Nailfold Videocapillaroscopic Alterations as Markers of Microangiopathy in COVID-19 Patients.

Nailfold videocapillaroscopic alterations have been described in COVID-19, but their correlations with biomarkers of inflammation, coagulation and endothelial perturbation are still unclear, and no information is available on nailfold histopathology. Nailfold videocapillaroscopy was performed on fifteen patients with COVID-19 in Milan, Italy and the signs of microangiopathy were correlated with plasma biomarkers of inflammation (C reactive protein [CRP], ferritin), coagulation (D-dimer, fibrinogen), endothelial perturbation (Von Willebrand factor [VWF]) and angiogenesis (vascular endothelial growth factor [VEGF]) along with genetic drivers of COVID-19 susceptibility. Histopathological analysis of autoptic nailfold excisions was performed on fifteen patients who died for COVID-19 in New Orleans, United States. All COVID-19 patients studied with videocapillaroscopy showed alterations rarely seen in healthy individuals consistent with microangiopathy, such as hemosiderin deposits (sign of microthrombosis and microhemorrhages) and enlarged loops (sign of endotheliopathy). The number of hemosiderin deposits correlated both with ferritin and CRP levels (r = 0.67, p = 0.008 for both) and the number of enlarged loops significantly correlated with the levels of VWF (r = 0.67, p = 0.006). Ferritin levels were higher in non-O groups, determined by the rs657152 C > A cluster, (median 619, min-max 551-3266 mg/dL) than in the O group (373, 44-581 mg/dL, p = 0.006). Nailfold histology revealed microvascular damage, i.e., mild perivascular lymphocyte and macrophage infiltration and microvascular ectasia in the dermal vessels of all cases, and microthrombi within vessels in five cases. Alterations in nailfold videocapillaroscopy and elevated biomarkers of endothelial perturbation that match histopathologic findings open new perspectives in the possibility of non-invasively demonstrating microangiopathy in COVID-19.

Complement activation predicts negative outcomes in COVID-19: The experience from Northen Italian patients.

Coronavirus disease 19 (COVID-19) may present as a multi-organ disease with a hyperinflammatory and prothrombotic response (immunothrombosis) in addition to upper and lower airway involvement. Previous data showed that complement activation plays a role in immunothrombosis mainly in severe forms. The study aimed to investigate whether complement involvement is present in the early phases of the disease and can be predictive of a negative outcome. We enrolled 97 symptomatic patients with a positive RT-PCR for SARS-CoV-2 presenting to the emergency room. The patients with mild symptoms/lung involvement at CT-scan were discharged and the remaining were hospitalized. All the patients were evaluated after a 4-week follow-up and classified as mild (n. 54), moderate (n. 17) or severe COVID-19 (n. 26). Blood samples collected before starting any anti-inflammatory/immunosuppressive therapy were assessed for soluble C5b-9 (sC5b-9) and C5a plasma levels by ELISA, and for the following serum mediators by ELLA: IL-1ß, IL-6, IL-8, TNFα, IL-4, IL-10, IL-12p70, IFNγ, IFNα, VEGF-A, VEGF-B, GM-CSF, IL-2, IL-17A, VEGFR2, BLyS. Additional routine laboratory parameters were measured (fibrin fragment D-dimer, C-reactive protein, ferritin, white blood cells, neutrophils, lymphocytes, monocytes, platelets, prothrombin time, activated partial thromboplastin time, and fibrinogen). Fifty age and sex-matched healthy controls were also evaluated. SC5b-9 and C5a plasma levels were significantly increased in the hospitalized patients (moderate and severe) in comparison with the non-hospitalized mild group. SC5b9 and C5a plasma levels were predictive of the disease severity evaluated one month later. IL-6, IL-8, TNFα, IL-10 and complement split products were higher in moderate/severe versus non-hospitalized mild COVID-19 patients and healthy controls but with a huge heterogeneity. SC5b-9 and C5a plasma levels correlated positively with CRP, ferritin values and the neutrophil/lymphocyte ratio. Complement can be activated in the very early phases of the disease, even in mild non-hospitalized patients. Complement activation can be observed even when pro-inflammatory cytokines are not increased, and predicts a negative outcome.

