Memory B cell responses to Omicron subvariants after SARS-CoV-2 mRNA breakthrough infection in humans.

Individuals who receive a third mRNA vaccine dose show enhanced protection against severe COVID-19, but little is known about the impact of breakthrough infections on memory responses. Here, we examine the memory antibodies that develop after a third or fourth antigenic exposure by Delta or Omicron BA.1 infection, respectively. A third exposure to antigen by Delta breakthrough increases the number of memory B cells that produce antibodies with comparable potency and breadth to a third mRNA vaccine dose. A fourth antigenic exposure with Omicron BA.1 infection increased variant-specific plasma antibody and memory B cell responses. However, the fourth exposure did not increase the overall frequency of memory B cells or their general potency or breadth compared to a third mRNA vaccine dose. In conclusion, a third antigenic exposure by Delta infection elicits strain-specific memory responses and increases in the overall potency and breadth of the memory B cells. In contrast, the effects of a fourth antigenic exposure with Omicron BA.1 are limited to increased strain-specific memory with little effect on the potency or breadth of memory B cell antibodies. The results suggest that the effect of strain-specific boosting on memory B cell compartment may be limited.

Humoral immunity to SARS-CoV-2 elicited by combination COVID-19 vaccination regimens.

The SARS-CoV-2 pandemic prompted a global vaccination effort and the development of numerous COVID-19 vaccines at an unprecedented scale and pace. As a result, current COVID-19 vaccination regimens comprise diverse vaccine modalities, immunogen combinations, and dosing intervals. Here, we compare vaccine-specific antibody and memory B cell responses following two-dose mRNA, single-dose Ad26.COV.2S, and two-dose ChAdOx1, or combination ChAdOx1/mRNA vaccination. Plasma-neutralizing activity, as well as the magnitude, clonal composition, and antibody maturation of the RBD-specific memory B cell compartments, showed substantial differences between the vaccination regimens. While individual monoclonal antibodies derived from memory B cells exhibited similar binding affinities and neutralizing potency against Wuhan-Hu-1 SARS-CoV-2, there were significant differences in epitope specificity and neutralizing breadth against viral variants of concern. Although the ChAdOx1 vaccine was inferior to mRNA and Ad26.COV.2S in several respects, biochemical and structural analyses revealed enrichment in a subgroup of memory B cell neutralizing antibodies with distinct RBD-binding properties resulting in remarkable potency and breadth.

Analysis of memory B cells identifies conserved neutralizing epitopes on the N-terminal domain of variant SARS-Cov-2 spike proteins.

SARS-CoV-2 infection or vaccination produces neutralizing antibody responses that contribute to better clinical outcomes. The receptor-binding domain (RBD) and the N-terminal domain (NTD) of the spike trimer (S) constitute the two major neutralizing targets for antibodies. Here, we use NTD-specific probes to capture anti-NTD memory B cells in a longitudinal cohort of infected individuals, some of whom were vaccinated. We found 6 complementation groups of neutralizing antibodies. 58% targeted epitopes outside the NTD supersite, 58% neutralized either Gamma or Omicron, and 14% were broad neutralizers that also neutralized Omicron. Structural characterization revealed that broadly active antibodies targeted three epitopes outside the NTD supersite including a class that recognized both the NTD and SD2 domain. Rapid recruitment of memory B cells producing these antibodies into the plasma cell compartment upon re-infection likely contributes to the relatively benign course of subsequent infections with SARS-CoV-2 variants, including Omicron.

Conserved Neutralizing Epitopes on the N-Terminal Domain of Variant SARS-CoV-2 Spike Proteins.

