Escalated complement activation during hospitalization is associated with higher risk of 60-day mortality in SARS-CoV-2-infected patients.

BACKGROUND: The complement system, an upstream recognition system of innate immunity, is activated upon SARS-CoV-2 infection. To gain a deeper understanding of the extent and duration of this activation, we investigated complement activation profiles during the acute phase of COVID-19, its persistence post-recovery and dynamic changes in relation to disease severity. METHODS: Serial blood samples were obtained from two cohorts of hospitalized COVID-19 patients (n = 457). Systemic complement activation products reflecting classical/lectin (C4d), alternative (C3bBbP), common (C3bc) and terminal pathway (TCC and C5a) were measured during hospitalization (admission, days 3-5 and days 7-10), at 3 months and after 1 year. Levels of activation and temporal profiles during hospitalization were related to disease severity defined as respiratory failure (PO2/FiO2 ratio <26.6 kPa) and/or admission to intensive care unit, 60-day total mortality and pulmonary pathology after 3 months. FINDINGS: During hospitalization, TCC, C4d, C3bc, C3bBbP and C5a were significantly elevated compared to healthy controls. Severely ill patients had significantly higher levels of TCC and C4d (p < 0.001), compared to patients with moderate COVID-19. Escalated levels of TCC and C4d during hospitalization were associated with a higher risk of 60-day mortality (p < 0.001), and C4d levels were additionally associated with chest CT changes at 3 months (p < 0.001). At 3 months and 1 year, we observed consistently elevated levels of most complement activation products compared to controls. CONCLUSION: Hospitalized COVID-19 patients display prominent and long-lasting systemic complement activation. Optimal targeting of the system may be achieved through enhanced risk stratification and closer monitoring of in-hospital changes of complement activation products.

High viral loads combined with inflammatory markers predict disease severity in hospitalized COVID-19 patients: Results from the NOR-Solidarity trial.

OBJECTIVES: To investigate temporal changes in the association between SARS-CoV2 viral load (VL) and markers of inflammation during hospitalization, as well as the ability of these markers alone or in combination to predict severe outcomes. METHODS: Serial oropharyngeal and blood samples were obtained from hospitalized COVID-19 patients (n = 160). Levels of inflammatory markers and oropharyngeal VL were measured during hospitalization (admission, days 3-5, and days 7-10) and related to severe outcomes (respiratory failure/intensive care unit admission). RESULTS: Elevated admission levels of IL (interleukin)-6, IL-33, IL-8, monocyte chemoattractant protein-1 (MCP-1), interferon-γ-induced protein 10 (IP-10), IL-1ß, and IL-1Ra were associated with severe outcomes during hospitalization. Although no inflammatory markers correlated with VL at baseline, there was a significant correlation between VL and levels of IP-10 and MCP-1 at days 3-5, accompanied by IL-8 and IL-6 at days 7-10. Finally, there was a seemingly additive effect of IP-10, MCP-1, and IL-6 in predicting severe outcomes when combined with high VL at baseline. CONCLUSIONS: An increasing number of inflammatory markers were associated with VL during the first 10 days of hospitalization, and several of these markers were associated with severe outcomes, in particular when combined with elevated VL. Future studies should assess the potential for combining antiviral and immunomodulatory treatment, preferably guided by viral and inflammatory biomarkers, for the selection of high-risk patients.

Changes in cardiopulmonary exercise capacity and limitations 3-12 months after COVID-19.

