Aspiration lung disease in children with severe neurodisability

Background: Children with severe neurodisability (ND) commonly suffer with respiratory disease and this is the leading cause of premature death. The nature of this respiratory disease is however, poorly understood. The underlying aetiology is often multifactorial, but aspiration (direct or reflux) is likely to play a key role. Unfortunately, diagnostic tests for reflux or direct aspiration are limited. There is a clinical need for high quality research on children with severe ND to define underlying mechanisms of respiratory disease and guide management. Aims: In this study, the aims were to (i) characterise respiratory symptoms and their relationship with lower airway inflammation in children with severe ND, (ii) explore available bronchial lavage (BAL) biomarkers of reflux aspiration, assessing their validity and their relationship to clinical and airway inflammatory data, (iii) develop a novel assay for the accurate detection/quantification of pepsin in BAL, (iv) investigate the validity of an alpha-amylase assay in paediatric BAL and explore the relationship of alpha-amylase levels with lower airway inflammation and clinical symptoms, and (v) investigate the effects of pH alteration and pepsin exposure on airway epithelial cells (AEC). Methods: Clinical data and BAL samples were collected from children with severe ND at times of stability and respiratory deterioration and also from healthy controls. BAL differential cell counts and cytokine measurements (ELISA) were performed. Lower airway microbial colonisation/infection was assessed. BAL pepsin measurement was attempted using a number of methods, and the feasibility of inhibitor affinity enrichment and LC-MRM-MS techniques to identify and quantify BAL pepsin was explored using SDS PAGE gel and mass spectrometry analysis. BAL alpha-amylase activity was measured using an ethylidene-pNP-G7 based assay. BEAS-2B cells were cultured in monolayer and the cytotoxic/inflammatory effects of pH alteration and pepsin exposure were explored through trypan blue staining ...