Lung Ultrasound Findings and Endothelial Perturbation in a COVID-19 Low-Intensity Care Unit.

Hypercoagulability and endothelial dysfunction related to inflammation have been clearly demonstrated in COVID-19. However, their influence on thromboembolism, lung alterations and mortality in low-intensity-care patients with COVID-19 is not completely clarified. Our aims were to evaluate the prevalence of deep vein thrombosis (DVT) with compressive ultrasound (CUS); to describe lung ultrasound (LUS) features; and to study coagulation, inflammatory and endothelial perturbation biomarkers in COVID-19 patients at low-intensity care unit admission. The predictive value of these biomarkers on mortality, need for oxygen support and duration of hospitalization was also evaluated. Of the 65 patients included, 8 were non-survivors. CUS was negative for DVT in all patients. LUS Soldati and Vetrugno scores were strongly correlated (rho = 0.95) with each other, and both significantly differed in patients who needed oxygen therapy vs. those who did not (Soldati p = 0.017; Vetrugno p = 0.023), with coalescent B lines as the most prevalent pattern in patients with a worse prognosis. Mean (SD) levels of thrombomodulin and VCAM-1 were higher in non-survivors than in survivors (7283.9 pg/mL (3961.9 pg/mL) vs. 4800.7 pg/mL (1771.0 pg/mL), p = 0.004 and 2299 ng/mL (730.35 ng/mL) vs. 1451 ng/mL (456.2 ng/mL), p < 0.001, respectively). Finally, in a multivariate analysis model adjusted for age, sex and Charlson score, VCAM-1 level increase was independently associated with death [OR 1.31 (1.06, 1.81; p = 0.036)]. In conclusion, in a cohort of mild COVID-19 patients, we found no DVT events despite the highly abnormal inflammatory, endothelial and coagulation parameters. The presence of lung alterations at admission could not predict outcome. The endothelial perturbation biomarker VCAM-1 emerged as a promising prognostic tool for mortality in COVID-19.

Effects of Antibody Responses to Pre-Existing Coronaviruses on Disease Severity and Complement Activation in COVID-19 Patients.

The severity of coronavirus disease 2019 (COVID-19) may be influenced by pre-existing immune responses against endemic coronaviruses, but conflicting data have been reported. We studied 148 patients who were hospitalised because of a confirmed diagnosis of COVID-19, classified mild in 58, moderate in 44, and severe in 46. The controls were 27 healthy subjects. At admission, blood samples were collected for the measurement of biomarkers of disease severity and levels of the IgG against the receptor-binding domain (RBD) of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and pre-existing coronaviruses OC43, HKU1, NL63 and 229E. Higher levels of IgG antibodies against the RBD of pre-existing coronavirus (with the highest significance for anti-HKU1 IgG, p = 0.01) were found in patients with mild disease, compared with those with moderate or severe disease. Multivariable logistic regression confirmed the association of high levels of antibodies to pre-existing coronavirus with mild disease and showed their associations with low levels of the complement activation marker SC5b-9 (p range = 0.007-0.05). High levels of anti-NL63 antibodies were associated with low levels of the coagulation activation marker D-dimer (p = 0.04), while high levels of IgG against 229E were associated with low levels of the endothelial activation marker von Willebrand factor (p = 0.05). Anti-SARS-CoV-2-neutralising activity of plasma positively correlated with anti-SARS-CoV-2 IgG (r = 0.53, p = 0.04) and with anti-HKU1 IgG (r = 0.51, p = 0.05). In hospitalised patients with COVID-19, high levels of antibodies to pre-existing coronaviruses are associated with mild disease, suggesting that their measurement could be useful in predicting the severity of the disease.

