PROLONGED SARS-CoV-2 VIRAL BURDEN IN SIV-SARS-CoV-2 COINFECTED MACAQUES

Background: Individuals living with HIV are at increased risk of morbidity and mortality from COVID-19. Furthermore, SARS-CoV-2 infection in immunocompromised HIV infected individuals poses a risk to prolonged infection and viral shedding and the emergence of new variants of concern (VOCs). Using the SIV macaque model for AIDS, we are investigating the hypothesis that immune dysfunction during HIV infection will prolong SARSCoV- 2 viral infection, promote enhanced COVID-19 disease, and accelerate viral evolution. Here, we report the impact of SIV-CoV-2 co-infection on immune responses and pathogenesis. Method(s): Eight female rhesus macaques (aged 7-15 years, 5.5-9.9kg) were infected with SIVmac251 via low dose intravaginal challenge and then inoculated with 6.5x105 TCID50/mL SARS-CoV-2 (WA-1) at 17-34 weeks post-SIV infection via combined intranasal and intratracheal routes. Blood, bronchoalveolar lavage (BAL), stool, and nasal, oral, and rectal swabs were collected pre-infection through 14 days post-infection (DPI) to measure immune responses and viremia. ELISAs, ELISPOT, qRT-PCR, lung pathology, cytokine multiplex, and virus neutralization assays were performed to measure viral loads, pathogenesis, and immune responses. Result(s): Three days post-SARS-CoV-2 infection, we observed a transient decrease in CD4 counts, but there were no changes in clinical symptoms or plasma SIV viral loads. However, SARS-CoV-2 replication persisted in the upper respiratory tract, but not the lower respiratory tract. In addition, SARS-CoV-2 IgG seroconversion was delayed and antigen-specific T-cell responses were dampened. Notably, viral RNA levels in nasal swabs were significantly higher 7-14 DPI in SIV+ compared to previously published results using the same SARS-CoV-2 challenge virus in SIV- rhesus (PMCID: PMC8462335, PMC8829873). In addition, SIV/CoV-2 co-infected animals exhibited elevated levels of myeloperoxidase (MPO), a marker of neutrophil activation and increased lung inflammation. Conclusion(s): Here we provide evidence for the utility of the rhesus macaque in modeling human HIV-SARS-CoV-2 co-infection. Our results suggest that immunosuppression during SIV infection impairs de novo generation of anti-SARS-CoV-2 immunity, that may contribute to prolonged SARS-CoV-2 viral shedding, increased transmission windows, altered disease pathogenesis, and lower protection against subsequent SARS-CoV-2 exposures. Studies in progress will determine if SARS-CoV-2 viral evolution is accelerated in SIV-infected macaques.

Myeloperoxidase Inhibition in Heart Failure With Preserved or Mildly Reduced Ejection Fraction: SATELLITE Trial Results.

BACKGROUND: Inflammation is a key driver of heart failure with preserved left ventricular ejection fraction. AZD4831 inhibits extracellular myeloperoxidase, decreases inflammation, and improves microvascular function in preclinical disease models. METHODS AND RESULTS: In this double-blind phase 2a study (Safety and Tolerability Study of AZD4831 in Patients With Heart Failure [SATELLITE]; NCT03756285), patients with symptomatic heart failure, left ventricular ejection fraction of ≥40%, and elevated B-type natriuretic peptides were randomized 2:1 to once-daily oral AZD4831 5 mg or placebo for 90 days. We aimed to assess target engagement (primary end point: myeloperoxidase specific activity) and safety of AZD4831. Owing to coronavirus disease 2019, the study was terminated early after randomizing 41 patients (median age 74.0 years, 53.7% male). Myeloperoxidase activity was decreased by more than 50% from baseline to day 30 and day 90 in the AZD4831 group, with a placebo-adjusted decreased of 75% (95% confidence interval, 48, 88, nominal P < .001). No improvements were noted in secondary or exploratory end points, apart from a trend in Kansas City Cardiomyopathy Questionnaire overall summary score. No deaths or treatment-related serious adverse events occurred. AZD4831 treatment-related adverse events were generalized maculopapular rash, pruritus, and diarrhea (all n = 1). CONCLUSIONS: AZD4831 inhibited myeloperoxidase and was well tolerated in patients with heart failure and left ventricular ejection fraction of 40% or greater. Efficacy findings were exploratory owing to early termination, but warrant further clinical investigation of AZD4831. LAY SUMMARY: Few treatments are available for patients with the forms of heart failure known as heart failure with preserved or mildly reduced ejection fraction. Current treatments do not target inflammation, which may play an important role in this condition. We tested a new drug called AZD4831 (mitiperstat), which decreases inflammation by inhibiting the enzyme myeloperoxidase. Among the 41 patients in our clinical trial, AZD4831 had a good safety profile and inhibited myeloperoxidase by the expected amount. Results mean we can conduct further trials to see whether AZD4831 decreases the symptoms of heart failure and improves patients' ability to participate in physical exercise.

Bioavailability of iron and its potential to improve the immune system and ward off COVID-19: a review

Iron is a mineral that plays an important role, especially to prevent anaemia through the production of red blood cells. Iron also plays a role in physiological processes, such as the activation of enzymes and hormones, as well as increasing the immune system in warding off various viral infections. Therefore, iron bioavailability needs to be considered to take the greatest benefit of iron. This review discussed the factors that can affect the bioavailability of iron, various technologies to increase the bioavailability, and its potential in enhancing the immune system. Iron bioavailability can be increased by fortification, fermentation, the addition of vitamin C, and iron encapsulation. Under conditions of adequate iron intake, iron plays an important role in enhancing the immune system through controlling lymphocytes and T cell proliferation. However, excess iron consumption can be at risk of weakening the host's immune response to viruses. Therefore, the appropriate level of iron intake must be maintained accurately to be used optimally and has the potential to ward off viral infections, including the Sars-CoV-2 virus as the cause of COVID-19.Copyright © 2023, Rynnye Lyan Resources. All rights reserved.