RATIONALE: To describe cardiopulmonary function during exercise 12 months after hospital discharge for coronavirus disease 2019 (COVID-19), assess the change from 3 to 12 months, and compare the results with matched controls without COVID-19. METHODS: In this prospective, longitudinal, multicentre cohort study, hospitalised COVID-19 patients were examined using a cardiopulmonary exercise test (CPET) 3 and 12 months after discharge. At 3 months, 180 performed a successful CPET, and 177 did so at 12 months (mean age 59.3 years, 85 females). The COVID-19 patients were compared with controls without COVID-19 matched for age, sex, body mass index and comorbidity. Main outcome was peak oxygen uptake (V'O2  peak). RESULTS: Exercise intolerance (V'O2  peak <80% predicted) was observed in 23% of patients at 12 months, related to circulatory (28%), ventilatory (17%) and other limitations including deconditioning and dysfunctional breathing (55%). Estimated mean difference between 3 and 12 months showed significant increases in V'O2  peak % pred (5.0 percentage points (pp), 95% CI 3.1-6.9 pp; p<0.001), V'O2  peak·kg-1 % pred (3.4 pp, 95% CI 1.6-5.1 pp; p<0.001) and oxygen pulse % pred (4.6 pp, 95% CI 2.5-6.8 pp; p<0.001). V'O2  peak was 2440 mL·min-1 in COVID-19 patients compared to 2972 mL·min-1 in matched controls. CONCLUSIONS: 1 year after hospital discharge for COVID-19, the majority (77%), had normal exercise capacity. Only every fourth had exercise intolerance and in these circulatory limiting factors were more common than ventilator factors. Deconditioning was common. V'O2  peak and oxygen pulse improved significantly from 3 months.

Circulating markers of extracellular matrix remodelling in severe COVID-19 patients.

BACKGROUND: Abnormal remodelling of the extracellular matrix (ECM) has generally been linked to pulmonary inflammation and fibrosis and may also play a role in the pathogenesis of severe COVID-19. To further elucidate the role of ECM remodelling and excessive fibrogenesis in severe COVID-19, we examined circulating levels of mediators involved in various aspects of these processes in COVID-19 patients. METHODS: Serial blood samples were obtained from two cohorts of hospitalised COVID-19 patients (n = 414). Circulating levels of ECM remodelling mediators were quantified by enzyme immunoassays in samples collected during hospitalisation and at 3-month follow-up. Samples were related to disease severity (respiratory failure and/or treatment at the intensive care unit), 60-day total mortality and pulmonary pathology after 3-months. We also evaluated the direct effect of inactivated SARS-CoV-2 on the release of the different ECM mediators in relevant cell lines. RESULTS: Several of the measured markers were associated with adverse outcomes, notably osteopontin (OPN), S100 calcium-binding protein A12 and YKL-40 were associated with disease severity and mortality. High levels of ECM mediators during hospitalisation were associated with computed tomography thorax pathology after 3-months. Some markers (i.e. growth differential factor 15, galectin 3 and matrix metalloproteinase 9) were released from various relevant cell lines (i.e. macrophages and lung cell lines) in vitro after exposure to inactivated SARS-CoV-2 suggesting a direct link between these mediators and the causal agent of COVID-19. CONCLUSION: Our findings highlight changes to ECM remodelling and particularly a possible role of OPN, S100A12 and YKL-40 in the pathogenesis of severe COVID-19.

High Circulating Levels of the Homeostatic Chemokines CCL19 and CCL21 Predict Mortality and Disease Severity in COVID-19.

BACKGROUND: Immune dysregulation is a major factor in the development of severe coronavirus disease 2019 (COVID-19). The homeostatic chemokines CCL19 and CCL21 have been implicated as mediators of tissue inflammation, but data on their regulation in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is limited. We thus investigated the levels of these chemokines in COVID-19 patients. METHODS: Serial blood samples were obtained from patients hospitalized with COVID-19 (n = 414). Circulating CCL19 and CCL21 levels during hospitalization and 3-month follow-up were analyzed. In vitro assays and analysis of RNAseq data from public repositories were performed to further explore possible regulatory mechanisms. RESULTS: A consistent increase in circulating levels of CCL19 and CCL21 was observed, with high levels correlating with disease severity measures, including respiratory failure, need for intensive care, and 60-day all-cause mortality. High levels of CCL21 at admission were associated with persisting impairment of pulmonary function at the 3-month follow-up. CONCLUSIONS: Our findings highlight CCL19 and CCL21 as markers of immune dysregulation in COVID-19. This may reflect aberrant regulation triggered by tissue inflammation, as observed in other chronic inflammatory and autoimmune conditions. Determination of the source and regulation of these chemokines and their effects on lung tissue is warranted to further clarify their role in COVID-19. CLINICAL TRIALS REGISTRATION: NCT04321616 and NCT04381819.

Respiratory dysfunction three months after severe COVID-19 is associated with gut microbiota alterations.

