Convalescent COVID-19 Patients Research Articles

IFN-γ decreases PD-1 in T lymphocytes from convalescent COVID-19 patients via the AKT/GSK3β signaling pathway

Post-COVID-19 syndrome may be associated with the abnormal immune status. Compared with the unexposed age-matched elder group, PD-1 in the CD8+ T cells from recovered COVID-19 patients was significantly lower. IFN-γ in the plasma of COVID-19 convalescent patients was increased, which inhibited PD-1 expression in CD8+ T cells from COVID-19 convalescent patients. scRNA-seq bioinformatics analysis revealed that AKT/GSK3β may regulate the INF-γ/PD-1 axis in CD8+ T cells from COVID-19 convalescent patients. In parallel, an IFN-γ neutralizing antibody reduced AKT and increased GSK3β in PBMCs. An AKT agonist (SC79) significantly decreased p-GSK3β. Moreover, AKT decreased PD-1 on CD8+ T cells, and GSK3β increased PD-1 on CD8+ T cells according to flow cytometry analysis. Collectively, we demonstrated that recovered COVID-19 patients may develop long COVID. Increased IFN-γ in the plasma of recovered Wuhan COVID-19 patients contributed to PD-1 downregulation on CD8+ T cells by regulating the AKT/GSK3β signaling pathway.

Single-Cell RNA Sequencing Reveals HIF1A as a Severity-Sensitive Immunological Scar in Circulating Monocytes of Convalescent Comorbidity-Free COVID-19 Patients.

COVID-19, caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), is characterized by a wide range of clinical symptoms and a poorly predictable disease course. Although in-depth transcriptomic investigations of peripheral blood samples from COVID-19 patients have been performed, the detailed molecular mechanisms underlying an asymptomatic, mild or severe disease course, particularly in patients without relevant comorbidities, remain poorly understood. While previous studies have mainly focused on the cellular and molecular dissection of ongoing COVID-19, we set out to characterize transcriptomic immune cell dysregulation at the single-cell level at different time points in patients without comorbidities after disease resolution to identify signatures of different disease severities in convalescence. With single-cell RNA sequencing, we reveal a role for hypoxia-inducible factor 1-alpha (HIF1A) as a severity-sensitive long-term immunological scar in circulating monocytes of convalescent COVID-19 patients. Additionally, we show that circulating complexes formed by monocytes with either T cells or NK cells represent a characteristic cellular marker in convalescent COVID-19 patients irrespective of their preceding symptom severity. Together, these results provide cellular and molecular correlates of recovery from COVID-19 and could help in immune monitoring and in the design of new treatment strategies.

Dynamics of serum cross-neutralization capacity against SARS-CoV-2 Delta variant in convalescent COVID-19 patients.

The magnitude and breadth of the neutralizing antibody response against variants of concern following natural infection would provide valuable insights regarding the immune response induced by severe acute respiratory syndrome-related coronavirus(SARS-CoV-2) infection. Herein, 25 patients were followed at 30 ±7 (Visit 1), 90± 15 (Visit 2), and 180 ± 15 (Visit 3) days post symptom onset (PSO). The neutralization titers against both Wuhan-Hu-1 (WT) and Delta variant were analyzed in parallel along with anti-Spike antibodies (anti-S1/S2 immunoglobulin G [IgG]). The median values of half-maximal neutralization titer (NT50 ) for the WT and Delta variants decreased by 75.8% and 82.2% at Visit 2 and by 85.4% and 81.4% at Visit 3, respectively. At Visit 1, the correlation between the anti-S1/S2 IgG and Nabs titers for the Delta variant was moderate for WT (r = 0.58) and weak for the Delta variant (r = 0.39). However, the correlation coefficient consistently remained above 0.7, with a very strong correlation at Visit 3 for both WT and Delta variants (r = 0.81). The dynamics of anti-S1/S2 IgG antibodies, NT50 , and cross-neutralization index correlated at different time points PSO. Longitudinal analysis of the cross-neutralization capacity of immune sera will inform upon the durability of the immune response against SARS CoV-2 variants.