Flexible Amperometric Immunosensor Based on Colloidal Quantum Dots for Detecting the Myeloperoxidase (MPO) Systemic Inflammation Biomarker.

Myeloperoxidase (MPO) has been demonstrated to be a biomarker of neutrophilic inflammation in various diseases. Rapid detection and quantitative analysis of MPO are of great significance for human health. Herein, an MPO protein flexible amperometric immunosensor based on a colloidal quantum dot (CQD)-modified electrode was demonstrated. The remarkable surface activity of CQDs allows them to bind directly and stably to the surface of proteins and to convert antigen-antibody specific binding reactions into significant currents. The flexible amperometric immunosensor provides quantitative analysis of MPO protein with an ultra-low limit of detection (LOD) (31.6 fg mL-1), as well as good reproducibility and stability. The detection method is expected to be applied in clinical examination, POCT (bedside test), community physical examination, home self-examination and other practical scenarios.

Inhibition of Myeloperoxidase Pro-Fibrotic Effect by Noscapine in Equine Endometrium.

Myeloperoxidase is an enzyme released by neutrophils when neutrophil extracellular traps (NETs) are formed. Besides myeloperoxidase activity against pathogens, it was also linked to many diseases, including inflammatory and fibrotic ones. Endometrosis is a fibrotic disease of the mare endometrium, with a large impact on their fertility, where myeloperoxidase was shown to induce fibrosis. Noscapine is an alkaloid with a low toxicity, that has been studied as an anti-cancer drug, and more recently as an anti-fibrotic molecule. This work aims to evaluate noscapine inhibition of collagen type 1 (COL1) induced by myeloperoxidase in equine endometrial explants from follicular and mid-luteal phases, at 24 and 48 h of treatment. The transcription of collagen type 1 alpha 2 chain (COL1A2), and COL1 protein relative abundance were evaluated by qPCR and Western blot, respectively. The treatment with myeloperoxidase increased COL1A2 mRNA transcription and COL1 protein, whereas noscapine was able to reduce this effect with respect to COL1A2 mRNA transcription, in a time/estrous cycle phase-dependent manner (in explants from the follicular phase, at 24 h of treatment). Our study indicates that noscapine is a promising drug to be considered as an anti-fibrotic molecule to prevent endometrosis development, making noscapine a strong candidate to be applied in future endometrosis therapies.

Nutritional deficiencies that may predispose to long COVID.

Multiple nutritional deficiencies (MND) confound studies designed to assess the role of a single nutrient in contributing to the initiation and progression of disease states. Despite the perception of many healthcare practitioners, up to 25% of Americans are deficient in five-or-more essential nutrients. Stress associated with the COVID-19 pandemic further increases the prevalence of deficiency states. Viral infections compete for crucial nutrients with immune cells. Viral replication and proliferation of immunocompetent cells critical to the host response require these essential nutrients, including zinc. Clinical studies have linked levels of more than 22 different dietary components to the likelihood of COVID-19 infection and the severity of the disease. People at higher risk of infection due to MND are also more likely to have long-term sequelae, known as Long COVID.

Antioxidant Strategies to Modulate NETosis and the Release of Neutrophil Extracellular Traps during Chronic Inflammation.

Extracellular traps are released by neutrophils and other immune cells as part of the innate immune response to combat pathogens. Neutrophil extracellular traps (NETs) consist of a mesh of DNA and histone proteins decorated with various anti-microbial granule proteins, such as elastase and myeloperoxidase (MPO). In addition to their role in innate immunity, NETs are also strongly linked with numerous pathological conditions, including atherosclerosis, sepsis and COVID-19. This has led to significant interest in developing strategies to inhibit NET release. In this study, we have examined the efficacy of different antioxidant approaches to selectively modulate the inflammatory release of NETs. PLB-985 neutrophil-like cells were shown to release NETs on exposure to phorbol myristate acetate (PMA), hypochlorous acid or nigericin, a bacterial peptide derived from Streptomyces hygroscopicus. Studies with the probe R19-S indicated that treatment of the PLB-985 cells with PMA, but not nigericin, resulted in the production of HOCl. Therefore, studies were extended to examine the efficacy of a range of antioxidant compounds that modulate HOCl production by MPO to prevent NETosis. It was shown that thiocyanate, selenocyanate and various nitroxides could prevent NETosis in PLB-985 neutrophils exposed to PMA and HOCl, but not nigericin. These results were confirmed in analogous experiments with freshly isolated primary human neutrophils. Taken together, these data provide new information regarding the utility of supplementation with MPO inhibitors and/or HOCl scavengers to prevent NET release, which could be important to more specifically target pathological NETosis in vivo.

Correlations of Myeloperoxidase (MPO), Adenosine deaminase (ADA), C-C motif chemokine 22 (CCL22), Tumour necrosis factor alpha (TNFα) and Interleukin-6 (IL-6) mRNA expression in the nasopharyngeal specimens with the diagnosis and severity of SARS-CoV-2 infections.