Urticarial vasculitis: Clinical and laboratory findings with a particular emphasis on differential diagnosis.

Urticarial vasculitis (UV) is a rare cutaneous vasculitis of small vessels characterized by recurrent episodes of wheal-like lesions that tend to last more than 24 hours, healing with a residual ecchymotic postinflammatory hyperpigmentation. The histopathologic pattern of UV is that of leukocytoclastic vasculitis, consisting of fibrinoid necrosis of dermal vessels' walls and neutrophil-rich perivascular inflammatory infiltrates. Although its etiopahogenesis remains still undefined, UV is now regarded as an immune complex-driven disease with activation of the complement cascade, leading to exaggerated production of anaphylatoxins that are responsible for neutrophil recruitment and activation. This condition can be categorized into 2 main entities according to serum complement levels: normocomplementemic UV and hypocomplementemic UV, the latter being associated with circulating anti-C1q autoantibodies and possible extracutaneous manifestations. Systemic multiorgan involvement may be seen particularly in syndromic hypocomplementemic UV, also known as McDuffie syndrome. This review summarizes the clinicopathological and laboratory features as well as the underlying pathophysiological mechanisms of UV. A focus on its main differential diagnoses is provided, that is, chronic spontaneous urticaria, bullous pemphigoid, IgA (Henoch-Schönlein purpura) and IgM/IgG immune complex vasculitis, lupus erythematous tumidus, Wells syndrome, erythema multiforme, cutaneous mastocytosis, cryopyrin-associated periodic syndromes, and coronavirus disease 2019-associated and anti-severe acute respiratory syndrome coronavirus 2-vaccine-associated urticarial eruptions.

Side effects among healthcare workers from a large Milan university hospital after second dose of BNT162b2 mRNA COVID-19 vaccine.

BACKGROUND: In Italy, healthcare workers (HCWs) were among the first to receive COVID-19 vaccination. Aim of the present study is to evaluate frequency and severity of adverse events (AEs) following the second dose of BNT162b2 vaccine among HCWs of a large university hospital in Milan, Italy. METHODS: One month after having received the second dose of vaccine, HCWs filled-in a form about type, severity, and duration of post-vaccination local and systemic symptoms. We calculated the overall frequency of AEs and used multivariable Poisson regression models (adjusted for sex, age, BMI, smoking, allergy history, previous SARS-CoV-2 infection, anti-hypertensive therapy, and occupation) to calculate risk ratios (RR) and 95% confidence intervals (CI) of AEs according to selected variables. RESULTS: We included 3659 HCWs. Overall, 2801 (76.6%) experienced at least one local event, with pain at injection site being the most frequent (2788, 76.2%). Systemic events were reported by 2080 (56.8%) HCWs, with fatigue (52.3%), muscle pain (42.2%), headache (37.7%), joint pain (31.9%), and fever (26.2%) being the most frequent. Risks of systemic events were associated with female gender (RR=1.14, CI: 1.06-1.23), age (strong decrease with increasing age, p-trend<0.001), allergy history (RR=1.13, CI: 1.05-1.20), and current smoking (RR=0.90, CI: 0.84-0.97). HCWs with previous SARS-CoV-2 infection (even if symptomatic) were not at increased risk. CONCLUSIONS: Both local and systemic acute effects after second dose of BNT162b2 vaccine were frequently reported. However, symptoms were mostly light/mild and of short duration. Thus, our findings support the safety of COVID-19 vaccination in adults in relatively good health.

Consumption of complement in a 26-year-old woman with severe thrombotic thrombocytopenia after ChAdOx1 nCov-19 vaccination.

