Analysis of nenatal immunity: innate lymphoid cells in cord blood and oxidative stress in exhaled breath condensate of children born preterm

Abstract

Project 1: Recent studies in mice and humans suggest that asthma is not just an adaptive IgE-mediated allergic inflammatory disease driven by T helper lymphocytes. There is considerable data proposing that the pathogenesis also involves a novel cell population of the innate immune system called type 2 innate lymphoid cells (ILC2s). These cells do not express the surface markers of T lymphocytes, but have the capacity to produce IL-4, IL-5 and IL-13. ILC2s are induced by the innate epithelial cytokines IL-33, IL-25 and TSLP and are considered important in initiating allergic type 2 immune responses and contribute to the persistent airway inflammation associated with severe asthma. Our laboratory has recently found that children with severe asthma and continuous symptoms despite high doses of corticosteroids have a greater number of ILC2s in bronchoalveolar lavage, sputum and blood compared to non-asthmatic children. It is proposed that persistent symptoms and eosinophilic inflammation in severe asthma that is relatively resistant to steroid therapy is mediated by ILC2s. We therefore aimed to culture a purified population of these cells to study their functional responses in vitro to innate epithelial cytokines with and without steroids. ILCs were quantified in tonsils and cord blood by means of flow cytometry. The proportion of ILCs identified in tonsils was very low and inadequate for culture. For this reason the experiments focused on ILCs isolated from cord blood. We showed: 1) a higher percentage of ILC2s in boys suggesting that ILC2s can be the reason why boys tend to have more atopic disease from birth until puberty; 2) ILC2s are capable of producing more IL-13 than CD4+ T cells and can be implicated in the Th2 response seen in pregnancy. When cord blood ILCs were cultured with different pro-inflammatory mediators with or without budesonide preliminary results suggest steroid sensitivity of ILCs.

Project 2: Oxidative stress, defined as an imbalance between antioxidant capacity and reactive oxygen species (ROS) generation, may play an important role in many diseases of prematurity. Respiratory distress syndrome (RDS) and eventually bronchopulmonary dysplasia (BPD) and pulmonary hypertension (PH), some of the most common complications of preterm newborns, may be mediated by the increased production of ROS. Furthermore, PH development may also depend on the reduced synthesis of airway nitric oxide (NO) caused by the endogenous inhibitor of NO endothelial synthase asymmetric dimethyl arginine (ADMA). To date, most of the studies performed in preterms have evaluated oxidative stress biomarkers and ADMA in peripheral sites such as cord blood or blood serum while in the airways a few has investigated oxidative stress and none ADMA. The purpose of the study was to validate exhaled breath condensate (EBC) collection in ventilated preterm babies at birth and in the first days of life with the aim of assessing oxidative stress biomarkers and ADMA values in their airways. EBC is a quick and non-invasive technique that can contribute in the biochemical and molecular understanding of some cellular processes occurring in the respiratory system. EBC biomarkers might help the clinicians to estimate the severity of pulmonary pathology and the risk of BPD and PH in children born preterm.