Extracellular Vesicles: New Players in the Mechanisms of Sepsis- and COVID-19-Related Thromboinflammation.

Sepsis and COVID-19 patients often manifest an imbalance in inflammation and coagulation, a complex pathological mechanism also named thromboinflammation, which strongly affects patient prognosis. Extracellular vesicles (EVs) are nanoparticles released by cells into extracellular space that have a relevant role in cell-to-cell communication. Recently, EVs have been shown to act as important players in a variety of pathologies, including cancer and cardiovascular disease. The biological properties of EVs in the mechanisms of thromboinflammation during sepsis and COVID-19 are still only partially known. Herein, we summarize the current experimental evidence on the role of EVs in thromboinflammation, both in bacterial sepsis and in COVID-19. A better understanding of EV involvement in these processes could be useful in describing novel diagnostic and therapeutic applications of EVs in these diseases.

A 4C mortality score based dichotomic rule supports Emergency Department discharge of COVID-19 patients.

BACKGROUND: For COVID-19 patients evaluated in the Emergency Department (ED), decision on hospital admission vs home discharge is challenging. The 4C mortality score (4CMS) is a prognostication tool integrating key demographic/clinical/biochemical data validated for COVID-19 inpatients. We sought to derive and validate a dichotomic rule based on 4CMS identifying patients with mild outcomes, suitable for safe ED discharge. METHODS: Derivation was performed in a prospective cohort of ED patients with suspected COVID-19 from two centers (April 2020). Validation was pursued in a prospective multicenter cohort of ED patients with confirmed COVID-19 from 6 centers (October 2020 to January 2021). Chest x-ray (CXR) images were independently scored. The primary composite outcome was all-cause 30-day mortality or hospital admission. Secondary outcomes were ED re-visit, oxygen therapy and ventilation. RESULTS: In a derivation cohort of 838 ED patients with suspected COVID-19, 4CMS ≤8 was associated with low outpatient mortality (0.4%) and was thus selected as a feasible discharge rule. In a validation cohort of 521 COVID-19 outpatients, the mean age was 51±17 years; 97 (18.6%) patients had ≥1 CXR infiltrate. The 4CMS had an AUC of 0.82 for the primary outcome and 0.93 for mortality, outperforming other scores (CURB-65, qCSI, qSOFA, NEWS) and CXR. In 474 (91%) patients with 4CMS≤8, the mortality rate was 0.2% and the hospital admission rate was 6.8%, versus 12.8% and 36.2% for 4CMS≥9 (P<0.001). CXR did not provide additional discrimination. CONCLUSIONS: COVID-19 outpatients with 4CMS≤8 have mild outcomes and can be safely discharged from the ED. [NCT0462918].

Feasibility of Self-Performed Lung Ultrasound with Remote Teleguidance for Monitoring at Home COVID-19 Patients.

During the COVID-19 pandemic, use of telemedicine with the aim of reducing the rate of viral transmission increased. This proof-of-concept observational study was planned to test the feasibility of a home-based lung ultrasound (LUS) follow-up performed by patients with mild COVID-19 infection on themselves. We enrolled patients presenting to the emergency department with SARS-CoV-2 infection without signs of pneumonia and indication to discharge. Each patient received a brief training on how to perform LUS and a handheld ultrasound probe. Then, patients were contacted on a daily basis, and LUS images were acquired by the patients themselves under "teleguidance" by the investigator. Twenty-one patients were enrolled with a median age of 44 years. All evaluations were of sufficient quality for a follow up. Probability of a better LUS quality was related to higher degree (odds ratio, OR, 1.42, 95% CI 0.5-3.99) and a lower quality to evaluation time (from 0.71, 95% CI 0.55-0.92 for less than 7 min, to 0.52, 95% CI 0.38-0.7, between 7 and 10 min, and to 0.29, 95% CI 0.2-0.43, for evaluations longer than 10 min). No effect related to gender or age was detected. LUS performed by patients and remotely overseen by expert providers seems to be a feasible and reliable telemedicine tool.

Thrombopoietin participates in platelet activation in COVID-19 patients.