Cytokine dynamics in patients with coronavirus disease 2019 (COVID-19) have been studied in blood but seldomly in respiratory specimens. We studied different cell markers and cytokines in fresh nasopharyngeal swab specimens for the diagnosis and for stratifying the severity of COVID-19. This was a retrospective case-control study comparing Myeloperoxidase (MPO), Adenosine deaminase (ADA), C-C motif chemokine ligand 22 (CCL22), Tumour necrosis factor alpha (TNFα) and Interleukin-6 (IL-6) mRNA expression in 490 (327 patients and 163 control) nasopharyngeal specimens from 317 (154 COVID-19 and 163 control) hospitalized patients. Of the 154 COVID-19 cases, 46 died. Both total and normalized MPO, ADA, CCL22, TNFα, and IL-6 mRNA expression levels were significantly higher in the nasopharyngeal specimens of infected patients when compared with controls, with ADA showing better performance (OR 5.703, 95% CI 3.424-9.500, p < 0.001). Receiver operating characteristics (ROC) curve showed that the cut-off value of normalized ADA mRNA level at 2.37 × 10-3 had a sensitivity of 81.8% and specificity of 83.4%. While patients with severe COVID-19 had more respiratory symptoms, and elevated lactate dehydrogenase, multivariate analysis showed that severe COVID-19 patients had lower CCL22 mRNA (OR 0.211, 95% CI 0.060-0.746, p = 0.016) in nasopharyngeal specimens, while lymphocyte count, C-reactive protein, and viral load in nasopharyngeal specimens did not correlate with disease severity. In summary, ADA appears to be a better biomarker to differentiate between infected and uninfected patients, while CCL22 has the potential in stratifying the severity of COVID-19.

Myeloperoxidase-derived hypochlorous acid targets human airway epithelial plasmalogens liberating protein modifying electrophilic 2-chlorofatty aldehydes.

Neutrophil and airway epithelial cell interactions are critical in the inflammatory response to viral infections including respiratory syncytial virus, Sendai virus, and SARS-CoV-2. Airway epithelial cell dysfunction during viral infections is likely mediated by the interaction of virus and recruited neutrophils at the airway epithelial barrier. Neutrophils are key early responders to viral infection. Neutrophil myeloperoxidase catalyzes the conversion of hydrogen peroxide to hypochlorous acid (HOCl). Previous studies have shown HOCl targets host neutrophil and endothelial cell plasmalogen lipids, resulting in the production of the chlorinated lipid, 2-chlorofatty aldehyde (2-ClFALD). We have previously shown that the oxidation product of 2-ClFALD, 2-chlorofatty acid (2-ClFA) is present in bronchoalveolar lavage fluid of Sendai virus-infected mice, which likely results from the attack of the epithelial plasmalogen by neutrophil-derived HOCl. Herein, we demonstrate small airway epithelial cells contain plasmalogens enriched with oleic acid at the sn-2 position unlike endothelial cells which contain arachidonic acid enrichment at the sn-2 position of plasmalogen. We also show neutrophil-derived HOCl targets epithelial cell plasmalogens to produce 2-ClFALD. Further, proteomics and over-representation analysis using the ω-alkyne analog of the 2-ClFALD molecular species, 2-chlorohexadecanal (2-ClHDyA) showed cell adhesion molecule binding and cell-cell junction enriched categories similar to that observed previously in endothelial cells. However, in contrast to endothelial cells, proteins in distinct metabolic pathways were enriched with 2-ClFALD modification, particularly pyruvate metabolism was enriched in epithelial cells and mitochondrial pyruvate respiration was reduced. Collectively, these studies demonstrate, for the first time, a novel plasmalogen molecular species distribution in airway epithelial cells that are targeted by myeloperoxidase-derived hypochlorous acid resulting in electrophilic 2-ClFALD, which potentially modifies epithelial physiology by modifying proteins.

Neutrophil extracellular traps have auto-catabolic activity and produce mononucleosome-associated circulating DNA.

BACKGROUND: As circulating DNA (cirDNA) is mainly detected as mononucleosome-associated circulating DNA (mono-N cirDNA) in blood, apoptosis has until now been considered as the main source of cirDNA. The mechanism of cirDNA release into the circulation, however, is still not fully understood. This work addresses that knowledge gap, working from the postulate that neutrophil extracellular traps (NET) may be a source of cirDNA, and by investigating whether NET may directly produce mono-N cirDNA. METHODS: We studied (1) the in vitro kinetics of cell derived genomic high molecular weight (gHMW) DNA degradation in serum; (2) the production of extracellular DNA and NET markers such as neutrophil elastase (NE) and myeloperoxidase (MPO) by ex vivo activated neutrophils; and (3) the in vitro NET degradation in serum; for this, we exploited the synergistic analytical information provided by specifically quantifying DNA by qPCR, and used shallow WGS and capillary electrophoresis to perform fragment size analysis. We also performed an in vivo study in knockout mice, and an in vitro study of gHMW DNA degradation, to elucidate the role of NE and MPO in effecting DNA degradation and fragmentation. We then compared the NET-associated markers and fragmentation size profiles of cirDNA in plasma obtained from patients with inflammatory diseases found to be associated with NET formation and high levels of cirDNA (COVID-19, N = 28; systemic lupus erythematosus, N = 10; metastatic colorectal cancer, N = 10; and from healthy individuals, N = 114). RESULTS: Our studies reveal that gHMW DNA degradation in serum results in the accumulation of mono-N DNA (81.3% of the remaining DNA following 24 h incubation in serum corresponded to mono-N DNA); "ex vivo" NET formation, as demonstrated by a concurrent 5-, 5-, and 35-fold increase of NE, MPO, and cell-free DNA (cfDNA) concentration in PMA-activated neutrophil culture supernatant, leads to the release of high molecular weight DNA that degrades down to mono-N in serum; NET mainly in the form of gHMW DNA generate mono-N cirDNA (2 and 41% of the remaining DNA after 2 h in serum corresponded to 1-10 kbp fragments and mono-N, respectively) independent of any cellular process when degraded in serum; NE and MPO may contribute synergistically to NET autocatabolism, resulting in a 25-fold decrease in total DNA concentration and a DNA fragment size profile similar to that observed from cirDNA following 8 h incubation with both NE and MPO; the cirDNA size profile of NE KO mice significantly differed from that of the WT, suggesting NE involvement in DNA degradation; and a significant increase in the levels of NE, MPO, and cirDNA was detected in plasma samples from lupus, COVID-19, and mCRC, showing a high correlation with these inflammatory diseases, while no correlation of NE and MPO with cirDNA was found in HI. CONCLUSIONS: Our work describes the mechanisms by which NET and cirDNA are linked. In doing so, we demonstrate that NET are a major source of mono-N cirDNA independent of apoptosis and establish a new paradigm of the mechanisms of cirDNA release in normal and pathological conditions. We also demonstrate a link between immune response and cirDNA.

