biology approaches in 3D commercially available reconstituted lung epithelial tissues.

Hypertension is a cardiovascular risk factor that has a profound influence on cardiovascular morbidity and mortality. While good progress has been made in terms of identifying and managing this risk factor for patient care, methods to assess the potential of chemical compounds to induce hypertension or to assess the efficacy of consumer products (e.g., e-cigarettes) targeted at reducing disease burden remain largely limited to epidemiological associations and in vivo studies. The field of toxicology has undergone a paradigm shift toward the replacement of in vivo testing in toxicological risk assessment. The adverse outcome pathway (AOP) framework could facilitate improved knowledge-based risk/benefit assessment of chemicals or consumer products, respectively, without the necessity of animal testing. Furthermore, to facilitate a more timely, cost effective, and ethical solution for risk/efficacy assessment purposes, integrated testing strategies are required, which do not heavily rely on in vivo studies. In this study, we present the supporting information on an AOP describing how vascular endothelial peptide oxidation leads to hypertension through perturbation of endothelial nitric oxide bioavailability, leading to impaired vasodilation. We also discuss the essentiality of the key events (KEs), and biological plausibility and empirical support of KE relationships, in accordance with the Organisation for Economic Cooperation and Development (OECD) handbook. This AOP could be a useful tool to serve as a foundation for a future integrated testing strategy for the regulatory assessment of the harm reduction potential of e-cigarettes relative to conventional tobacco products and other consumer products, which aim to reduce cardiovascular disease risk.