BACKGROUND: The pathogenesis of coronavirus disease 2019 (COVID-19) is characterized by enhanced platelet activation and diffuse hemostatic alterations, which may contribute to immunothrombosis/thromboinflammation and subsequent development of target-organ damage. Thrombopoietin (THPO), a growth factor essential to megakariocyte proliferation, is known to prime platelet activation and leukocyte-platelet interaction. In addition, THPO concentrations increase in several critical diseases, such as acute cardiac ischemia and sepsis, thus representing a potential diagnostic and prognostic biomarker. Furthermore, several data suggest that interleukin (IL)-6 is one of the most important inflammatory mediators involved in these phenomena, which led to explore the potential therapeutic role of IL-6 inhibitors. In this prospective cohort study, we aimed to study THPO and IL-6 concentrations in COVID-19 patients at the time of first clinical evaluation in the Emergency Department (ED), and to investigate their potential use as diagnostic and prognostic biomarkers. In addition, we sought to explore the role of THPO contained in plasma samples obtained from COVID-19 patients in priming in vitro platelet activation and leukocyte-platelet interaction. METHODS: We enrolled 66 patients presenting to the ED with symptoms suggestive of COVID-19, including 47 with confirmed COVID-19 and 19 in whom COVID-19 was excluded (Non-COVID-19 patients). As controls, we also recruited 18 healthy subjects. In vitro, we reproduced the effects of increased circulating THPO on platelet function by adding plasma from COVID-19 patients or controls to platelet-rich plasma or whole blood obtained by healthy donors, and we indirectly studied the effect of THPO on platelet activation by blocking its biological activity. FINDINGS: THPO levels were higher in COVID-19 patients than in both Non-COVID-19 patients and healthy subjects. Studying THPO as diagnostic marker for the diagnosis of COVID-19 by receiver-operating-characteristic (ROC) statistics, we found an area under the curve (AUC) of 0.73, with an optimal cut-off value of 42.60 pg/mL. IL-6 was higher in COVID-19 patients than in healthy subjects, but did not differ between COVID-19 and Non-COVID-19 patients. THPO concentrations measured at the time of diagnosis in the ED were also higher in COVID-19 patients subsequently developing a severe disease than in those with mild disease. Evaluating THPO as biomarker for severe COVID-19 using ROC analysis, we found an AUC of 0.71, with an optimal cut-off value of 57.11 pg/mL. IL-6 was also higher in severe than in mild COVID-19 patients, with an AUC for severe COVID-19 of 0.83 and an optimal cut-off value of 23 pg/ml. THPO concentrations correlated with those of IL-6 (r=0.2963; p=0.043), and decreased 24 h after the administration of tocilizumab, an IL-6 receptor blocking antibody, showing that the increase of THPO levels depends on IL-6-stimulated hepatic synthesis. In vitro, plasma obtained from COVID-19 patients, but not from healthy subjects, primed platelet aggregation and leukocyte-platelet binding, and these effects were reduced by inhibiting THPO activity. INTERPRETATION: Increased THPO may be proposed as an early biomarker for the diagnosis of COVID-19 and for the identification of patients at risk of developing critical illness. Elevated THPO may contribute to enhance platelet activation and leukocyte-platelet interaction in COVID-19 patients, thus potentially participating in immunothrombosis/thromboinflammation. FUNDING: This work was supported by Ministero dell'Università e della Ricerca Scientifica e Tecnologica (MURST) ex 60% to GM and EL.

A phase 2 randomized, double-blinded, placebo-controlled, multicenter trial evaluating the efficacy and safety of raloxifene for patients with mild to moderate COVID-19.

Background: Current available therapeutic options for Coronavirus Disease-2019 (COVID-19) are primarily focused on treating hospitalized patients, and there is a lack of oral therapeutic options to treat mild to moderate outpatient COVID-19 and prevent clinical progression. Raloxifene was found as a promising molecule to treat COVID-19 due to its activity to modulate the replication of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and act as an immunomodulator to decrease proinflammatory cytokines. Methods: This was a phase 2 multicenter, randomized, placebo-controlled trial to evaluate the efficacy and safety of raloxifene in adult patients with mild to moderate COVID-19 between October 2020 to June 2021 in five centers located in Italy. This was a planned 2/3 adaptive study, but due to operational difficulties, the study was discontinued during the phase 2 study segment. Participants were randomized 1:1:1 to receive oral placebo, raloxifene 60 mg, or raloxifene 120 mg by self-administration for a maximum of two weeks. The primary outcomes were the proportion of patients with undetectable SARS-CoV-2 via nasopharyngeal swabs at day 7 and the proportion of patients who did not require supplemental oxygen therapy or mechanical ventilation on day 14. Safety was assessed. The trial is registered (EudraCT 2021-002,476-39, and ClinicalTrials.gov: NCT05172050). Findings: A total of 68 participants were enrolled and randomized to placebo (n = 21), raloxifene 60 mg (n = 24), and raloxifene 120 mg (n = 23). The proportion of participants with undetectable SARS-CoV-2 after seven days of treatment with raloxifene 60 mg [36.8%, 7/19 vs. 0.0%, 0/14] and 120 mg [22.2%, 4/18 vs. 0.0%, 0/14] was better compared to placebo, [risk difference (RD) = 0·37 (95% C.I.:0·09-0·59)] and [RD = 0·22 (95% C.I.: -0·03-0·45)], respectively. There was no evidence of effect for requirement of supplemental oxygen and/or mechanical ventilation with effects for raloxifene 60 mg and raloxifene 120 mg over placebo, [RD = 0·09 (95% C.I.: -0·22-0·37)], and [RD = 0·03 (95% C.I.: -0·28-0·33)], respectively. Raloxifene was well tolerated at both doses, and there was no evidence of any difference in the occurrence of serious adverse events. Interpretation: Raloxifene showed evidence of effect in the primary virologic endpoint in the treatment of early mild to moderate COVID-19 patients shortening the time of viral shedding. The safety profile was consistent with that reported for other indications. Raloxifene may represent a promising pharmacological option to prevent or mitigate COVID-19 disease progression. Funding: The study was funded by Dompé Farmaceutici SpA and supported by the funds from the European Commission - Health and Consumers Directorate General, for the Action under the Emergency Support Instrument- Grant to support clinical testing of repurposed medicines to treat SARS-COV-2 patients (PPPA-ESI-CTRM-2020-SI2.837140), and by the COVID-2020-12,371,675 Ricerca finalizzata and line 1 Ricerca Corrente COVID both funded by Italian Ministry of Health.