Neutralizing and enhancing monoclonal antibodies in SARS-CoV-2 convalescent patients: lessons from early variant infection and impact on shaping emerging variants

ABSTRACT Serological studies of COVID-19 convalescent patients have identified polyclonal lineage-specific and cross-reactive antibodies (Abs), with varying effector functions against virus variants. Individual specificities of anti-SARS-CoV-2 Abs and their impact on infectivity by other variants have been little investigated to date. Here, we dissected at a monoclonal level neutralizing and enhancing Abs elicited by early variants and how they affect infectivity of emerging variants. B cells from 13 convalescent patients originally infected by D614G or Alpha variants were immortalized to isolate 445 naturally-produced anti-SARS-CoV-2 Abs. Monoclonal antibodies (mAbs) were tested for their abilities to impact the cytopathic effect of D614G, Delta, and Omicron (BA.1) variants. Ninety-eight exhibited robust neutralization against at least one of the three variant types, while 309 showed minimal or no impact on infectivity. Thirty-eight mAbs enhanced infectivity of SARS-CoV-2. Infection with D614G/Alpha variants generated variant-specific (65 neutralizing Abs, 35 enhancing Abs) and cross-reactive (18 neutralizing Abs, 3 enhancing Abs) mAbs. Interestingly, among the neutralizing mAbs with cross-reactivity restricted to two of the three variants tested, none demonstrated specific neutralization of the Delta and Omicron variants. In contrast, cross-reactive mAbs enhancing infectivity (n = 3) were found exclusively specific to Delta and Omicron variants. Notably, two mAbs that amplified in vitro the cytopathic effect of the Delta variant also exhibited neutralization against Omicron. These findings shed light on functional diversity of cross-reactive Abs generated during SARS-CoV-2 infection and illustrate how the balance between neutralizing and enhancing Abs facilitate variant emergence.

Association between tinnitus and anxiety in the convalescent COVID-19 patients: A single-center observational study

Association between tinnitus and anxiety in the convalescent COVID-19 patients: A single-center observational study

Rapid generation of human recombinant monoclonal antibodies from antibody-secreting cells using ferrofluid-based technology.

Monoclonal antibodies (mAbs) are one of the most important classes of biologics with high therapeutic and diagnostic value, but traditional methods for mAbs generation, such as hybridoma screening and phage display, have limitations, including low efficiency and loss of natural chain pairing. To overcome these challenges, novel single B cell antibody technologies have emerged, but they also have limitations such as in vitro differentiation of memory B cells and expensive cell sorters. In this study, we present a rapid and efficient workflow for obtaining human recombinant monoclonal antibodies directly from single antigen-specific antibody secreting cells (ASCs) in the peripheral blood of convalescent COVID-19 patients using ferrofluid technology. This process allows the identification and expression of recombinant antigen-specific mAbs in less than 10 days, using RT-PCR to generate linear Ig heavy and light chain gene expression cassettes, called "minigenes", for rapid expression of recombinant antibodies without cloning procedures. This approach has several advantages. First, it saves time and resources by eliminating the need for in vitro differentiation. It also allows individual antigen-specific ASCs to be screened for effector function prior to recombinant antibody cloning, enabling the selection of mAbs with desired characteristics and functional activity. In addition, the method allows comprehensive analysis of variable region repertoires in combination with functional assays to evaluate the specificity and function of the generated antigen-specific antibodies. Our approach, which rapidly generates recombinant monoclonal antibodies from single antigen-specific ASCs, could help to identify functional antibodies and deepen our understanding of antibody dynamics in the immune response through combined antibody repertoire sequence analysis and functional reactivity testing.

Evidence of a Sjögren's disease-like phenotype following COVID-19 in mice and humans.

Sjögren's Disease (SjD) is a systemic autoimmune disease characterized by lymphocytic inflammation of the lacrimal and salivary glands (SG), dry eyes and mouth, and systemic symptoms. SARS-CoV-2 may trigger the development or progression of autoimmune diseases. To test this, we used a mouse model of SARS-CoV-2 infection and convalescent patients' blood and SG in order to understand the development of SjD-like autoimmunity after infection. First, SARS-CoV-2-infected human angiotensin-converting enzyme 2 (ACE2) transgenic mice exhibited decreased salivation, elevated antinuclear antibodies (ANA), and lymphocytic infiltration in the lacrimal and SG. The sera from patients with COVID-19 sera showed increased ANA (i.e., anti-SSA [Sjögren's-syndrome-related antigen A]/anti-Ro52 and anti-SSB [SS-antigen B]/anti-La). Male patients showed elevated anti-SSA compared with female patients, and female patients exhibited diverse ANA patterns. SG biopsies from convalescent COVID-19 patients were microscopically similar to SjD SG with focal lymphocytic infiltrates in 4 of 6 patients and 2 of 6 patients exhibiting focus scores of at least 2. Lastly, monoclonal antibodies produced in recovered patients blocked ACE2/spike interaction and cross-reacted with nuclear antigens. Our study shows a direct association between SARS-CoV-2 and SjD. Hallmark features of SjD-affected SGs were histologically indistinguishable from convalescent COVID-19 patients. The results implicate that SARS-CoV-2 could be an environmental trigger for SjD.

