Phd in PHARMACOLOGY AND TOXICOLOGY

Thesis title: Plant polyphenols and herbal extracts against damage induced by respiratory toxicants: novel strategies to improve antioxidant and antinflammatory host defences

Respiratory diseases pose a significant socio-economic burden, with viral infections and airway illnesses often associated with immune dysregulation and excessive inflammation driven by oxidative stress. In this study, polyphenol-based herbal extracts and their mixtures were investigated for their protective role against oxidative and inflammatory stress in airway models and their potential as novel preventive or adjuvant therapies to support antioxidant defense and immune response. To this end, high-quality extracts obtained from Humulus lupulus L. cones (hop) and Sambucus nigra L. flowers (elderberry), medicinal plants traditionally used for respiratory health, were evaluated in vitro for their antioxidant, anti-inflammatory, and immunomodulatory properties. Their antiviral effects against influenza A virus (PR8/H1N1) were also assessed in airway cell models. Herbal extracts are characterized by intrinsic chemical complexity and variability so, in order to promote reproducible effects, a phytochemical characterization, with a focus on phenolic compounds, was performed. To enhance their bioactivity, the extracts were formulated into mixtures at optimized ratios and concentrations. Combining different herbal extracts allowed for synergistic effects, improving efficacy while reducing doses and potential side effects. Special attention was given to hop-containing mixtures, as hop extracts exhibit potent antioxidant and antiviral properties but face market limitations due to agricultural challenges and climate-related production variability. The protective role of the extracts and their mixtures was assessed using in vitro airway cells and macrophages exposed to oxidative and inflammatory stressors, such as tert-butyl hydroperoxide (tBOOH), lipopolysaccharide (LPS) and influenza A virus (PR8/H1N1). Key parameters, including cell viability, intracellular reactive oxygen species (ROS) levels, pro-inflammatory cytokine release, macrophage phagocytic activity, and influenza viral, infective capacity and expression of several viral proteins, involved in different phases of virus life cycle titer, have been evaluated. Furthermore, the modulation of Nrf2, a key transcription factor regulating antioxidant defenses and interacting with NF-κB in inflammatory pathways, was investigated as a potential mechanism of action. Our results demonstrated that herbal extracts, particularly their mixtures, exhibited cytoprotective, anti-inflammatory, and antiviral properties, by effectively counteracting oxidative and inflammatory stress and modulating macrophage activity. Notably, hop-elderberry mixtures showed enhanced bioactivity compared to individual extracts, making them promising candidates for novel adjuvant or preventive strategies against respiratory diseases. These findings have led to a patent application by EPO s.r.l., which provided the studied extracts. Although further studies are needed to deepen toxicological profile and influence of in vivo metabolism, these results reinforce our interest in herbal extract combinations as multi-targeted strategies for the management of respiratory diseases.