BACKGROUND: Although coronavirus disease 2019 (COVID-19) is primarily a respiratory infection, mounting evidence suggests that the gastrointestinal tract is involved in the disease, with gut barrier dysfunction and gut microbiota alterations being related to disease severity. Whether these alterations persist and are related to long-term respiratory dysfunction remains unknown. METHODS: Plasma was collected during hospital admission and after 3 months from the NOR-Solidarity trial (n = 181) and analyzed for markers of gut barrier dysfunction and inflammation. At the 3-month follow-up, pulmonary function was assessed by measuring the diffusing capacity of the lungs for carbon monoxide (DLCO ). Rectal swabs for gut microbiota analyses were collected (n = 97) and analyzed by sequencing the 16S rRNA gene. RESULTS: Gut microbiota diversity was reduced in COVID-19 patients with respiratory dysfunction, defined as DLCO below the lower limit of normal 3 months after hospitalization. These patients also had an altered global gut microbiota composition, with reduced relative abundance of 20 bacterial taxa and increased abundance of five taxa, including Veillonella, potentially linked to fibrosis. During hospitalization, increased plasma levels of lipopolysaccharide-binding protein (LBP) were strongly associated with respiratory failure, defined as pO2 /fiO2 (P/F ratio) <26.6 kPa. LBP levels remained elevated during and after hospitalization and were associated with low-grade inflammation and respiratory dysfunction after 3 months. CONCLUSION: Respiratory dysfunction after COVID-19 is associated with altered gut microbiota and persistently elevated LBP levels. Our results should be regarded as hypothesis generating, pointing to a potential gut-lung axis that should be further investigated in relation to long-term pulmonary dysfunction and long COVID.

Anti-PF4/polyanion antibodies in COVID-19 patients are associated with disease severity and pulmonary pathology.

Thromboembolic events are frequent and associated with poor outcome in severe COVID-19 disease. Anti-PF4/polyanion antibodies are related to heparin-induced thrombocytopenia (HIT) and thrombus formation, but data on these antibodies in unselected COVID-19 populations are scarce. We assessed the presence of anti-PF4/polyanion antibodies in prospectively collected serum from an unselected cohort of hospitalized COVID-19 patients and evaluated if elevated levels could give prognostic information on ICU admission and respiratory failure (RF), were associated with markers of inflammation, endothelial activation, platelet activation, coagulation and fibrosis and were associated with long-term pulmonary CT changes. Five out of 65 patients had anti-PF4/polyanion reactivity with OD ≥0.200. These patients had more severe disease as reflected by ICU admission without any evidence of HIT. They also had signs of enhanced inflammation and fibrinogenesis as reflected by elevated ferritin and osteopontin, respectively, during the first 10 days of hospitalization. Increased ferritin and osteopontin persisted in these patients at 3 months follow-up, concomitant with pulmonary CT pathology. Our finding shows that the presence of anti-PF4/polyanion antibodies in unselected hospitalized COVID-19 patients was not related to HIT, but was associated with disease severity, inflammation, and pulmonary pathology after 3 months.

Cardiopulmonary exercise capacity and limitations 3†months after COVID-19 hospitalisation.

BACKGROUND: This study aimed to describe cardiopulmonary function during exercise 3 months after hospital discharge for COVID-19 and compare groups according to dyspnoea and intensive care unit (ICU) stay. METHODS: Participants with COVID-19 discharged from five large Norwegian hospitals were consecutively invited to a multicentre, prospective cohort study. In total, 156 participants (mean age 56.2 years, 60 females) were examined with a cardiopulmonary exercise test (CPET) 3 months after discharge and compared with a reference population. Dyspnoea was assessed using the modified Medical Research Council (mMRC) dyspnoea scale. RESULTS: Peak oxygen uptake (V'O2  peak) <80% predicted was observed in 31% (n=49). Ventilatory efficiency was reduced in 15% (n=24), while breathing reserve <15% was observed in 16% (n=25). Oxygen pulse <80% predicted was found in 18% (n=28). Dyspnoea (mMRC ≥1) was reported by 47% (n=59). These participants had similar V'O2  peak (p=0.10) but lower mean±sd V'O2  peak·kg-1 % predicted compared with participants without dyspnoea (mMRC 0) (76±16% versus 89±18%; p=0.009) due to higher body mass index (p=0.03). For ICU- versus non-ICU-treated participants, mean±sd V'O2  peak % predicted was 82±15% and 90±17% (p=0.004), respectively. Ventilation, breathing reserve and ventilatory efficiency were similar between the ICU and non-ICU groups. CONCLUSIONS: One-third of participants experienced V'O2  peak <80% predicted 3 months after hospital discharge for COVID-19. Dyspnoeic participants were characterised by lower exercise capacity due to obesity and lower ventilatory efficiency. Ventilation and ventilatory efficiency were similar between ICU- and non-ICU-treated participants.