SARS-CoV-2 infection or vaccination produces neutralizing antibody responses that contribute to better clinical outcomes. The receptor binding domain (RBD) and the N-terminal domain (NTD) of the spike trimer (S) constitute the two major neutralizing targets for the antibody system. Neutralizing antibodies targeting the RBD bind to several different sites on this domain. In contrast, most neutralizing antibodies to NTD characterized to date bind to a single supersite, however these antibodies were obtained by methods that were not NTD specific. Here we use NTD specific probes to focus on anti-NTD memory B cells in a cohort of pre-omicron infected individuals some of which were also vaccinated. Of 275 NTD binding antibodies tested 103 neutralized at least one of three tested strains: Wuhan-Hu-1, Gamma, or PMS20, a synthetic variant which is extensively mutated in the NTD supersite. Among the 43 neutralizing antibodies that were further characterized, we found 6 complementation groups based on competition binding experiments. 58% targeted epitopes outside the NTD supersite, and 58% neutralized either Gamma or Omicron, but only 14% were broad neutralizers. Three of the broad neutralizers were characterized structurally. C1520 and C1791 recognize epitopes on opposite faces of the NTD with a distinct binding pose relative to previously described antibodies allowing for greater potency and cross-reactivity with 7 different variants including Beta, Delta, Gamma and Omicron. Antibody C1717 represents a previously uncharacterized class of NTD-directed antibodies that recognizes the viral membrane proximal side of the NTD and SD2 domain, leading to cross-neutralization of Beta, Gamma and Omicron. We conclude SARS-CoV-2 infection and/or Wuhan-Hu-1 mRNA vaccination produces a diverse collection of memory B cells that produce anti-NTD antibodies some of which can neutralize variants of concern. Rapid recruitment of these cells into the antibody secreting plasma cell compartment upon re-infection likely contributes to the relatively benign course of subsequent infections with SARS-CoV-2 variants including omicron.

Intrinsic DNA Shape Accounts for Affinity Differences between Hox-Cofactor Binding Sites.

Transcription factors bind to their binding sites over a wide range of affinities, yet how differences in affinity are encoded in DNA sequences is not well understood. Here, we report X-ray crystal structures of four heterodimers of the Hox protein AbdominalB bound with its cofactor Extradenticle to four target DNA molecules that differ in affinity by up to ∼20-fold. Remarkably, despite large differences in affinity, the overall structures are very similar in all four complexes. In contrast, the predicted shapes of the DNA binding sites (i.e., the intrinsic DNA shape) in the absence of bound protein are strikingly different from each other and correlate with affinity: binding sites that must change conformations upon protein binding have lower affinities than binding sites that have more optimal conformations prior to binding. Together, these observations suggest that intrinsic differences in DNA shape provide a robust mechanism for modulating affinity without affecting other protein-DNA interactions.

Homophilic and Heterophilic Interactions of Type II Cadherins Identify Specificity Groups Underlying Cell-Adhesive Behavior.

Type II cadherins are cell-cell adhesion proteins critical for tissue patterning and neuronal targeting but whose molecular binding code remains poorly understood. Here, we delineate binding preferences for type II cadherin cell-adhesive regions, revealing extensive heterophilic interactions between specific pairs, in addition to homophilic interactions. Three distinct specificity groups emerge from our analysis with members that share highly similar heterophilic binding patterns and favor binding to one another. Structures of adhesive fragments from each specificity group confirm near-identical dimer topology conserved throughout the family, allowing interface residues whose conservation corresponds to specificity preferences to be identified. We show that targeted mutation of these residues converts binding preferences between specificity groups in biophysical and co-culture assays. Our results provide a detailed understanding of the type II cadherin interaction map and a basis for defining their role in tissue patterning and for the emerging importance of their heterophilic interactions in neural connectivity.

Electric cardioversion of atrial fibrillation resulting in pulmonary oedema in patient with apical hypertrophic cardiomyopathy.