Lung Ultrasound Improves Outcome Prediction over Clinical Judgment in COVID-19 Patients Evaluated in the Emergency Department.

In the Emergency Department (ED), the decision to hospitalize or discharge COVID-19 patients is challenging. We assessed the utility of lung ultrasound (LUS), alone or in association with a clinical rule/score. This was a multicenter observational prospective study involving six EDs (NCT046291831). From October 2020 to January 2021, COVID-19 outpatients discharged from the ED based on clinical judgment were subjected to LUS and followed-up at 30 days. The primary clinical outcome was a composite of hospitalization or death. Within 393 COVID-19 patients, 35 (8.9%) reached the primary outcome. For outcome prognostication, LUS had a C-index of 0.76 (95%CI 0.68-0.84) and showed good performance and calibration. LUS-based classification provided significant differences in Kaplan-Meier curves, with a positive LUS leading to a hazard ratio of 4.33 (95%CI 1.95-9.61) for the primary outcome. The sensitivity and specificity of LUS for primary outcome occurrence were 74.3% (95%CI 59.8-88.8) and 74% (95%CI 69.5-78.6), respectively. The integration of LUS with a clinical score further increased sensitivity. In patients with a negative LUS, the primary outcome occurred in nine (3.3%) patients (p < 0.001 vs. unselected). The efficiency for rule-out was 69.7%. In unvaccinated ED patients with COVID-19, LUS improves prognostic stratification over clinical judgment alone and may support standardized disposition decisions.

Diaphragmatic Point-of-Care Ultrasound in COVID-19 Patients in the Emergency Department-A Proof-of-Concept Study.

BACKGROUND: Lung Ultrasound Evaluation (LUS) is usefully applied in the Emergency Department (ED) to patients with suspected or confirmed COVID-19. Diaphragmatic Ultrasound (DUS) may provide additional insight into ventilatory function. This proof-of-concept study aimed to evaluate the feasibility of LUS and DUS in a third level ED during the COVID-19 pandemic. METHODS: Adult patients presenting with COVID-19 symptoms were eligible. After the physical examination, both LUS and DUS (i.e., diaphragmatic motion and thickness) were performed. All patients were followed after 30 days to determine their need for ventilation, admission, and/or a new ED evaluation after discharge. The diagnostic accuracies of diaphragm measurements in assessing the risk of the 30-day outcome were calculated as well as the measurements' usefulness. Bland-Altman plots were used for comparing bedside and off-line diaphragm measurements. RESULTS: 118 patients were enrolled. Median thickness and motion were 1.7 mm (iqr 0.4) and 1.8 cm (iqr 0.7), respectively, with a mean difference of 0.009 mm (95% CI -0.037-0.056 mm) and -0.051 cm (95% CI -0.108-0.006 cm), respectively. The 30-day outcome was associated with an increase in thickness (OR 5.84, 95% CI 0.96-35.4), and a lower motion (OR 0.49, 95% CI 0.2-1.21). CONCLUSION: DUS seemed to be feasible and reliable in the ED in a population of patients presenting with symptoms related to COVID-19 infection.

After the first wave and beyond lockdown: long-lasting changes in emergency department visit number, characteristics, diagnoses, and hospital admissions.