Dyspnoea, lung function and CT findings 3†months after hospital admission for COVID-19.

The long-term pulmonary outcomes of coronavirus disease 2019 (COVID-19) are unknown. We aimed to describe self-reported dyspnoea, quality of life, pulmonary function and chest computed tomography (CT) findings 3 months following hospital admission for COVID-19. We hypothesised outcomes to be inferior for patients admitted to intensive care units (ICUs), compared with non-ICU patients.Discharged COVID-19 patients from six Norwegian hospitals were enrolled consecutively in a prospective cohort study. The current report describes the first 103 participants, including 15 ICU patients. The modified Medical Research Council (mMRC) dyspnoea scale, the EuroQol Group's questionnaire, spirometry, diffusing capacity of the lung for carbon monoxide (D LCO), 6-min walk test, pulse oximetry and low-dose CT scan were performed 3 months after discharge.mMRC score was >0 in 54% and >1 in 19% of the participants. The median (25th-75th percentile) forced vital capacity and forced expiratory volume in 1 s were 94% (76-121%) and 92% (84-106%) of predicted, respectively. D LCO was below the lower limit of normal in 24% of participants. Ground-glass opacities (GGO) with >10% distribution in at least one of four pulmonary zones were present in 25% of participants, while 19% had parenchymal bands on chest CT. ICU survivors had similar dyspnoea scores and pulmonary function as non-ICU patients, but higher prevalence of GGO (adjusted OR 4.2, 95% CI 1.1-15.6) and lower performance in usual activities.3 months after admission for COVID-19, one-fourth of the participants had chest CT opacities and reduced diffusing capacity. Admission to ICU was associated with pathological CT findings. This was not reflected in increased dyspnoea or impaired lung function.

Patients with Cheyne-Stokes respiration and heart failure: patient tolerance after three-month discontinuation of treatment with adaptive servo-ventilation.

The recent SERVE HF study concluded that patients with chronic heart failure (CHF) and Cheyne-Stokes respiration (CSR) have increased mortality when treated with adaptive servo-ventilation (ASV). We, therefore, wanted to explore if these patients tolerated discontinuation of ASV treatment. The study was a prospective post-ASV treatment observational design with a 3-month follow-up period. 14 patients from our outpatient clinic, all male, were originally diagnosed with CHF and Cheyne-Stokes respiration, which is a clinical form of central sleep apnea. Left ventricular ejection fraction (LVEF) was ≤45% when ASV treatment was initiated. Median machine use was 68 (42-78) months when the patients were instructed to terminate ASV treatment. The patients were then followed during conventional CHF treatment for 3 months. Study baseline was set the last ASV treatment day. Sleep data were collected from the machine the last day of use. Apnea-hypopnea index (AHI), LVEF, 6-min walk test and 24-h ambulatory electrocardiogram recordings were performed at baseline and at study end. Life quality data were obtained using The Minnesota Living with Heart Failure Questionaire (MLHFQ). New York Heart Association Functional Classification (NYHA) was registered. An ambulatory sleep screening was performed at study end. AHI increased significantly after 3 months without ASV treatment [from 1.6 (0.8-3.2) to 39.2 (24.3-44.1, p = 0.001)]. Quality of life (QOL) decreased significantly: 30 (13-54) at discontinuation of ASV vs. 46 (24-67) (MLHFQ) at study end, p = 0.04. Though there was no significant change in NYHA functional class, patients especially reported increased shortness of breath, reduced concentration and reduced memory after discontinuation of ASV treatment. There were no significant differences in LVEF, heart rhythm data and physical capacity. Left ventricular function was preserved indicating that discontinuation of ASV in heart failure patients does not affect cardiac capacity. There was a significant decrement in QOL that must be considered in further treatment of these patients.