Withania somnifera (L.) Dunal as Add-On Therapy for COPD Patients: A Randomized, Placebo-Controlled, Double-Blind Study.

Background: The current gold-standard therapies for chronic obstructive pulmonary disease (COPD) lack disease-modifying potential and exert adverse side effects. Moreover, COPD patients are at a higher risk of severe outcomes if they get infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus, the cause of the current epidemic. This is the first study to document clinical research on an adaptogenic and steroidal activity-containing herb as a complementary medicine for COPD treatment. Objective: We aimed to evaluate the efficacy of Withania somnifera (L.) Dunal [Solanaceae] (WS) as an add-on therapy for COPD patients. Methods: A randomized, placebo-controlled, and double-blind clinical study was conducted. A total of 150 patients were randomly assigned to three groups: control, placebo, and WS group. In addition to conventional medicines, WS root capsules or starch capsules were given twice a day to the WS group and the placebo group, respectively. Their lung functioning, quality of life, exercise tolerance, systemic oxidative stress (OS), and systemic inflammation were assessed before and after 12 weeks of intervention. WS root phytochemicals were identified by LC-ESI-MS. The inhibitory activity of these phytochemicals against angiotensin-converting enzyme 2 (ACE-2); the SARS-CoV-2 receptor; myeloperoxidase (MPO); and interleukin-6 (IL-6) was evaluated by in silico docking to investigate the mechanism of action of WS. Results: The pulmonary functioning, quality of life, and exercise tolerance improved, and inflammation reduced notably the most in the WS group. Systemic oxidative stress subsided significantly only in the WS group. Although a minor placebo effect was observed in the SGRQ test, but it was not present in other tests. Withanolides found in the WS roots demonstrated substantial inhibitory activity against the proteins ACE-2, MPO, and IL-6, compared to that of a standard drug or known inhibitor. Moreover, FEV1% predicted had significant correlation with systemic antioxidative status (positive correlation) and malondialdehyde (MDA, negative correlation), suggesting that the antioxidative potential of WS has significant contribution to improving lung functioning. Conclusion: Our study clinically demonstrated that WS root when given along with conventional drugs ameliorated COPD significantly more in comparison to the conventional drugs alone, in GOLD 2 and 3 categories of COPD patients. In silico, it has potent inhibitory activity against SARS-CoV-2 receptor, ACE-2, MPO, and IL-6.

Clinical significance of neutrophil extracellular traps biomarkers in thrombosis.

Neutrophil extracellular traps (NETs) may be associated with the development of thrombosis. Experimental studies have confirmed the presence of NETs in thrombi specimens and potential role of NETs in the mechanisms of thrombosis. Clinical studies also have demonstrated significant changes in the levels of serum or plasma NETs biomarkers, such as citrullinated histones, myeloperoxidase, neutrophil elastase, nucleosomes, DNA, and their complexes in patients with thrombosis. This paper aims to comprehensively review the currently available evidence regarding the change in the levels of NETs biomarkers in patients with thrombosis, summarize the role of NETs and its biomarkers in the development and prognostic assessment of venous thromboembolism, coronary artery diseases, ischemic stroke, cancer-associated thromboembolism, and coronavirus disease 2019-associated thromboembolism, explore the potential therapeutic implications of NETs, and further discuss the shortcomings of existing NETs biomarkers in serum and plasma and their detection methods.