The direct-current electric shock is considered to be safe treatment of arrhythmias and rarely leads to serious hemodynamic complications. A 62-year-old patient was admitted to the hospital due to a first symptomatic episode of atrial fibrillation. Patient was diagnosed with apical hypertrophic cardiomyopathy 20 years ago. Transoesophageal echocardiography was performed to exclude an atrial thrombus followed by electrical cardioversion with restoration of sinus rhythm. After 6 hours symptoms of pulmonary oedema developed. The patient's condition improved after furosemide administration. As the possible cause of the oedema, inotropic effect of administered propafenone and atrial stunning were considered. The atria seem to be responsible for important part of forward cardiac output even during AF, especially in cardiomyopathies. Contractility deterioration of the left atrium (stunning) along with earlier resumption of the right atrium contractile function could be associated with hemodynamic instability causing pulmonary oedema in subjects with hypertrophied myocardium. It is necessary to take into consideration the atrial function while administrating antiarrhythmic drugs, especially those with negative inotropic effect.

Structural and energetic determinants of adhesive binding specificity in type I cadherins.

Type I cadherin cell-adhesion proteins are similar in sequence and structure and yet are different enough to mediate highly specific cell-cell recognition phenomena. It has previously been shown that small differences in the homophilic and heterophilic binding affinities of different type I family members can account for the differential cell-sorting behavior. Here we use a combination of X-ray crystallography, analytical ultracentrifugation, surface plasmon resonance and double electron-electron resonance (DEER) electron paramagnetic resonance spectroscopy to identify the molecular determinants of type I cadherin dimerization affinities. Small changes in sequence are found to produce subtle structural and dynamical changes that impact relative affinities, in part via electrostatic and hydrophobic interactions, and in part through entropic effects because of increased conformational heterogeneity in the bound states as revealed by DEER distance mapping in the dimers. These findings highlight the remarkable ability of evolution to exploit a wide range of molecular properties to produce closely related members of the same protein family that have affinity differences finely tuned to mediate their biological roles.

Echocardiographic evaluation of external iliac artery Doppler waveform in patients with coronary artery disease.

Visual interpretation of the Doppler waveform in the common femoral or distal external iliac artery (EIA) was reported to be useful in screening for proximal peripheral artery occlusive disease (PAOD) in patients with lower limb ischemia. Commonly patients with coronary artery disease (CAD) referred for echocardiography have coexistent arterial pathology. Therefore, we decided to study whether echocardiographic evaluation of the distal EIA flow can be useful for detection of PAOD in patients with CAD. We studied 150 consecutive patients (pts) with CAD referred for echocardiography. At the end of an echocardiographic examination, evaluation of the flow in the distal EIA with an echocardiographic probe was performed. The Doppler waveform was classified as normal-with early diastolic flow reversal or abnormal-without early diastolic flow reversal. Echocardiographic findings were compared in a blinded fashion with the results of the ankle brachial index measurements (ABI). Based on the ABI ≤ 0.9, peripheral artery disease was diagnosed in 54 pts (36%) and abnormal external iliac Doppler waveform was found in 27 pts (18%). Sensitivity of abnormal external iliac Doppler waveform in predicting PAOD was 48%, specificity 99%, positive predictive value (PPV) 96%, and negative predictive value 77%. Peripheral arterial occlusive disease is common in patients with CAD referred for echocardiographic study. Echocardiographic assessment of distal EIA Doppler waveform has low sensitivity, but high specificity and high PPV in the diagnosis of peripheral arterial occlusive disease.

Echocardiographic evaluation of aorto-iliac occlusive disease.