Taste dysfunction after COVID-19: Analysis with functional near-infrared spectroscopy.

<br><b>Introduction:</b> According to official data, COVID-19 emerged in China in December 2019 and has spread worldwide since then.</br> <br><b>Aim:</b> The aim of this study was to investigate differences in functional near-infrared spectroscopy (fNIRS) recordings between convalescent COVID-19 patients and a healthy control group, which could help to clarify the pathomechanism of dysgeusia in COVID-19.</br> <br><b>Material and methods:</b> The study included 16 participants, comprised of 8 convalescent COVID-19 patients and 8 healthy controls. All participants were examined with fNIRS. The amplitude of changes in oxyhemoglobin (oxyHb) concentration in the cerebral cortex was analyzed statistically (for the test and control groups after stimulation with a taste stimulus - citric acid solution).</br> <br><b>Results:</b> The differences in the amplitude of changes in oxyHb concentration in the cerebral cortex were not statistically significant between the groups. Discussion: Using fNIRS, a strong stimulation of the visual cortex was discovered in response to the taste stimulus, consisting of large, repetitive changes in oxyHb concentration occurring in parallel with stimulation of areas of the taste cortex. This phenomenon has not, to our knowledge, been described previously in the scientific literature. The exact location of the primary taste cortex is controversial, but to date the occipital cortex has not been considered to be involved.</br> <br><b>Conclusions: No difference was observed in the dynamics of changes in oxyHb in the examined areas of the cerebral cortex between convalescent COVID-19 patients and healthy controls. However, the determination of the role of the occipital cortex in the perception of taste requires further research.</br&gt.

Hemostatic imbalance underlying preterm delivery in COVID-19 convalescent patients

Aim: to study the role of the hemostatic system in pretem delivery in pregnant women who have had COVID-19 in the gestation period from 14 to 16 weeks.Materials and Methods. A prospective single-center observational study was conducted by enrolling 63 pregnant women with verified COVID-19 at 14–16 weeks of gestation. The main group consisted of 37 patients with preterm birth (PB), comparison group – 26 patients labour activity that occurred at least at gestational age of 37 weeks. Clinical and anamnestic data and dynamic changes in fibrinogen and D-dimer level, activity of tissue factor (TF), tissue factor pathway inhibitor (TFPI), plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator (t-PA), urokinase plasminogen activator (u-PA) were analyzed; thrombin generation assay (TGA) was performed.Results. It was found that severity of COVID-19 infection did not determine the timing of delivery that depended on patient comorbid condition. All PB observations (37 out of 63, 58.7 %) were caused by decompensated placental function manifested by acute obstetrical complications: increasing intrauterine fetal hypoxia (64.9 %) along with intrauterine growth retardation (51.4 %), severe preeclampsia (13.5 %) and premature abruption of the normally located placenta (5.0 %). In both study groups, COVID-19 experienced at 14–16 weeks of pregnancy was associated with coagulation and fibrinolytic imbalances. At the same time, at least 6 weeks post-COVID-19 infection, patients with PB had higher level of the “Peak thrombin” vs. comparison group (3050 vs. 2527 pmol/L; p = 0.0433). Also, patients with term vs. preterm delivery had TF activity decreased significantly: by 47.1% and 28.1%, respectively (p = 0.0546). Patients in preterm delivery group were characterized by fibrinolytic imbalance. At the first time point, suppressed fibrinolysis (PAI-1 level – 18.4 vs. 12.5 ng/ml in the comparison group; p = 0.0209) was concomitant with elevated level of u-PA (1.5 vs. 0.55 ng/ml in comparison group, p = 0.0015), which suggests a potential prolonged immunoinflammatory response in patients with PB. Magnitude of fibrinogen concentration and D-dimer level during post-COVID-19 follow-up study was within the reference values specific to gestational age.Conclusion. A significant increase in coagulation potential was found and verified by elevated activity of tissue factor and potential to thrombin generation in COVID-19 convalescent patients. In the case of preterm delivery, there was an imbalance in fibrinolysis system revealed by decreased blood fibrinolytic activity elevating along with increasing gestational age.