CT abnormalities 3 and 12 months after hospitalization for COVID-19 and association with disease severity: A prospective cohort study.

OBJECTIVES: To investigate changes in chest CT between 3 and 12 months and associations with disease severity in patients hospitalized for COVID-19 during the first wave in 2020. MATERIALS AND METHODS: Longitudinal cohort study of patients hospitalized for COVID-19 in 2020. Chest CT was performed 3 and 12 months after admission. CT images were evaluated using a CT severity score (CSS) (0-12 scale) and recoded to an abbreviated version (0-3 scale). We analyzed determinants of the abbreviated CSS with multivariable mixed effects ordinal regression. RESULTS: 242 patients completed CT at 3 months, and 124 (mean age 62.3±13.3, 78 men) also at 12 months. Between 3 and 12 months (n = 124) CSS (0-12 scale) for ground-glass opacities (GGO) decreased from median 3 (25th-75th percentile: 0-12) at 3 months to 0.5 (0-12) at 12 months (p<0.001), but increased for parenchymal bands (p<0.001). In multivariable analysis of GGO, the odds ratio for more severe abbreviated CSS (0-3 scale) at 12 months was 0.11 (95%CI 0.11 0.05 to 0.21, p<0.001) compared to 3 months, for WHO severity category 5-7 (high-flow oxygen/non-invasive ventilation/ventilator) versus 3 (non-oxygen use) 37.16 (1.18 to 43.47, p = 0.032), and for age ≥60 compared to <60 years 4.8 (1.33 to 17.6, p = 0.016). Mosaicism was reduced at 12 compared to 3 months, OR 0.33 (95%CI 0.16 to 0.66, p = 0.002). CONCLUSIONS: GGO and mosaicism decreased, while parenchymal bands increased from 3 to 12 months. Persistent GGO were associated with initial COVID-19 severity and age ≥60 years.

A prospective study of pulmonary outcomes and chest computed tomography in the first year after COVID-19.

COVID-19 primarily affects the respiratory system. We aimed to evaluate how pulmonary outcomes develop after COVID-19 by assessing participants from the first pandemic wave prospectively 3 and 12 months following hospital discharge. Pulmonary outcomes included self-reported dyspnoea assessed with the modified Medical Research Council dyspnoea scale, 6-min walk distance (6MWD), spirometry, diffusing capacity of the lung for carbon monoxide (D LCO), body plethysmography and chest computed tomography (CT). Chest CT was repeated at 12 months in participants with pathological findings at 3 months. The World Health Organization (WHO) ordinal scale for clinical improvement defined disease severity in the acute phase. Of 262 included COVID-19 patients, 245 (94%) and 222 (90%) participants attended the 3- and 12-month follow-up, respectively. Self-reported dyspnoea and 6MWD remained unchanged between the two time points, while D LCO and total lung capacity improved (0.28 mmol·min-1·kPa-1, 95% CI 0.12-0.44, and 0.13 L, 95% CI 0.02-0.24, respectively). The prevalence of fibrotic-like findings on chest CT at 3 and 12 months in those with follow-up chest CT was unaltered. Those with more severe disease had worse dyspnoea, D LCO and total lung capacity values than those with mild disease. There was an overall positive development of pulmonary outcomes from 3 to 12 months after hospital discharge. The discrepancy between the unaltered prevalence of self-reported dyspnoea and the improvement in pulmonary function underscores the complexity of dyspnoea as a prominent factor of long-COVID. The lack of increase in fibrotic-like findings from 3 to 12 months suggests that SARS-CoV-2 does not induce a progressive fibrotic process in the lungs.

The course and determinants of post-traumatic stress over 12 months after hospitalization for COVID-19.