SEVERE IMMUNE-MEDIATED DIFFUSE ALVEOLAR HEMORRHAGE IN THE AGE OF COVID-19 AND EXTRACORPOREAL MEMBRANE OXYGENATION

SESSION TITLE: Pulmonary Manifestations of Infections SESSION TYPE: Case Reports PRESENTED ON: 10/17/2022 03:15 pm - 04:15 pm INTRODUCTION: Diffuse alveolar hemorrhage (DAH) due to an undiagnosed autoimmune condition is rare and can be life-threatening. Veno-venous extracorporeal membrane oxygenation (VV-ECMO) has been described as a viable rescue therapy in severe cases, providing time to establish a diagnosis and begin remission induction therapy (1). We report a patient who presented during the Omicron surge with hypoxemic respiratory failure due to pulmonary hemorrhage ultimately diagnosed with antineutrophil cytoplasmic antibody (ANCA) associated vasculitis (AAV) who was supported with VV-ECMO without systemic anticoagulation. CASE PRESENTATION: A 46-year-old woman presented with subacute fatigue and two days of cough and brown sputum. She was found to have normocytic anemia (hemoglobin 3.5 g/dL), renal failure (serum creatinine 17.4 µmol/L), and bilateral pulmonary infiltrates on chest roentgenogram. Though vaccinated, nasal molecular testing detected SARS-CoV-2. She was intubated for progressive hypoxic respiratory failure and bronchoalveolar lavage fluid was consistent with DAH. She received empiric antibiotics, remdesivir, and pulse dose intravenous methylprednisolone as well as continuous renal replacement therapy and plasma exchange. Due to refractory hypoxemia she was cannulated for VV-ECMO. Systemic anticoagulation was deferred due to concerns that it may exacerbate her underlying pathology and due to a small subcortical bleed seen on computed tomography of the head. Perinuclear ANCA (titer >1:1280) was confirmed by immunofluorescence analysis with elevated myeloperoxidase serologies and cyclophosphamide was initiated. Glomeruli with cellular crescent formation consistent with AAV was later identified on renal biopsy. Her course was complicated by recurrent DAH while tapering steroids and an iliac vein thrombus, extracted during decannulation. Her respiratory failure resolved and she was discharged to rehab. DISCUSSION: Traditionally, VV-ECMO obligates systemic anticoagulation to prevent circuit thrombosis, however this may be viewed as a barrier to its use in patients with prohibitive bleeding risk and may contribute to the therapy's overall morbidity. Some institutions have begun to demonstrate the safety of ECMO with low- or prophylactic doses of anticoagulation (2), but this practice remains controversial. Detection of SARS-CoV-2 posed diagnostic and management challenges and its significance to this case remains uncertain. There are many past examples of infectious triggers for both DAH and AAV, and there is emerging evidence for an association between SARS-CoV-2 and ANCA (3). Concerns regarding the risk of B-cell depletion influenced the selection of remission induction therapy. CONCLUSIONS: In the case described, a patient with severe DAH was successfully supported with VV-ECMO. Withholding systemic anticoagulation did not prevent recurrent bleeding and may have contributed to a deep vein thrombosis. Reference #1: Arnold S, Deja M, Nitschke M, Bohnet S, Wallis S, Humrich JY, Riemekasten G, Steinhoff J, Lamprecht P. Extracorporeal membrane oxygenation in ANCA-associated vasculitis. Autoimmun Rev. 2021 Jan;20(1):102702. doi: 10.1016/j.autrev.2020.102702. Epub 2020 Nov 11. PMID: 33188916. Reference #2: Kurihara C, Walter JM, Karim A, et al. Feasibility of Venovenous Extracorporeal Membrane Oxygenation Without Systemic Anticoagulation. Ann Thorac Surg. 2020;110(4):1209-1215. doi:10.1016/j.athoracsur.2020.02.011 Reference #3: Kadkhoda, K., Laurita, K. Antineutrophil cytoplasmic antibodies and their association with clinical outcomes in hospitalized COVID-19 patients. Cell Death Discov. 7, 277 (2021). DISCLOSURES: No relevant relationships by Nathaniel Nelson No relevant relationships by Radu Postelnicu no disclosure on file for Antonio Velez;

A CASE OF MICROSCOPIC POLYANGIITIS (MPA) FOLLOWING SARS-COV-2 INFECTION

SESSION TITLE: Unique Inflammatory and Autoimmune Complications of COVID-19 Infections SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: ANCA-associated vasculitis (AAV) is a systemic disease that causes inflammation of small vessels in various organs, such as the lungs, kidneys, and nervous system. We report a case of AAV following SARS-CoV-2 infection. CASE PRESENTATION: A 64-year-old female from Albania with no known medical history, presented with intermittent low-grade hemoptysis and fever for 1 week prior to admission. She had chills, myalgias, fatigue, and poor appetite 6 weeks prior. On arrival, she was febrile (101F), and hypoxic (Spo2 92% on 3L O2). Labs were significant for anemia [Hb 6.8 g/dl], acute kidney injury (AKI) [Cr 2.5 mg/dl]. She was found to be Positive for Sars-CoV-2 by PCR. Chest X-Ray showed patchy bilateral airspace opacities with peripheral and lower lobe predominance, concerning for atypical pneumonia (Fig 1). Urinalysis was significant for proteinuria (2+) and hematuria (2+). CT (Computed Tomography) thorax showed extensive bilateral airspace disease (dense consolidations and ground-glass opacities) favoring multifocal infection and mediastinal lymphadenopathy(Fig 2). Due to the chronicity of her symptoms and atypical imaging for viral pneumonia, other diagnoses were explored including bacterial superinfection, Tuberculosis, and autoimmune disease. Sputum studies were negative for infections including Acid Fast Bacilli. Workup revealed elevated Antimyeloperoxidase antibodies (MPO) and positive COVID-19 Ig G. She was started on methylprednisolone 1g for AAV. Renal biopsy revealed pauci-immune glomerulonephritis with features of cellular crescent consistent with microscopic polyangiitis (Fig 3). Follow-up CT chest showed improved airspace abnormalities and mediastinal lymphadenopathy. After induction therapy with Rituximab was initiated, she continued to recover and was discharged home. DISCUSSION: The pathogenesis of AAV is believed to be an aberrant pathogenic autoimmune response that follows an initial insult which can include infections. SARS-CoV-2 has been associated with the emergence of autoimmune diseases in susceptible patients(1). The proposed mechanism is linked to elevated levels of circulating neutrophil extracellular traps (NETs) observed in covid infection. These NETS are covered with proteins including neutrophilic enzymes which can activate complement pathways causing tissue destruction and vasculitis. The diagnosis of new-onset AAV can be challenging in COVID-19 patients as symptoms and clinical manifestations of both diseases can overlap. AAV should be considered strongly in patients who are currently infected or have been infected with SARS-CoV-2 and present with atypical or non-resolving pneumonia and other organ involvement such as AKI to avoid permanent organ damage. CONCLUSIONS: The presence of non-resolving or atypical pneumonia and AKI in a patient with SARS-CoV-2 should prompt evaluation of immunological markers to assess or rule out AAV for early diagnosis and treatment. Reference #1: Caso F, Costa L, Ruscitti P, Navarini L, Del Puente A, Giacomelli R, Scarpa R. Could Sars-coronavirus-2 trigger autoimmune and/or autoinflammatory mechanisms in genetically predisposed subjects? Autoimmun Rev. 2020;19(5):102524. doi: 10.1016/j.autrev.2020.102524 DISCLOSURES: No relevant relationships by Tarik Al-Bermani No relevant relationships by Anant Jain No relevant relationships by Ian Kaplan No relevant relationships by Alina Kifayat No relevant relationships by Lisa Paul

