E-cigarettes (E-cigs), also known as electronic cigarettes or vapes, have risen in popularity as alternatives to traditional tobacco smoking. Marketed as safer choices, they have garnered significant attention for their perceived health advantages. Nonetheless, recent research indicates inherent risks associated with e-cigs. Therefore, understanding the toxicological effects of e-cigs in vitro (i.e., cells), in vivo (i.e., animals), and in humans is crucial for evaluating their overall safety and developing effective regulatory measures. While the in vitro system refers to experiments conducted outside of living organisms such as in culture flasks, petri dishes, or test tubes, the in vivo system offers more complex experimentation in living organisms such as in animals or humans. Furthermore, using an e-cig that contains nicotine increases the expression of cytokines, hyperreactivity of the airways, and damage to lung tissue. This chapter also investigates various mechanisms of toxicity from the perspectives of pulmonary, inflammation, oxidative stress, DNA damage, apoptosis, transcriptome alterations, and epithelial-mesenchymal transition (EMT), all of which are directly linked to the effects of e-cigs. As for humans, studies have shown that using e-cigs increases the risk of multiorgan involvement, particularly respiratory, cardiovascular, neurodevelopment, immune system, and heightened cancer risk.