The first wave (FW) of COVID-19 led to a rapid reduction in total emergency department (ED) visits and hospital admissions for other diseases. Whether this represented a transient "lockdown and fear" phenomenon, or a more persisting trend, is unknown. We divided acute from post-wave changes in ED flows, diagnoses, and hospital admissions, in an Italian city experiencing a FW peak followed by nadir. This multicenter, retrospective, cross-sectional study involved five general EDs of a large Italian city (January-August 2020). Percent changes were calculated versus 2019, using four 14-day periods (FW peak, early/mid/late post-wave). ED visits were 147,446 in 2020, versus 214,868 in 2019. During the FW peak, visits were reduced by 66.4% (P < 0.001). The drop was maximum during daytime (69.8%) and for pediatric patients (89.4%). Critical triage codes were unchanged. Reductions were found for all non-COVID-19 diagnoses. Non-COVID-19 hospital admissions were reduced by 39.5% (P < 0.001), involving all conditions except hematologic, metabolic/endocrine, respiratory diseases, and traumas. In the early, mid, and late post-wave periods, visits were reduced by 25.4%, 25.3% and 23.5% (all P < 0.001) respectively. In the late period, reduction was greater for female (27.9%) and pediatric patients (44.6%). Most critical triage codes were unchanged. Oncological, metabolic/endocrine, and hematological diagnoses were unchanged, while other diagnoses had persistent reductions. Non-COVID-19 hospital admissions were reduced by 12.8% (P = 0.001), 6.3% (P = 0.1) and 12.2% (P = 0.001), respectively. Reductions in ED flows, led by non-critical codes, persisted throughout the summer nadir of COVID-19. Hospital admissions for non-COVID-19 diseases had transient changes.

Diagnostic accuracy of physician's gestalt in suspected COVID-19: Prospective bicentric study.

OBJECTIVES: Physicians' gestalt is central in the diagnostic pipeline of suspected COVID-19, due to the absence of a single tool allowing conclusive rule in or rule out. The aim of this study was to estimate the diagnostic test characteristics of physician's gestalt for COVID-19 in the emergency department (ED), based on clinical findings or on a combination of clinical findings and bedside imaging results. METHODS: From April 1 to April 30, 2020, patients with suspected COVID-19 were prospectively enrolled in two EDs. Physicians prospectively dichotomized patients in COVID-19 likely or unlikely twice: after medical evaluation of clinical features (clinical gestalt [CG]) and after evaluation of clinical features and results of lung ultrasound or chest x-ray (clinical and bedside imaging-integrated gestalt [CBIIG]). The final diagnosis was adjudicated after independent review of 30-day follow-up data. RESULTS: Among 838 ED enrolled patients, 193 (23%) were finally diagnosed with COVID-19. The area under the curve (AUC), sensitivity, and specificity of CG and CBIIG for COVID-19 were 80.8% and 91.6% (p < 0.01), 82.9% and 91.4% (p = 0.01), and 78.6% and 91.8% (p < 0.01), respectively. CBIIG had similar AUC and sensitivity to reverse transcription-polymerase chain reaction (RT-PCR) for SARS-CoV-2 on the first nasopharyngeal swab per se (93.5%, p = 0.24; and 87%, p = 0.17, respectively). CBIIG plus RT-PCR had a sensitivity of 98.4% for COVID-19 (p < 0.01 vs. RT-PCR alone) compared to 95.9% for CG plus RT-PCR (p = 0.05). CONCLUSIONS: In suspected COVID-19, CG and CBIIG have fair diagnostic accuracy, in line with physicians' gestalt for other acute conditions. Negative RT-PCR plus low probability based on CBIIG can rule out COVID-19 with a relatively low number of false-negative cases.

Clinical and epidemiological characteristics associated with pneumonia at disease onset in patients admitted for COVID-19 to the Emergency Department of a large Hospital in Piedmont (North-Western Italy).