Extremely rare reactions characterized by thrombosis and thrombocytopenia have been described in subjects that received ChAdOx1 nCoV-19 vaccination 5-16 days earlier. Although patients with vaccine-induced thrombotic thrombocytopenia (VITT) have high levels of antibodies to platelet factor 4 (PF4)-polyanion complexes, the exact mechanism of the development of thrombosis is still unknown. Here we reported serum studies as well as proteomics and genomics analyses demonstrating a massive complement activation potentially linked to the presence of anti-PF4 antibodies in a patient with severe VITT. At admission, complement activity of the classical and lectin pathways were absent (0% for both) with normal levels of the alternative pathway (73%) in association with elevated levels of the complement activation marker sC5b-9 (630 ng/mL [n.v. 139-462 ng/mL]) and anti-PF4 IgG (1.918 OD [n.v. 0.136-0.300 OD]). The immunoblotting analysis of C2 showed the complete disappearance of its normal band at 110 kDa. Intravenous immunoglobulin treatment allowed to recover complement activity of the classical pathway (91%) and lectin pathway (115%), to reduce levels of sC5b-9 (135 ng/mL) and anti-PF4 IgG (0.681 OD) and to normalize the C2 pattern at immunoblotting. Proteomics and genomics analyses in addition to serum studies showed that the absence of complement activity during VITT was not linked to alterations of the C2 gene but rather to a strong complement activation leading to C2 consumption. Our data in a single patient suggest monitoring complement parameters in other VITT patients considering also the possibility to target complement activation with specific drugs.

Increased Risk of Urticaria/Angioedema after BNT162b2 mRNA COVID-19 Vaccine in Health Care Workers Taking ACE Inhibitors.

Urticarial eruptions and angioedema are the most common cutaneous reactions in patients undergoing mRNA COVID-19 vaccinations. The vasoactive peptide bradykinin has long been known to be involved in angioedema and recently also in urticaria. Bradykinin is mainly catabolized by angiotensin-converting enzyme (ACE), which is inhibited by ACE inhibitors, a commonly employed class of antihypertensive drugs. We evaluated the risk of developing urticaria/angioedema after inoculation with the BNT162b2 mRNA COVID-19 vaccine in a population of 3586 health care workers. The influences of ACE inhibitors and selected potential confounding variables (sex, age, previous SARS-CoV-2 infection, and allergy history) were evaluated by fitting univariate and multivariable Poisson regression models. The overall cumulative incidence of urticaria/angioedema was 1.8% (65 out of 3586; 95% CI: 1.4-2.3%). Symptoms were mild, and no subject consulted a physician. Subjects taking ACE inhibitors had an adjusted three-fold increased risk of urticaria/angioedema (RR 2.98, 95% CI: 1.12-7.96). When we restricted the analysis to those aged 50 years or more, the adjusted RR was 3.98 (95% CI: 1.44-11.0). In conclusion, our data indicate that subjects taking ACE inhibitors have an increased risk of urticaria/angioedema after vaccination with the BNT162b2 mRNA COVID-19 vaccine. Symptoms are mild and self-limited; however, they should be considered to adequately advise subjects undergoing vaccination.

Mortality in Patients with COVID-19 on Renin Angiotensin System Inhibitor Long-Term Treatment: An Observational Study Showing that Things Are Not Always as They Seem.

INTRODUCTION: At the beginning of the coronavirus disease 2019 (COVID-19) pandemic, controversial data were reported concerning angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs) that induced a number of physicians to stop using them in patients with COVID-19. Although large-scale studies have ruled out this concern, it is common experience that patients with COVID-19 taking ACE inhibitors or ARBs are at increased risk of death. The aim of this study was to investigate the reasons for this apparently high mortality rate. METHODS: During the first wave of the pandemic, we conducted a field study of 427 consecutive patients with COVID-19 upon their admission to the emergency department of a hospital in one of the most severely hit cities in northern Italy, and 30 days later. The disease was defined as being mild, moderate or severe on the basis of clinical, laboratory and imaging data, and a multivariate model was used to analyse the determinants of mortality. RESULTS: Within 30 days of admission, 31.6% of the patients treated with ACE inhibitors or ARBs and 15.2% of those not treated with these drugs had died. Multivariate analysis showed that the determinants of mortality were age (p = 0.0001), hypertension (p = 0.0120) and diabetes (p = 0.0129), whereas ACE inhibitors or ARBs had no effect on mortality. There was no significant difference between the patients treated with ACE inhibitors and those treated with ARBs. CONCLUSION: The apparently increased mortality of patients with COVID-19 receiving long-term treatment with ACE inhibitors or ARBs is not due to the drugs themselves, but to the conditions associated with their use.