Rapid transient and longer-lasting innate cytokine changes associated with adaptive immunity after repeated SARS-CoV-2 BNT162b2 mRNA vaccinations.

Cytokines and chemokines play an important role in shaping innate and adaptive immunity in response to infection and vaccination. Systems serology identified immunological parameters predictive of beneficial response to the BNT162b2 mRNA vaccine in COVID-19 infection-naïve volunteers, COVID-19 convalescent patients and transplant patients with hematological malignancies. Here, we examined the dynamics of the serum cytokine/chemokine responses after the 3rd BNT162b2 mRNA vaccination in a cohort of COVID-19 infection-naïve volunteers. We measured serum cytokine and chemokine responses after the 3rd dose of the BNT162b2 mRNA (Pfizer/BioNtech) vaccine in COVID-19 infection-naïve individuals by a chemiluminescent assay and ELISA. Anti-Spike binding antibodies were measured by ELISA. Anti-Spike neutralizing antibodies were measured by a pseudotype assay. Comparison to responses found after the 1st and 2nd vaccinations showed persistence of the coordinated responses of several cytokine/chemokines including the previously identified rapid and transient IL-15, IFN-γ, CXCL10/IP-10, TNF-α, IL-6 signature. In contrast to the transient (24hrs) effect of the IL-15 signature, an inflammatory/anti-inflammatory cytokine signature (CCL2/MCP-1, CCL3/MIP-1α, CCL4/MIP-1β, CXCL8/IL-8, IL-1Ra) remained at higher levels up to one month after the 2nd and 3rd booster vaccinations, indicative of a state of longer-lasting innate immune change. We also identified a systemic transient increase of CXCL13 only after the 3rd vaccination, supporting stronger germinal center activity and the higher anti-Spike antibody responses. Changes of the IL-15 signature, and the inflammatory/anti-inflammatory cytokine profile correlated with neutralizing antibody levels also after the 3rd vaccination supporting their role as immune biomarkers for effective development of vaccine-induced humoral responses. These data revealed that repeated SARS-Cov-2 BNT162b2 mRNA vaccination induces both rapid transient as well as longer-lasting systemic serum cytokine changes associated with innate and adaptive immune responses. Clinicaltrials.gov, identifier NCT04743388.

449. Elevated Lactate Dehydrogenase in Convalescent Severe COVID-19 Patients Reflects Delayed Clinical Recovery and Activated Th1 and Th17 Responses

Abstract Background Elevated lactate dehydrogenase (LD) levels during acute phase of COVID-19 is known to reflect multiple organ injuries and clinical severity. However, clinical implication and origin of elevated LD after convalescence have not been elucidated. Methods We prospectively enrolled non-vaccinated severe COVID-19 patients with an oxygen demand greater than FiO2 0.4 and followed them up to one year after discharge. Laboratory values and modified Medical Research Council (mMRC) dyspnea scale were collected at each outpatient visit. To investigate the potential association between elevated LD levels and CD4+T cell activities, fluorescence-activated cell sorting (FACS) and intracellular cytokine staining (ICS) of INF-γ, TNF-α, IL-4, and IL-17A were conducted. Results A total of 74 patients were included, of which 46 (62%) were male, the median age was 59 years (IQR 52–69), and peak FiO2 during hospitalization was 0.65 (IQR 0.50–0.80). At discharge, median absolute lymphocyte count (ALC) was 1.29 x103/μL (IQR 0.93–1.75), median C-reactive protein (CRP) was 0.23 mg/dL (IQR 0.06–0.52), and median LD was 334 IU/L (IQR 276–450). After discharge, ALC increased (R2 = 0.1434, P < 0.001) and LD decreased (R2 = 0.0838, P < 0.001) overtime, but CRP level did not show time-dependent changes (Figure 1A-C). Increased LD (R2 = 0.160, P < 0.001) and CRP (R2 = 0.028, P = 0.024) levels were significantly associated with higher grade of mMRC scales, while ALC was not associated with mMRC scale (Fig 1D-F). To investigate the potential association between elevated LD level and CD4+T cell activation, serial 38 PBMC specimens from 10 patients were further investigated. CD4+T cells were identified using FACS gating and then stained for intracellular cytokines. The proportion of TNF-α+CD4+T cells (R2 = 0.547, P < 0.001; Fig 2A) and IL-17A+CD4+T cells (R2 = 0.419, P = 0.009; Fig 2B) after spike protein stimulation were positively correlated with LD levels, while the proportion of Treg did not.Figure 1.Time-dependent changes in ALC, CRP and LD levels and their association with the degree of dyspnea (mMRC scale) after discharge.(A) Linear regression with ALC and time. (B) Linear regression with CRP and time. (C) Linear regression with LD and time. (D) Linear regression with ALC and mMRC. (E) Linear regression with CRP and mMRC. (F) Linear regression with LD and mMRC. *Statistically significant. ALC, absolute lymphocyte count; CRP, C-reactive protein; LD, lactate dehydrogenase; WBC, white blood cell, mMRC, modified Medical Research Council.Figure 2.FACS gating for CD4+T cells and ICS correlation with serum LD levels after convalescence.FACS gating for CD3+CD4+T cells and ICS for TNF-α (A) and IL-17A (B) were conducted. Serum LD levels after convalescence showed significant correlations with the proportion of TNF-α+CD4+T cells (A) and that of IL-17A+CD4+T cells (B). FACS, fluorescence-activated cell sorting; ICS, intracellular cytokine staining; LD, lactate dehydrogenase. Conclusion Elevated LD in convalescent severe COVID-19 patients reflects delayed clinical recovery and activated Th1 and Th17 responses Disclosures All Authors: No reported disclosures