Several studies demonstrated feasibility of visual assessment of the common femoral artery Doppler waveform, in an indirect evaluation of aorto-iliac segment stenosis. Patients with cardiac diseases referred for echocardiography often have coexistent arterial pathology. Since many of them are potential candidates for endovascular procedures, we decided to study, whether echocardiography can be useful for detection of aorto-iliac occlusive disease. We evaluated 92 patients with abdominal aortic aneurysm or peripheral artery occlusive disease, referred from the vascular surgery department for cardiac evaluation before surgery. At the end of an echocardiographic examination, evaluation of flow in the distal external iliac arteries with an echocardiographic probe was performed. The Doppler waveform was classified into normal--with early diastolic flow reversal or abnormal--without early diastolic flow reversal. Echocardiographic results were compared in a blinded fashion with reports from computed tomography angiography. Overall there were 58 iliac segments with significant (≥70%) area stenosis or occlusion and 126 iliac segments without significant disease on computed tomography angiography. Abnormal Doppler waveform was found in 56 out of 58 abnormal iliac segments-sensitivity 97%, and normal waveform was found in 106 out of 126 normal iliac segments-specificity 84%. Positive predictive value of abnormal Doppler waveform for significant iliac disease was 74%, and negative predicting value was 98%. Detection of significant stenoses in aorto-iliac segments is feasible with echocardiography. Further studies are necessary to evaluate its potential utility in a population of patients with cardiac disease referred for echocardiographic study.

Electrocardiographic differentiation between acute pulmonary embolism and non-ST elevation acute coronary syndromes at the bedside.

BACKGROUND: Clinical picture of acute pulmonary embolism (APE), with wide range of electrocardiographic (ECG) abnormalities can mimic acute coronary syndromes. OBJECTIVES: Assessment of standard 12-lead ECG usefulness in differentiation at the bedside between APE and non-ST elevation acute coronary syndrome (NSTE-ACS). METHODS: Retrospective analysis of 143 patients: 98 consecutive patients (mean age 63.4 +/- 19.4 year, 45 M) with APE and 45 consecutive patients (mean age 72.8 +/- 10.8 year, 44 M) with NSTE-ACS. Standard ECGs recorded on admission were compared in separated groups. RESULTS: Right bundle branch block (RBBB) and S(1)S(2)S(3) or S(1)Q(3)T(3) pattern were found in similar frequency in both groups (10 [11%] APE patients vs 6 [14%] NSTE-ACS patients, 27 [28%] patients vs 7 [16%] patients, respectively, NS). Negative T waves in leads V(1-3) together with negative T waves in inferior wall leads II, III, aVF (OR 1.3 [1.14-1.68]) significantly indicated APE with a positive predictive value of 85% and specificity of 87%. However, counterclockwise axis rotation (OR 4.57 [2.74-7.61]), ventricular premature beats (OR 2.60 [1.60-4.19]), ST depression in leads V(1-3) (OR 2.25 [1.43-3.56]), and negative T waves in leads V(5-6) (OR 2.08 [1.31-3.29]) significantly predicted NSTE-ACS. CONCLUSIONS: RBBB, S(1)S(2)S(3), or S(1)Q(3)T(3) pattern described as characteristic for APE were not helpful in the differentiation between APE and NSTE-ACS in studied group. Coexistence of negative T waves in precordial leads V(1-3) and inferior wall leads may suggest APE diagnosis.

B-type natriuretic peptide in acute pulmonary embolism.

Myocardial stretch leads to the natriuretic peptides release in acute or chronic left ventricular dysfunction. However, there is an accumulating evidence that B-type natriuretic peptide (BNP) and its N-terminal fragment (NT-proBNP) may originate from right ventricle and their concentrations are elevated in patients with acute pulmonary embolism (APE) especially when resulting in right ventricular dysfunction (RVD). Recently it is underlined that severity assessment of APE as well as the risk stratification and therapy selection is based both on patients' hemodynamic status and markers of myocardial injury and RVD. BNP and NT-proBNP are helpful in identifying patients with RVD in APE, emerging as an adjunctive tool to echocardiography. Elevated BNP or NT-proBNP levels are also significant predictors of death and/or complicated clinical course in APE.

Elevated D-dimer concentration identifies patients with incomplete recanalization of pulmonary artery thromboemboli despite 6 months anticoagulation after the first episode of acute pulmonary embolism.

