Inflammation and fibrosis analysis on a systemic and tissue level in adult patients with a systemic right ventricle

Abstract Background Adults with a systemic right ventricle (sRV) are prone to develop heart failure (HF) with poor prognosis due to lack of evidence for classic HF therapies and extensive fibrosis of non-ischemic origin. We previously reported more systemic inflammation in sRV patients with HF. Purpose To assess systemic and tissue-level inflammation- and fibrosis-related pathology in sRV-HF patients in comparison to patients with a normal heart anatomy and HF (systemic left ventricle-heart failure, sLV-HF). Methods Samples from sRV (n=87) and sLV-HF (n=30) patients were examined from a prospectively enrolled cohort as well as retrospective analysis of biobanked tissue at a secondary site. Plasma samples from volunteers age- and sex-matched to the sRV cohort (n=42) were analysed, in addition to cardiac tissue samples from non-failing individuals obtained commercially or through a local biobank. Tissue was analysed by Sirius Red, hematoxylin and eosin (H&E), and immuno- staining. In plasma, a panel of proinflammatory cytokines were measured along collagen markers. Additionally, neutrophil activation was assessed for degranulation (free myeloperoxidase (MPO)) or neutrophil extracellular trap release (DNA-complexed MPO or citrullinated histones (MPO-DNA or H3Cit-DNA complexes)). Kruskal-Wallis tests with post-hoc Dunn’s correction were performed. Results Histological analysis for fibrosis showed high collagen levels in the sRV, as well as in the subpulmonary LV as well as interventricular septum (Figure 1A). Identified patterns- interstitial, perivascular, replacement, and subendocardial fibrosis were comparable to sLV-HF patients. H&E staining revealed areas of granulocytic infiltration and intravascular accumulation (Figure 1B), confirmed by immunostaining which showed neutrophil-rich areas in less fibrotic tissue while more fibrotic regions had few neutrophils. We next aimed to quantify biomarkers in circulation. Plasma samples showed elevated levels of Type III procollagen peptide (PIIINP) in both sRV and sLV-HF cohorts compared to healthy controls (Figure 2A), indicating collagen turnover. These levels were already high in sRV patents without HF. Findings for proinflammatory cytokines were unremarkable, so we aimed to assess neutrophilic inflammation in more detail. MPO was significantly higher in both sRV and sLV-HF patient groups compared to healthy controls and were highest in sLV-HF patients (Figure 2B). Interestingly, NET biomarkers (H3Cit-DNA and MPO-DNA) were significantly higher in sRV patients compared to sLV-HF patients and healthy controls (Figure 2C). For both MPO and NETs, these elevated levels were independent of HF in sRV. Conclusions Extensive cardiac fibrosis is evident in sRV patients, as well as local inflammatory cell recruitment likely preceding fibrosis development. sRV patients have increased NET biomarkers, while sLV-HF patients have elevated MPO suggesting different neutrophil activation profiles in sRV patients.Figure 1 FIgure 2