Mechanistic Advances in the Therapeutic Application of Bixin for Lung Inflammation In Vitro and In Vivo

<b>Background:</b> Nrf2 plays a key role in regulating the antioxidant response against oxidative stress. Therefore, it is imperative to examine the advantages of Nrf2 activation by new small molecules capable of inhibiting the Nrf2-Keap1 protein interaction that do not present electrophilic sites, since electrophilic compounds have intrinsic toxicity. The bixin pigment has been used as a form of treatment and prevention of several pathological conditions in animal models since it was described as an Nrf2 activator without electrophilic sites. This study aims to synthetize a soluble derivate KBx (potassium bixinate) and evaluate its ability to activate Nrf2/ARE in a model of exposure to cigarette smoke extract (CSE; in vitro) and intranasal LPS administration (in vivo). <b>Methods:</b> In the in vivo study, C57BL/6 mice were pretreated with 200 mg/kg of KBx (gavage) during 5 consecutive days and then challenged with 60 µg of LPS i.n. for 16 h. Bronchoalveolar lavage was collected to examine cytokines dosage. In the in vitro study, RAW 264.7 macrophages were exposed to CSE and post-treated with KBx to evaluate their ability to revert the redox imbalance caused by the stressor. <b>Results:</b> KBx was characterized using mass spectrometry (433.1778 <i>m</i>/<i>z</i>). KC levels were increased in the LPS group (<i>p</i> = 0.021), and KBx inhibited this (<i>p</i> = 0.001). IL-10 levels were decreased (<i>p</i> = 0.055) in the LPS group that was prevented when pretreated with KBx (<i>p</i> = 0.037). The in vitro study showed KBx to be a more potent derivate of bixin through its ability to intercept ROS formation with three-fold more potency, and it showed an anti-inflammatory propriety by reducing the nuclear translocation of p65 (<i>p</i> < 0.001). <b>Conclusions:</b> In conclusion, these data suggest that KBx was able to activate the Nrf2/ARE pathway and intercept ROS formation induced by CSE and LPS in both in vivo and in vitro studies.