ATYPICAL VARIANT OF GRANULOMATOSIS WITH POLYANGIITIS WITH EOSINOPHILIA

SESSION TITLE: Critical Systemic Disease Case Report Posters SESSION TYPE: Case Report Posters PRESENTED ON: 10/19/2022 12:45 pm - 01:45 pm INTRODUCTION: Granulomatosis with polyangiitis (GPA) is a necrotizing granulomatous vasculitis affecting small-to-medium sized blood vessels. GPA is highly associated with antineutrophil cytoplasmic antibodies (ANCAs) and often triggered by environmental factors such as medications and infectious agents. Tracheobronchial stenosis and diffuse alveolar hemorrhage are serious complications of GPA. CASE PRESENTATION: A 35-year-old Caucasian male with a history of chronic sinusitis requiring balloon sinuplasty and recent tympanostomy had presented multiple times to the emergency room due to dyspnea and cough with pinkish sputum production. This was associated with sore throat and fever, which were attributed to his COVID-19 infection and treated with supportive care. Due to persistent drainage through his tympanostomy he was prescribed levofloxacin by his ENT specialist. After the second dose of levofloxacin, he developed Raynaud's phenomenon, diffuse purpuric lesions and swelling over his lower extremity, eyelids, and elbows. Four days later he developed worsening hemoptysis and dyspnea for which he was admitted for further evaluation. Laboratory findings were remarkable for peripheral eosinophilia, elevated ESR 19mm/hr, CRP 9.2mg/dl, c-ANCA 1:320 titer, positive proteinase-3 antibodies and normal p-ANCA titers. Urinalysis with microscopic hematuria. Chest CT scan showed ground glass opacity, consolidative infiltrate with subpleural sparing and minimal left bronchial stenosis. Bronchoscopy suggestive of diffuse alveolar hemorrhage. Limited lung biopsy showed ulcer and granulation tissue with abundant eosinophils, but no granulomatous inflammation noted. Pulse dose steroids and Rituximab were initiated, and rapid clinical improvement was noted. Patient was discharged on prednisone taper and Pneumocystis jiroveci prophylaxis. DISCUSSION: We believe that GPA may have been triggered by recent COVID-19 infection and levofloxacin use. Mild peripheral blood and tissue eosinophilia (<12%) has been described in GPA, however it is a rare finding. GPA and eosinophilic granulomatosis with polyangiitis (EGPA) are both ANCA vasculitis that involve lungs and kidneys. GPA presents with sinusitis, alveolar hemorrhage and high titers of PR-3 antibodies. EGPA presents with a history of atopic, asthma and high titers of myeloperoxidase-ANCA along with abundant peripheral eosinophils. Our patient best fits the diagnostic criteria for GPA with eosinophilia variant rather than EGPA. Our patient had no history of asthma or atopic disease and p-ANCA was normal, which also points away from EGPA. CONCLUSIONS: Clinicians should recognize the differential diagnosis for eosinophils in ANCA vasculitis. Early diagnosis of ANCA vasculitis and initiation of appropriate treatment is important to decrease morbidity and mortality. Reference #1: Potter MB, Fincher RK, Finger DR. Eosinophilia in Wegener's Granulomatosis. Chest 116: 1480-1483, 1999 Reference #2: Krupsky, Meir et al. Wegener's Granulomatosis With Peripheral Eosinophilia. CHEST, Volume 104, Issue 4, 1290 - 1292 Reference #3: Kitching AR, Anders HJ, et al. ANCA-associated vasculitis. Nat Rev Dis Primers. 2020 Aug 27;6(1):71. doi: 10.1038/s41572-020-0204-y. PMID: 32855422. DISCLOSURES: No relevant relationships by Afoma King No relevant relationships by Joshuam Ruiz Vega No relevant relationships by Krishna Shah no disclosure on file for Milos Tucakovic;