BACKGROUND: Despite long-term anticoagulation in some patients after acute pulmonary embolism (APE) pulmonary thrombi are not completely resolved. We hypothesized that elevated D-dimer concentration reflecting increased endogenous fibrinolysis may indicate incomplete pulmonary thrombi resolution after the first episode of PE. METHODS: 55 patients aged 54.7+/-18.6 years were anticoagulated for 6 months with acenocumarol (74.5% patients) or low molecular weight heparin (25.5% patients) when control spiral computed tomography (sCT), lung perfusion scintigraphy and D-dimer assessment were performed. RESULTS: Incomplete recanalization of pulmonary circulation was found in 39 (70.9%) patients - thrombi at sCT and/or > or =1 wedge-shaped perfusion defect at scintigraphy. Age, sex, rate of unprovoked APE, malignancies, thrombolysis in the acute phase and type of long-term anticoagulation were similar in patients without and with complete recanalization. D-dimer at follow-up but not on admission was higher in patients with then without incomplete recanalization (median 340 (80-2280) vs 160 (60-390) ng/mL, p=0.02). All 11 (20%) patients with D-dimer level >500 ng/mL at follow-up did not resolve thromboemboli completely. ROC analysis showed that D-dimer at follow-up identified patients with incomplete recanalization (AUC 0.709, 95% CI (0.560-0.831), p=0.007). Multivariable analysis confirmed that D-dimer >350 ng/mL at follow-up and right ventricle dysfunction at the diagnosis were independent predictors of incomplete recanalization (OR 18.58 (95% CI 1.97-175.19) and 7.03 (95% CI 1.43-34.6), respectively, p=0.0006). CONCLUSION: Elevated D-dimer after 6 months anticoagulation and right ventricular dysfunction at the diagnosis predict incomplete recanalization of pulmonary circulation after first episode of APE.

Right ventricle injury during acute pulmonary embolism leads to its remodeling.

Right ventricular (RV) overload and hypoxia in acute pulmonary embolism (APE) may lead to RV myocardium injury reflected by elevated cardiac troponin levels. We studied 26 patients aged 57.2+/-17.8 years with first episode of APE. On admission troponin T (TnT) was measured. Transthoracic echocardiography was performed after 6 months of anticoagulation. Myocardial injury (TnT > or =0.03 ng/ml) was observed in 8 (30.8%) patients at the diagnosis. At follow up RV diastolic area tended to be larger in group with myocardial injury (25.0 (20.8-38.6) vs 18.4 (17.7-23.3) cm(2), p=0.06). Tricuspid annulus systolic velocity at tissue Doppler was lower in group with myocardial injury (0.12 (0.11-0.13) vs 0.15 (0.13-0.21) m/s, p=0.04), while no such a relationship was found for mitral annulus systolic velocity. TnT concentration correlated with RV diastolic area (r=0.61) and tricuspid annulus systolic velocity (r=-0.58) although not significantly (p=0.08 and p=0.09. respectively). Our data suggest that RV injury in acute phase of PE may lead to its remodeling.

[Pulmonary embolism: a difficult diagnostic problem]. / Zatorowosc plucna--trudny problem diagnostyczny.

INTRODUCTION: The mortality of untreated pulmonary embolism (PE) is estimated at approximately 30% of patients, whereas treatment decreases it to 2-8%. A specific combination of symptoms present in PE may suggest other cardiac or lung disorder. OBJECTIVES: To evaluate frequencies of clinical symptoms and changes in diagnostic investigations misleading to the recognition of acute coronary syndrome (ACS) or lung diseases (Ld) in PE patients. PATIENTS AND METHODS: Retrospective analysis of 154 records of individuals with recognized PE allowed to divide patients into groups suggestive of ACS (min. 2 of: chest pain, ischemic changes on electrocardiogram (ECG) and elevated cardiac troponin T level [cTnT >0.01 ng/ml]) or suggestive of the Ld (min. 2 of: dyspnea, cough, fever, lung consolidations on chest radiograph). RESULTS: Fifty-five (36%) patients were classified to the ACS group and 54 (35%) to Ld group, while 69 (45%) patients were not included to either group. Twenty-four (16%) patients fulfilled criteria of both groups. There were no significant differences in the frequency of coronary heart disease, heart failure, atrial fibrillation and chronic obstructive pulmonary disease between groups. Elevated troponin level was observed in 68% of patients with chest pain and changes on ECG, and in 26% of patients without coexistence of these symptoms (p < 0.05). CONCLUSIONS: In most patients with final diagnosis of PE, symptoms and initial investigation results can mislead to the diagnosis of ACS or lung disease. The chest pain and ischemic changes on ECG are frequently associated with the myocardial injury resulting in increased troponin levels in PE patients.

