Acrolein (CH2=CHCHO) is a highly-reactive, volatile chemical that may be released accidentally or intentionally, resulting in eye, skin, and respiratory toxicity. Based on acrolein’s chemistry, it is known to target biological nucleophiles including guanine in DNA, as well as cysteine, lysine, histidine and arginine residues in critical regions of transcription factors, proteases and other proteins. Consequently, with such a broad range of potential biological targets it has been difficult to be certain of the most sensitive targets in vivo on which to base possible therapeutic interventions. Acrolein has been reported to be an agonist for the Ca2+-permeable transient receptor potential cation channel, subfamily A, member 1 (TRPA1). Therefore, acrolein targeting TRPA1 on lung macrophages would be expected to increase inflammatory cytokine production. However, studies have reported that acrolein inhibits macrophage cytokine production. Acrolein is expected to target thiol groups that are common critical sites for activity of protein tyrosine phosphatases (PTP) making them an important and logical family of target proteins that would explain inhibition of macrophage cytokine production. Of the various PTP, PTP1B has been reported to be irreversibly inhibited by acrolein, but there are no reports that PTP1B is a target of acrolein toxicity in vivo. Since PTP1B is a key regulatory PTP involved in critical pathways and diseases and as a target of acrolein exposure could explain a number of the reported outcomes of acrolein exposure especially at sublethal concentrations. Since the precise mechanism of acrolein toxicity remains uncertain, this project will fill an important gap of knowledge to define in vitro and demonstrate for the first time in vivo, whether inhibition of PTP1B catalytic activity helps define the mechanism of acrolein-induced respiratory toxicity at sublethal concentrations. Studies in Aim 1 will Demonstrate that acrolein blocks the activity of PTP1B in macrophages and epithelial cells in vitro and will be compared to acrolein’s effects on TRPA1. Studies in Aim 2 will demonstrate that sublethal in vivo exposures of acrolein blocks the activity of PTP1B in lung macrophages to decrease inflammatory cytokine release from macrophages. These studies will provide new mechanistic information on acrolein toxicity in vitro and in vivo that will help explain the reported adverse impacts of acrolein exposure and provide guidance to develop medical countermeasures to alleviate the adverse effects of acrolein exposures.