A RARE CASE OF ANTINEUTROPHIL CYTOPLASMIC ANTIBODIES (ANCA)-ASSOCIATED VASCULITIS IN THE SETTING OF COVID-19 INFECTION

SESSION TITLE: Management of COVID-19-Induced Complications SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: Anti-neutrophil cytoplasmic antibody-associated vasculitis (ANCA-AV) is an autoimmune mediated inflammation of small and medium sized vessel walls. The occurrence of this autoimmune vasculitis is typically associated with underlying infection, medications, and genetic predisposition.(1) The objective of this case report is to describe a rare presentation of ANCA-AV in the setting of COVID-19 infection. CASE PRESENTATION: A 67-year-old male presented to the hospital with a three-week history of cough productive of brown sputum, epistaxis, fatigue, decreased appetite, and unintentional weight loss. During the previous week, he experienced worsening dyspnea and bilateral lower extremity swelling. On physical examination, he was hypoxic requiring 4L of supplemental oxygen to maintain saturations greater than 90%. Diffuse and bilateral wheezes were heard on auscultation of his lungs. A tender petechial rash was dispersed over his limbs, trunk, oropharynx, and nasopharynx. A basic metabolic panel revealed a mild, acute renal impairment. Urinalysis showed new onset proteinuria and hemoglobinuria. Nasopharyngeal swab was positive for SARS-COV-2. Contrast-enhanced computed tomography of the chest revealed diffuse, bilateral ground glass opacities and interstitial changes. Therapy with piperacillin-tazobactam was started for presumed superimposed bacterial community acquired pneumonia in the setting of COVID-19 infection. On day three of hospitalization, the petechial rash progressed to hemorrhagic blisters. His oral petechiae were now ulcerated. A punch biopsy of the affected skin showed leukocytoclastic vasculitis. Anti-Proteinase 3 (PR3) antibodies were positive. Subsequent renal biopsy showed pauci-immune focal necrotizing crescentic glomerulonephritis consistent with ANCA-AV. Therapy with intravenous pulse dose corticosteroids led to improvement in his rash and body aches, and he was discharged home on oral steroids ten days after admission. DISCUSSION: This report describes a rare case of ANCA-AV in the setting of recent COVID-19 infection. Differentiation of ANCA-AV, bacterial and COVID-19 pneumonia can be challenging on chest imaging alone.(1) New onset renal impairment, hematuria, proteinuria and the presence of the petechial rash were suspicious for co-existing ANCA-AV in this patient. COVID-19- associated cytokine storm and formation of neutrophil extracellular traps (NETs) is postulated to be the underlying cause.(1-3) NETs present myeloperoxidase (MPO) and PR3 antigens to the immune system. Formation of auto-antibodies to MPO and PR3 lead to the development of ANCA-AV. The findings of NETs on kidney biopsy specimens in patients with ANCA-AV supports this hypothesis.(1,2) CONCLUSIONS: To avoid the misdiagnosis of COVID-19-induced vasculitis, a low threshold to investigate co-existing vasculitis in patients with COVID-19 and associated clinical findings is highly recommended. Reference #1: Izci Duran T, Turkmen E, Dilek M, Sayarlioglu H, Arik N. ANCA-associated vasculitis after COVID-19. Rheumatol Int. 2021;41(8):1523-1529. Reference #2: Uppal NN, Kello N, Shah HH, et al. De Novo ANCA-Associated Vasculitis With Glomerulonephritis in COVID-19. Kidney Int Rep. 2020;5(11):2079-2083. Reference #3: Cobilinschi C, Cobilinschi C, Constantinescu A, Draniceanu I, Ionescu R. New-Onset ANCA-Associated Vasculitis in a Patient with SARS-COV2. Balkan Med J. 2021;38(5):318-320. DISCLOSURES: No relevant relationships by Andrei Hastings No relevant relationships by Jason Lane No relevant relationships by Tanya Marshall No relevant relationships by Palak Rath No relevant relationships by Sterling Shriber No relevant relationships by inderprit Singh No relevant relationships by Samuel Wiles

Rapid Colorimetric Detection of Wound Infection with a Fluidic Paper Device.

Current procedures for the assessment of chronic wound infection are time-consuming and require complex instruments and trained personnel. The incidence of chronic wounds worldwide, and the associated economic burden, urge for simple and cheap point-of-care testing (PoCT) devices for fast on-site diagnosis to enable appropriate early treatment. The enzyme myeloperoxidase (MPO), whose activity in infected wounds is about ten times higher than in non-infected wounds, appears to be a suitable biomarker for wound infection diagnosis. Herein, we develop a single-component foldable paper-based device for the detection of MPO in wound fluids. The analyte detection is achieved in two steps: (i) selective immunocapture of MPO, and (ii) reaction of a specific dye with the captured MPO, yielding a purple color with increasing intensity as a function of the MPO activity in infected wounds in the range of 20-85 U/mL. Ex vivo experiments with wound fluids validated the analytic efficiency of the paper-based device, and the results strongly correlate with a spectrophotometric assay.

Pathogenetic and prognostic significance of inflammation and altered ADAMTS-13/vWF axis in patients with severe COVID-19

Introduction. Currently, endothelial dysfunction caused by inflammation and immunothrombosisis considered as one of the crucial mechanisms in developing the SARS-CoV-2 virus-mediated coronavirus disease 2019 (COVID-19). A mass endothelial damage followed by release of untypical large quantity of von Willebrand factor (vWF) multimers and subsequent consumption of metalloproteinase ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13) is described during severe COVID-19. The activation of innate immune cells including neutrophils results in formation of neutrophil extracellular traps (NETs) and myeloperoxidase (MPO) release that, in turn, contributes to spread of inflammation and microvascular thrombosis. Aim: to evaluate a pathogenetic role and predictive significance for serum markers of inflammation, endothelial dysfunction and hemostatis activation such as vWF, ADAMTS-13 and MPO for in-hospital mortality in severe COVID-19 patients requiring mechanical lung ventilation. Materials and Methods. There was performed a single-center observational study with 129 severe COVID-19 patients on mechanical lung ventilation at the intensive care unit, by assessing serum in all subjects vWF, ADAMTS-13 as well as in 79 patients MPO level along with other potential predictors for in-hospital mortality. Results. A multivariate analysis revealed that increased serum level for vWF antigen (vWF:Ag) and MPO antigen (MPO:Ag) were significantly and independently related to high mortality probability: vWF:Ag (IU/ml) - adjusted odds ratio (OR) = 3.360;95 % confidence interval (95 % Cl) = 1.562-7,228 (р = 0,0019);MPO:Ag (ng/ml) - adjusted OR = 1.062;95 % = 1.024-1.101 (p = 0.0011). Such data allowed to obtained a simplified mortality score for categorizing patients as those having a higher or lower score compared with the median score level: a high score was associated with lower cumulative survival rate (p < 0.0001), with 50 % of the cases linked to lethal outcome on day 13 post-hospital admission. Conclusion. Severe COVID-19 patients requiring mechanical lung ventilation were found to have elevated level of serum MPO activity and vWF correlating with poor survival.