OBJECTIVES: to explore clinical and epidemiological characteristics associated with an imaging feature of COVID-19 pneumonia at disease onset, in order to identify factors that may be evaluable by general practitioners at patient's home, and which may lead to identify a more severe disease, needing hospitalization. DESIGN: this is a retrospective/prospective observational hospital cohort. SETTING AND PARTICIPANTS: the study population includes all patients consecutively admitted to the emergency department of Città della salute e della scienza University Hospital from 01.03 to 31.05.2020 with a confirmed diagnosis of SARS-CoV-2 infection. MAIN OUTCOME MEASURES: patients were classified in two groups according to the findings of X-ray imaging, lung ultrasound and chest computer tomography, as pneumonia or not pneumonia patients. RESULTS: in multivariable analysis, factors most strongly associated with emergency department admission with pneumonia were age, oxygen saturation <90% (adj OR 4.16 ;95%CI 1.44-12.07), respiratory rate >24 breaths/min (adj OR 6.50; 95%CI 2.36-17.87), fever ≥38° (adj OR 3.05; 95%CI 1.53-6.08) and the presence of gastroenteric symptoms (vomiting and diarrhea). A delay (> 7 days) between the appearance of the initial lung symptoms (cough and dyspnea) and the admission to the emergency department was also related to a higher probability of receiving a positive imaging report (OR 4.99; 95%CI 2,02-12,34). CONCLUSIONS: in order to reorganize the management of COVID-19 patients in Italy, in view of the risk of a second wave of epidemic or of local outbreaks, it would be desirable to relocate the triage, and possibly the patient's care, from hospital to home. In this scenario it is important to identify all symptoms and signs associated with COVID-19 pneumonia that would facilitate the decision-making process of GPs leading to patients hospitalization.

Lung Ultrasonography for the Diagnosis of SARS-CoV-2 Pneumonia in the Emergency Department.

STUDY OBJECTIVE: Accurate diagnostic testing to identify severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is critical. Although highly specific, SARS-CoV-2 reverse transcriptase-polymerase chain reaction (RT-PCR) has been shown in clinical practice to be affected by a noninsignificant proportion of false-negative results. This study seeks to explore whether the integration of lung ultrasonography with clinical evaluation is associated with increased sensitivity for the diagnosis of coronavirus disease 2019 pneumonia, and therefore may facilitate the identification of false-negative SARS-CoV-2 RT-PCR results. METHODS: This prospective cohort study enrolled consecutive adult patients with symptoms potentially related to SARS-CoV-2 infection who were admitted to the emergency department (ED) of an Italian academic hospital. Immediately after the initial assessment, a lung ultrasonographic evaluation was performed and the likelihood of SARS-CoV-2 infection, based on both clinical and lung ultrasonographic findings ("integrated" assessment), was recorded. RT-PCR SARS-CoV-2 detection was subsequently performed. RESULTS: We enrolled 228 patients; 107 (46.9%) had SARS-CoV-2 infection. Sensitivity and negative predictive value of the clinical-lung ultrasonographic integrated assessment were higher than first RT-PCR result (94.4% [95% confidence interval {CI} 88.2% to 97.9%] versus 80.4% [95% CI 71.6% to 87.4%] and 95% [95% CI 89.5% to 98.2%] versus 85.2% [95% CI 78.3% to 90.6%], respectively). Among the 142 patients who initially had negative RT-PCR results, 21 tested positive at a subsequent molecular test performed within 72 hours. All these false-negative cases were correctly identified by the integrated assessment. CONCLUSION: This study suggests that, in patients presenting to the ED with symptoms commonly associated with SARS-CoV-2 infection, the integration of lung ultrasonography with clinical evaluation has high sensitivity and specificity for coronavirus disease 2019 pneumonia and it may help to identify false-negative results occurring with RT-PCR.

Multi-organ point-of-care ultrasound for COVID-19 (PoCUS4COVID): international expert consensus.

COVID-19 has caused great devastation in the past year. Multi-organ point-of-care ultrasound (PoCUS) including lung ultrasound (LUS) and focused cardiac ultrasound (FoCUS) as a clinical adjunct has played a significant role in triaging, diagnosis and medical management of COVID-19 patients. The expert panel from 27 countries and 6 continents with considerable experience of direct application of PoCUS on COVID-19 patients presents evidence-based consensus using GRADE methodology for the quality of evidence and an expedited, modified-Delphi process for the strength of expert consensus. The use of ultrasound is suggested in many clinical situations related to respiratory, cardiovascular and thromboembolic aspects of COVID-19, comparing well with other imaging modalities. The limitations due to insufficient data are highlighted as opportunities for future research.

Self-Performed Lung Ultrasound for Home Monitoring of a Patient Positive for Coronavirus Disease 2019.

A subset of patients with coronavirus disease 2019 (COVID-19) and lung involvement pose a disposition challenge, particularly when hospital resources are constrained. Those not in respiratory failure are sent home, often with phone monitoring and/or respiratory rate and oxygen saturation monitoring. Hypoxemia may be a late presentation and is often preceded by abnormal lung findings on ultrasound. Early identification of pulmonary progression may preempt emergency hospitalization for respiratory decompensation and facilitate more timely admission. With the goal of safely isolating infected patients while providing advanced monitoring, we present a first report of patient self-performed lung ultrasound in the home with a hand-held device under the guidance of a physician using a novel teleguidance platform.