Chromosome 3 cluster rs11385942 variant links complement activation with severe COVID-19.

BACKGROUND: Genetic variation at a multigene cluster at chromosome 3p21.31 and the ABO blood group have been associated with the risk of developing severe COVID-19, but the mechanism remains unclear. Complement activation has been associated with COVID-19 severity. OBJECTIVE: The aim of this study was to examine whether chromosome 3p21.31 and the ABO variants are linked to the activation of the complement cascade in COVID-19 patients. METHODS: We considered 72 unrelated European hospitalized patients with genetic data and evaluation of circulating C5a and soluble terminal complement complex C5b-9 (SC5b-9). Twenty-six (36.1%) patients carried the rs11385942 G>GA variant and 44 (66.1%) non-O blood group associated with increased risk of severe COVID-19. RESULTS: C5a and SC5-b9 plasma levels were higher in rs11385949 GA carriers than in non-carriers (P = 0.041 and P = 0.012, respectively), while C5a levels were higher in non-O group than in O group patients (P = 0.019). The association between rs11385949 and SC5b-9 remained significant after adjustment for ABO and disease severity (P = 0.004) and further correction for C5a (P = 0.018). There was a direct relationship between upper airways viral load and SC5b-9 in carriers of the rs11385949 risk allele (P = 0.032), which was not observed in non-carriers. CONCLUSIONS: The rs11385949 G>GA variant, tagging the chromosome 3 gene cluster variation and predisposing to severe COVID-19, is associated with enhanced complement activation, both with C5a and terminal complement complex, while non-O blood group with C5a levels. These findings provide a link between genetic susceptibility to more severe COVID-19 and complement activation.

Complement activation and endothelial perturbation parallel COVID-19 severity and activity.

BACKGROUND: Animal models and few clinical reports suggest the involvement of the complement system in the onset of severe manifestations of coronavirus disease-2019 (COVID-19). However, complement contribution to endotheliopathy and hypercoagulability has not been elucidated yet. OBJECTIVE: To evaluate the association among complement activation, endothelial damage and disease severity or activity in COVID-19 patients. METHODS: In this single-centre cohort study, 148 patients with COVID-19 of different severity were evaluated upon hospital admission and 30 days later. Markers of complement activation (SC5b-9 and C5a) and endothelial perturbation (von Willebrand factor [vWF], tissue-type plasminogen activator [t-PA], plasminogen activator inhibitor-1 [PAI-1], soluble thrombomodulin [sTM], and soluble endothelial selectin [sE-selectin]) were measured in plasma. RESULTS: The patients had high plasma levels of SC5b-9 and C5a (p = 0.0001 for both) and vWF, t-PA and PAI-1 (p = 0.0001 for all). Their SC5b-9 levels correlated with those of vWF (r = 0.517, p = 0.0001) and paralleled disease severity (severe vs mild p = 0.0001, severe vs moderate p = 0.026 and moderate vs mild p = 0.001). The levels of sE-selectin were significantly increased only in the patients with severe disease. After 30 days, plasma SC5b-9, C5a and vWF levels had significantly decreased (p = 0.0001 for all), and 43% of the evaluated patients had normal levels. CONCLUSIONS: Complement activation is boosted during the progression of COVID-19 and dampened during remission, thus indicating its role in the pathophysiology of the disease. The association between complement activation and the biomarkers of endothelial damage suggests that complement may contribute to tissue injury and could be the target of specific therapy.