ENDOGENOUS REGULATION OF NEUTROPHIL EXTRACELLULAR TRAPS IN ACUTE ISCHAEMIC STROKE DUE TO VACCINE-INDUCED IMMUNE THROMBOTIC THROMBOCYTOPENIA

Background and aims: Vaccination is one of the most important countermeasures in the ongoing COVID19-pandemic. Pharmacovigilance concerns, however, emerged after very rare, but potentially disastrous thrombotic complications following vaccination with ChAdOx1. The underlying pathology was described as platelet factor 4 antibody mediated vaccine-induced immune thrombotic thrombocytopenia (VITT). Mechanisms of immunothrombosis including the regulation of neutrophil extracellular traps (NETs) might be of crucial importance in VITT. Methods: In this study, we had the exceptional opportunity to investigate blood and thrombus samples of a patient who suffered severe ischaemic stroke due to VITT undergoing successful mechanical thrombectomy (MT) in comparison to 13 control stroke patients with similar clinical characteristics without VITT. Cerebral thrombi were stained for complement factors, NETs-markers, DNase and LL-37 and were histologically assessed. In blood samples before MT and 7 days later, cell-free DNA, myeloperoxidase-histone complexes, myeloperoxidase activity, DNase activity, LL-37 and inflammatory cytokines were determined. Results: We identified NETs-markers in thrombi of all patients. The thrombus of the VITT-patient, however, exclusively revealed complement factors and high amounts of NETs-degrading DNase and LL-37, which protects NETs from degradation. In accordance, ex vivo and in vitro studies also implied a disturbed regulation of NETs in VITT-samples. Moreover, we found markedly pronounced temporal changes of specific circulating cytokines in the VITT-case. Conclusions: The results of this study suggest a disturbed endogenous regulation of NETs in VITT involving a pro-inflammatory response and fulminant clinical course. Further studies are warranted to confirm our results and to provide deeper insights into the causative mechanisms.

DE NOVO DOUBLE GLOMERULOPATHY (MEMBRANOUS NEPHROPATHY, MN AND COLLAPSING FOCAL SEGMENTAL GLOMERULOSCLEROSIS, CFSGS) ASSOCIATED TO POSITIVE MYELOPEROXIDASE-O (MPO) ANTIBODY FOLLOWING PFIZER - BIONTECH MRNA VACCINATION COVID 19

BACKGROUND AND AIMS: In the COVID 19 pandemic era, anti SARS-CoV-2 vaccination showed high efficacy at preventing the infection and its most severe complications. The aim of this report is to describe an unusual double glomerulopathy related to anti SARS-CoV-2 vaccination and the good results obtained with the immunosoppressive treatment. METHOD: An 80-year-old caucasian woman developed a nephrotic syndrome, progressive renal insufficiency and microhematuria. The patient presented a medical history of thrombocytopenic purpura treated and resolved by steroids in 2013, hypothyroidism, hypertension, ischaemic heart disease treated with surgical bypass in 2019 and pacemaker in 2020 for atrial ventricular block. Due to pandemic COVID 19 status, she received two doses of the Pfizer BioNTech mRNA COVID-19 vaccine in March 2021. Two weeks after the second dose her weight increased of 23 kg. The family physician added furosemide to her therapy for generalized edema with no diuretic effect. In April, creatinine was 1.38 mg/dL (versus 0.8 mg/dL 1 year before);urinalysis showed proteinuria (300 mg/dL) and microscopic hematuria;serum total cholesterol level was 218 mg/dL and triglycerides 178 mg/dL;then it was suggested to increase the doses of furosemide. In May 2021, creatinine resulted 2 mg/dL, serum albumin 2 g/dL, and urinalysis confirmed proteinuria and microscopic hematuria;proteinuria was 10 g/day. Abdomen ultrasound showed normal liver, kidneys and spleen, not ascites. Lower limb eco-Doppler showed right superficial femoral artery stenosis of 60% and absence of venous thrombosis. The physical examination evidenced anasarca. The patients were admitted to the nephrology unit;hepatitis B surface antigen, hepatitis C antibody and human immunodeficiency virus antigen and antibody were negative. Both complement C3 and C4 levels resulted within the normal range. Cryoglobulins were absent. Urinary Bence Jones, antinuclear antibody (ANA), anti-extractable nuclear antigen (ENA), anti-double stranded DNA (nDNA) antibodies were negative. Antineutrophil cytoplasm antibodies (ANCA) were 1:2560 with Perinuclear pattern and anti-MPO positivity (716 UA/mL);anti-proteinase-3 antibodies (PR3) were negative. Antiphosholipase A2 receptor antibody (PLA2R Ab) was positive with high titre. A kidney biopsy was performed showing a double nephropathy: a focal segmental glomerulosclerosis (FSGS) with some collapsing features, superimposed on membranous glomerulonephritis (Fig. 1). RESULTS: We started the Ponticelli regimen (alternate months steroids and cyclophosphamide). After the first month of therapy, blood tests revealed creatinine 1.7 mg/dL, haemoglobin 11.7 g/dL;serum albumin 2.7 g/dL and urinalysis without microscopic haematuria. At the third month of therapy, the patient developed atrial fibrillation and started anticoagulation;blood tests were as follows: creatinine 1.1 mg/dL, serum albumin 3.0 g/dL, Ab anti-MPO 7 UA/mL and PLA2R Ab was absent. A left ocular, frontal and parietal herpes zoster induced a short discontinuation of therapy and responded well to Acyclovir;then we concluded the fourth month of therapy. At the fifth month, a SARS CoV 2 RT PCR unexpectedly resulted positive;the patient remained asymptomatic, but we stopped definitively the therapy. One month later, blood tests showed: creatinine 1 mg/dL, serum albumin 4 g/dL, proteinuria 0.7 g/die, MPO 2 UA/mL and PLA2R Ab absent. CONCLUSION: To our knowledge, this is the first case of nephrotic sindrome secondary to a De novo MN and FSGS, associated with positive MPO antibody, following Pfizer-BioNTech mRNA vaccination COVID 19;the patient responded well to immunosoppression going in remission and regaining renal function. (Figure Presented).