Objective: To assess the trajectory of symptoms and symptom-defined post-traumatic stress disorder (PTSD) from 1.5 to 12 months after hospitalization for COVID-19 and determine risk factors for persistent symptoms and PTSD. Methods: This was a prospective cohort study of consecutive patients discharged after hospitalization for COVID-19 before 1 June 2020 in six hospitals in Southern Norway. Symptom-defined PTSD was assessed by the post-traumatic stress disorder (PTSD) checklist for DSM-5 (PCL-5) at 1.5, 3 and/or 12 months after hospitalization, using DSM-5 criteria. Changes in PCL-5 symptom score and the prevalence of PTSD were analyzed with multivariable mixed models. Results: In total, 388 patients were discharged alive, and 251 (65%) participated. Respondents had a mean (SD) age of 58.4 (14.2) years, and 142 (57%) were males. The prevalence of symptom-defined PTSD was 14, 8, and 9% at 1.5, 3, and 12 months, respectively. WHO disease severity for COVID-19 was not associated with PCL-5 scores. Female sex, lower age and non-Norwegian origin were associated with higher PCL-5 scores. The odds ratio (OR) (95%CI) for PTSD was 0.32 (0.12 to 0.83, p = 0.019) at 3 months and 0.38 (0.15 to 0.95, p = 0.039) at 12 months compared to 1.5 months. There was no association between PTSD and WHO severity rating. Conclusions: The level of PTSD symptoms decreased from 1.5 to 3 months after hospitalization, but did not decrease further to 12 months, and there was no association between PTSD symptoms and COVID-19 disease severity.

Markers of cellular senescence is associated with persistent pulmonary pathology after COVID-19 infection.

BACKGROUND: The lungs are the organ most likely to sustain serious injury from coronavirus disease 2019 (COVID-19). However, the mechanisms for long-term complications are not clear. Patients with severe COVID-19 have shorter telomere lengths and higher levels of cellular senescence, and we hypothesized that circulating levels of the telomere-associated senescence markers chitotriosidase, ß-galactosidase, cathelicidin antimicrobial peptide and stathmin 1 (STMN1) were elevated in hospitalized COVID-19 patients compared to controls and could be associated with pulmonary sequelae following hospitalization. METHODS: Ninety-seven hospitalized patients with COVID-19 who underwent assessment for pulmonary sequelae at three-month follow-up were included in the study. ß-Galactosidase and chitotriosidase were analysed by fluorescence; stathmin 1 and cathelicidin antimicrobial peptide were analysed by enzyme immuno-assay in plasma samples from the acute phase and after three-months. In addition, the classical senescence markers cyclin-dependent kinase inhibitor 1A and 2A were analysed by enzyme immuno-assay in peripheral blood mononuclear cell lysate after three months. RESULTS: We found elevated plasma levels of the senescence markers chitotriosidase and stathmin 1 in patients three months after hospitalization with COVID-19, and these markers in addition to protein levels of cyclin-dependent kinase inhibitor 2A in cell lysate, were associated with pulmonary pathology. The elevated levels of these markers seem to reflect both age-dependent (chitotriosidase) and age-independent (stathmin 1, cyclin-dependent kinase inhibitor 2A) processes. CONCLUSIONS: We suggest that accelerated ageing or senescence could be important for long-term pulmonary complications of COVID-19, and our findings may be relevant for future research exploring the pathophysiology and management of these patients.

Persistent pulmonary pathology after COVID-19 is associated with high viral load, weak antibody response, and high levels of matrix metalloproteinase-9.

The association between pulmonary sequelae and markers of disease severity, as well as pro-fibrotic mediators, were studied in 108 patients 3 months after hospital admission for COVID-19. The COPD assessment test (CAT-score), spirometry, diffusion capacity of the lungs (DLCO), and chest-CT were performed at 23 Norwegian hospitals included in the NOR-SOLIDARITY trial, an open-labelled, randomised clinical trial, investigating the efficacy of remdesivir and hydroxychloroquine (HCQ). Thirty-eight percent had a CAT-score ≥ 10. DLCO was below the lower limit of normal in 29.6%. Ground-glass opacities were present in 39.8% on chest-CT, parenchymal bands were found in 41.7%. At admission, low pO2/FiO2 ratio, ICU treatment, high viral load, and low antibody levels, were predictors of a poorer pulmonary outcome after 3 months. High levels of matrix metalloproteinase (MMP)-9 during hospitalisation and at 3 months were associated with persistent CT-findings. Except for a negative effect of remdesivir on CAT-score, we found no effect of remdesivir or HCQ on long-term pulmonary outcomes. Three months after hospital admission for COVID-19, a high prevalence of respiratory symptoms, reduced DLCO, and persistent CT-findings was observed. Low pO2/FiO2 ratio, ICU-admission, high viral load, low antibody levels, and high levels of MMP-9 were associated with a worse pulmonary outcome.