Identification of autoantigens and their potential post-translational modification in EGPA and severe eosinophilic asthma.

BACKGROUND: The chronic airway inflammation in severe eosinophilic asthma (SEA) suggests potential autoimmune aetiology with unidentified autoantibodies analogous to myeloperoxidase (MPO) in ANCA-positive EGPA (eosinophilic granulomatosis with polyangiitis). Previous research has shown that oxidative post-translational modification (oxPTM) of proteins is an important mechanism by which autoantibody responses may escape immune tolerance. Autoantibodies to oxPTM autoantigens in SEA have not previously been studied. METHODS: Patients with EGPA and SEA were recruited as well as healthy control participants. Autoantigen agnostic approach: Participant serum was incubated with slides of unstimulated and PMA-stimulated neutrophils and eosinophils, and autoantibodies to granulocytes were identified by immunofluorescence with anti-human IgG FITC antibody. Target autoantigen approach: Candidate proteins were identified from previous literature and FANTOM5 gene set analysis for eosinophil expressed proteins. Serum IgG autoantibodies to these proteins, in native and oxPTM form, were detected by indirect ELISA. RESULTS: Immunofluorescence studies showed that serum from patients with known ANCA stained for IgG against neutrophils as expected. In addition, serum from 9 of 17 tested SEA patients stained for IgG to PMA-stimulated neutrophils undergoing NETosis. Immunofluorescent staining of eosinophil slides was evident with serum from all participants (healthy and with eosinophilic disease) with diffuse cytoplasmic staining except for one SEA individual in whom subtle nuclear staining was evident. FANTOM5 gene set analysis identified TREM1 (triggering receptor expressed on myeloid cells 1) and IL-1 receptor 2 (IL1R2) as eosinophil-specific targets to test for autoantibody responses in addition to MPO, eosinophil peroxidase (EPX), and Collagen-V identified from previous literature. Indirect ELISAs found high concentrations of serum autoantibodies to Collagen-V, MPO, and TREM1 in a higher proportion of SEA patients than healthy controls. High concentrations of serum autoantibodies to EPX were evident in serum from both healthy and SEA participants. The proportion of patients with positive autoantibody ELISAs was not increased when examining oxPTM compared to native proteins. DISCUSSION: Although none of the target proteins studied showed high sensitivity for SEA, the high proportion of patients positive for at least one serum autoantibody shows the potential of more research on autoantibody serology to improve diagnostic testing for severe asthma. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, identifier, NCT04671446

Deleterious Effects of Cold Air Inhalation on Coronary Physiological Indices in Patients With Obstructive Coronary Artery Disease

Background Cold air inhalation during exercise increases cardiac mortality, but the pathophysiology is unclear. During cold and exercise, dual‐sensor intracoronary wires measured coronary microvascular resistance (MVR) and blood flow velocity (CBF), and cardiac magnetic resonance measured subendocardial perfusion. Methods and Results Forty‐two patients (62±9 years) undergoing cardiac catheterization, 32 with obstructive coronary stenoses and 10 without, performed either (1) 5 minutes of cold air inhalation (5°F) or (2) two 5‐minute supine‐cycling periods: 1 at room temperature and 1 during cold air inhalation (5°F) (randomized order). We compared rest and peak stress MVR, CBF, and subendocardial perfusion measurements. In patients with unobstructed coronary arteries (n=10), cold air inhalation at rest decreased MVR by 6% (P=0.41), increasing CBF by 20% (P<0.01). However, in patients with obstructive stenoses (n=10), cold air inhalation at rest increased MVR by 17% (P<0.01), reducing CBF by 3% (P=0.85). Consequently, in patients with obstructive stenoses undergoing the cardiac magnetic resonance protocol (n=10), cold air inhalation reduced subendocardial perfusion (P<0.05). Only patients with obstructive stenoses performed this protocol (n=12). Cycling at room temperature decreased MVR by 29% (P<0.001) and increased CBF by 61% (P<0.001). However, cold air inhalation during cycling blunted these adaptations in MVR (P=0.12) and CBF (P<0.05), an effect attributable to defective early diastolic CBF acceleration (P<0.05) and associated with greater ST‐segment depression (P<0.05). Conclusions In patients with obstructive coronary stenoses, cold air inhalation causes deleterious changes in MVR and CBF. These diminish or abolish the normal adaptations during exertion that ordinarily match myocardial blood supply to demand

Cardiac sympathetic dysfunction in the prehypertensive spontaneously hypertensive rat.

Recent studies in prehypertensive spontaneously hypertensive rats (SHR) have shown larger calcium transients and reduced norepinephrine transporter (NET) activity in cultured stellate neurons compared with Wistar-Kyoto (WKY) controls, although the functional significance of these results is unknown. We hypothesized that peripheral sympathetic responsiveness in the SHR at 4 wk of age would be exaggerated compared with the WKY. In vivo arterial pressure (under 2% isoflurane) was similar in SHRs (88 ± 2/50 ± 3 mmHg, n = 18) compared with WKYs (88 ± 3/49 ± 4 mmHg, n = 20). However, a small but significant (P &lt; 0.05) tachycardia was observed in the young SHR despite the heart rate response to vagus stimulation (3 and 5 Hz) in vivo being similar (SHR: n = 12, WKY: n = 10). In isolated atrial preparations there was a significantly greater tachycardia during right stellate stimulation (5 and 7 Hz) in SHRs (n = 19) compared with WKYs (n = 16) but not in response to exogenous NE (0.025-5 μM, SHR: n = 10, WKY: n = 10). There was also a significantly greater release of [(3)H]NE to field stimulation (5 Hz) of atria in the SHR (SHR: n = 17, WKY: n = 16). Additionally, plasma levels of neuropeptide Y sampled from the right atria in vivo were also higher in the SHR (ELISA, n = 12 for both groups). The difference in [(3)H]NE release between SHR and WKY could be normalized by the NET inhibitor desipramine (1 μM, SHR: n = 10, WKY: n = 8) but not the α2-receptor antagonist yohimbine (1 μM, SHR: n = 7, WKY: n = 8). Increased cardiac sympathetic neurotransmission driven by larger neuronal calcium transients and reduced NE reuptake translates into enhanced cardiac sympathetic responsiveness at the end organ in prehypertensive SHRs