Bronchopulmonary dysplasia (BPD) is described as the multiple external injurious stimuli (oxygen toxicity, baro- and volu-trauma, and infection/inflammation) damage the immature, developing lung, and arrest lung development (Abman et al., 2017; Principi et al., 2018; Lignelli et al., 2019). A key histopathological feature of BPD is an alveolar simplification, which is characterized by fewer and larger alveoli (Hwang et al., 2018; Kalikkot et al., 2017). The pulmonary disfunction of BPD infants persists into adulthood, suggesting that lung injuries early in life may have lifelong consequences (Baraldi et al., 2007; Lignelli et al., 2019). BPD is no longer a disease limited to lung. BPD infants present decreased right ventricular function, increased cardiovascular risk, and higher incidence of arterial hypertension in adolescence and adulthood (Wilson-Costello et al., 2009; Lewandowski et al., 2013; Sipola-Leppänen et al., 2014; Gough et al., 2014). In addition, BPD is an independent risk factor for long-term neurodevelopmental impairment (Jensen et al., 2014; Baraldi et al., 2007; Nakanishi et al., 2016). Among 10-year-old children born extremely preterm, those who had BPD were at increased risk of cognitive and academic achievement (Sriram et al., 2018). Lowering the high incidence of BPD may be vital to improving long-term outcomes after extremely or very preterm birth. Unfortunately, current prevention strategies such as corticosteroids (Doyle et al., 2017), surfactant (Isayama et al., 2016), less invasive respiratory strategies (Sweet et al., 2019), careful oxygen usage (Saugstad Ola et al., 2018),and antioxidants (Poggi et al., 2014) have not been able to diminish the BPD incidence.
For the developing lung, oxygen toxicity is a significant trigger for BPD (Morty et al., 2018; Bhandari et al., 2010; Wang et al., 2018). Lung development progresses in five distinct stages: embryonic, pseudo glandular, canalicular, saccular, and alveolar (Kotecha et al., 2000). Human preterm infants who develop BPD are primarily born in the saccular stage of lung development (human 24–38 weeks of gestation age). In rodent models, the saccular stage of lung development begins at embryonic day 18 and continues through postnatal (PN) day 5 (Joshi et al., 2007). The rodent pups from birth exposed to high concentration oxygen have similar histological changes with hyperoxia-exposed human preterm infants (Nardiello et al., 2017; O'Reilly et al., 2014; Berger et al., 2014; Buczynski et al., 2013). Therefore, hyperoxia-exposure neonatal rat models are essential for better understanding the molecular mechanisms of BPD and testing the efficacy of potential therapeutic agents.
The molecular mechanisms of BPD are complex. The newborns pass from the hypoxic environment of the womb to the relatively hyperoxic extrauterine environment, and they experience increased oxidative stress. Preterm neonates are more susceptible to oxidative stress because of developmental deficits in antioxidant defenses (Kinsella et al., 2006; Negi et al., 2012). In addition, an additive oxygen therapy for the treatment of respiratory instabilities can increase oxidative stress (Perrone et al., 2016). Although premature infants are usually exposed to only the least required amount of supplemental oxygen, studies show considerable evidence of oxidant stress (Ozsurekci et al., 2016; Perrone et al., 2016). The excessive reactive oxygen species (ROS) trigger the oxidative stress response, and damage the developing lung. And oxidative stress can activate inflammatory responses, representing increased pro-inflammatory cytokines and chemokines, and inflammatory cell infiltration in lung tissue. The inflammatory cells produce more ROS to worsens the oxidative toxicity, which in turn recruits more inflammatory cells to the lung (Rosanna et al., 2012). These complex and cross-talk processes between oxidative stress and inflammation ultimately lead to lung injury and impairment of lung development, which contribute to BPD (Rosanna et al., 2012; Balany et al., 2015). However, the mechanism of the interaction between oxidative stress and inflammation has not yet been conclusively clarified.
The NLRP3 (NOD-, LRR-, and pyrin-domain containing protein 3) inflammasome is the most extensively studied inflammasome complex (Agostini et al., 2004), and NLRP3 inflammasome can be activated during oxidative stress and systemic infections (Groslambert et al., 2018; Yang et al., 2019). NLRP3 recruits ASC (apoptosis-associated Specklike protein containing CARD) and caspase-1, and format the NLRP3 inflammasome. The NLRP3 inflammasome mediates the post-translational processing of IL-1β by active caspase-1, which cleaves pro-IL-1β to produce mature IL-1β. Mature IL-1β interacts with its receptor, IL1R, to signal inflammatory pathways. Activation of the NLRP3 inflammasome is considered to be a two-step process that requires two signals: the first priming signal triggers nuclear factor-κB (NF-κB)-dependent upregulation of NLRP3 and pro-IL-1β expression (Bauernfeind et al., 2009); the second signal is to induce NLRP3 activation (Swanson et al., 2019; Yang et al., 2019). ROS has been considered as a trigger of NLRP3 inflammasome activation (Teng et al., 2020; Wei et al., 2020), and many chemical compounds are reported to activate NLRP3 inflammasome via increasing intracellular ROS levels (Zhong et al., 2013). In addition, ROS can activate the NF-κB signaling pathway, resulting in the amplification of inflammatory response (Zha et al., 2014). ROS-mediated NF-κB signaling plays an essential role in activating the NLRP3 inflammasome (Teng et al., 2020; Peng et al., 2020). Therefore, ROS/NF-κB/NLRP3 inflammasome signaling is closely associated with the overlapping or feedback pathways between oxidative stress and inflammatory response.
Caffeine is a kind of plant alkaloid, and exhibits practical benefit in the treatment or prevention of BPD in several clinical studies (Coulter et al., 2006; Sanchez-Solis et al., 2020; Pakvasa et al., 2018). Endesfelder et al. reported that Caffeine prevented oxygen-induced inflammatory lung injury in neonatal rats (Endesfelder et al., 2020). Chen et al. reported that Caffeine treatment could protect hyperoxia-induced mice lung from oxidative damage by inhibiting NLRP3 inflammasome and NF-κB pathway (Chen et al., 2020). Besides Caffeine, others plant-derived compounds are beneficial to human health. Glycyrrhiza radix is one of the most popular phytochemicals, widely used in the medicine and food industries (Pastorino et al., 2018; El-Saber et al., 2020). Triterpene glycoside 18β-glycyrrhetinic acid (18β-GA), extracted from Glycyrrhiza radix, is a promising bioactive compound with anti-oxidative and anti-inflammatory properties (Pastorino et al., 2018; El-Saber et al., 2020). Animal studies have confirmed that 18β-GA has a protective effect against pulmonary diseases, such as asthma, pulmonary hypertension, pulmonary infection, and pulmonary fibrosis (Kim et al., 2017; Zhang et al., 2019; Chen et al., 2017; Kao et al., 2010). All of these studies have reflected that 18β-GA has beneficial effects on lungs.
Based on these considerations, we hypothesized that 18β-GA had a protective effect in neonatal rats with hyperoxia exposure. In this study, we investigated the role of ROS/NF-κB/NLRP3 inflammasome in this process.