Introduction: Because of the rising popularity of novel tobacco products, there has been an increasing interest in the biological effects of these products. This study was performed to investigate the biological effects of vapor from a novel tobacco vapor product (NTV) in comparison with cigarette smoke (CS) from a 3R4F reference cigarette by using two types of in vitro model. Materials and Methods: A two-dimensional (2D) culture system composed of a submerged monolayer of BEAS-2B cells was used to detect cellular oxidative stress responses. A three-dimensional (3D) culture system composed of a pseudostratified air-liquid interface culture of normal human bronchial epithelial cells was used to detect tissue inflammation. In both culture systems, exposures were conducted with an aqueous extract (AqE) of NTV vapor or CS. Results: In the 2D culture system, exposure to CS AqE induced glutathione oxidization and antioxidant response element reporter gene activation. NTV vapor AqE also induced reporter gene activity, but the concentration required for activation was higher than that of CS AqE. In the 3D culture system, exposure to CS AqE increased the secretion of various inflammatory mediators. In contrast, the effect of NTV vapor AqE on tissue inflammation was limited, and only interferon-c showed a slight increase in expression. Conclusions: Overall, these results demonstrate that NTV vapor has fewer effects on cellular oxidative stress and tissue inflammatory responses as compared with CS in in vitro models using human cells.