Persistent NT-proBNP elevation in acute pulmonary embolism predicts early death.

AIMS: Low levels of brain natriuretic peptides on admission identify low-risk patients with acute pulmonary embolism (APE) through their high NPV for mortality. However, increased natriuretic peptide values on admission are less helpful for identifying high-risk patients due to their low PPV. The aim of the study was to test whether the PPV for mortality can be improved by performing serial NT-proBNP measurements on admission, at 12 h, and at 24 h. METHODS AND RESULTS: We prospectively included 113 consecutive patients with APE (mean age 63+/-18 years), of whom 10 had clinically massive APE. Thirty-day mortality was 15% (95% CI: 8%-22%). In survivors, median NT-proBNP levels decreased within 24 h from 1895 ng/L (range: 16-33,340) to 1007 ng/L (range: 9-33,243) (p<0.001). In non-survivors, median NT-proBNP levels at baseline (11,491 ng/L, range: 618-60,958) remained elevated at 24 h (8139 ng/L, range: 35-70,018; p=NS). The 30-day mortality rate in the group of 18 patients with NT-proBNP >7500 ng/L and less than 50% decrease of NT-proBNP within 24 h was 61% (95% CI: 39%-84%). PPV and NPV of NT-proBNP >7500 ng/L on admission and less than 50% decrease of NT-proBNP within 24 h were 61% and 94%, respectively. CONCLUSION: Persistent elevation of plasma NT-proBNP levels within 24 h after APE diagnosis indicates ongoing right ventricular dysfunction with a poor prognosis. These critically ill patients may be candidates for rapid aggressive intervention, including thrombolysis, catheter thrombectomy, or surgical embolectomy.

Plasma heart-type fatty acid binding protein is superior to troponin and myoglobin for rapid risk stratification in acute pulmonary embolism.

BACKGROUND: Irreversible right ventricular (RV) failure with myocardial damage may precipitate fatal outcome in acute pulmonary embolism (APE). Cytoplasmic heart-type fatty acid binding protein (H-FABP) is a sensitive and specific biomarker of myocardial damage. We assessed which biomarker of myocardial damage or RV stretching is the most useful for short-term risk stratification in APE. METHODS: We analyzed 77 patients (51 F, 26 M) aged 65.3+/-16.0 years with confirmed APE. On admission, systemic blood pressure and transthoracic echocardiography (for RV overload) were recorded and plasma concentrations of myoglobin (Mb), cardiac troponin T (cTnT), N-terminal fragment of proBNP (NT-proBNP) and H-FABP were evaluated. RESULTS: Fifteen (19.5%) patients died and 24 (31.2%) experienced complicated clinical course (CCC)-death/thrombolysis/cardiopulmonary resuscitation/intravenous vasopressors. Hazard ratio analysis demonstrated that plasma H-FABP, Mb, cTnT and NT-proBNP concentrations predicted fatal outcome. When only APE-related deaths were considered, plasma H-FABP concentrations indicated fatal outcome. Multivariate hazard ratio analysis revealed H-FABP as the only 30-day mortality predictor (HR 1.02 CI 95% 1.01-1.05). CONCLUSIONS: H-FABP measured on admission is useful for short-term risk stratification in APE. It appears to be superior to cTnT, NT-proBNP and Mb in the prediction of 30-day APE-related mortality.