Salivary IgA and vimentin differentiate in vitro SARS-CoV-2 infection: A study of 290 convalescent COVID-19 patients

SARS-CoV-2 initially infects cells in the nasopharynx and oral cavity. The immune system at these mucosal sites plays a crucial role in minimizing viral transmission and infection. To develop new strategies for preventing SARS-CoV-2 infection, this study aimed to identify proteins that protect against viral infection in saliva. We collected 551 saliva samples from 290 healthcare workers who had tested positive for COVID-19, before vaccination, between June and December 2020. The samples were categorized based on their ability to block or enhance infection using in vitro assays. Mass spectrometry and enzyme-linked immunosorbent assay experiments were used to identify and measure the abundance of proteins that specifically bind to SARS-CoV-2 antigens. Immunoglobulin (Ig)A specific to SARS-CoV-2 antigens was detectable in over 83% of the convalescent saliva samples. We found that concentrations of anti-receptor-binding domain IgA >500 pg/µg total protein in saliva correlate with reduced viral infectivity in vitro. However, there is a dissociation between the salivary IgA response to SARS-CoV-2, and systemic IgG titers in convalescent COVID-19 patients. Then, using an innovative technique known as spike-baited mass spectrometry, we identified novel spike-binding proteins in saliva, most notably vimentin, which correlated with increased viral infectivity in vitro and could serve as a therapeutic target against COVID-19.

Blood T cell phenotypes correlate with fatigue severity in post-acute sequelae of COVID-19.

Post-acute sequelae of COVID-19 (PASC) affect approximately 10% of convalescent patients. The spectrum of symptoms is broad and heterogeneous with fatigue being the most often reported sequela. Easily accessible blood biomarkers to determine PASC severity are lacking. Thus, our study aimed to correlate immune phenotypes with PASC across the severity spectrum of COVID-19. A total of 176 originally immunonaïve, convalescent COVID-19 patients from a prospective cohort during the first pandemic phase were stratified by initial disease severity and underwent clinical, psychosocial, and immune phenotyping around 10weeks after first COVID-19 symptoms. COVID-19-associated fatigue dynamics were assessed and related to clinical and immune phenotypes. Fatigue and severe fatigue were commonly reported irrespective of initial COVID-19 severity or organ-specific PASC. A clinically relevant increase in fatigue severity after COVID-19 was detected in all groups. Neutralizing antibody titers were higher in patients with severe acute disease, but no association was found between antibody titers and PASC. While absolute peripheral blood immune cell counts in originally immunonaïve PASC patients did not differ from unexposed controls, peripheral CD3+CD4+ T cell counts were independently correlated with fatigue severity across all strata in multivariable analysis. Patients were at similar risk of self-reported PASC irrespective of initial disease severity. The independent correlation between fatigue severity and blood T cell phenotypes indicates a possible role of CD4+ T cells in the pathogenesis of post-COVID-19 fatigue, which might serve as a blood biomarker.

Gag Virus-like Particles Functionalized with SARS-CoV-2 Variants: Generation, Characterization and Recognition by COVID-19 Convalescent Patients' Sera.

The robustness, safety, versatility, and high immunogenicity of virus-like particles (VLPs) make them a promising approach for the generation of vaccines against a broad range of pathogens. VLPs are recombinant macromolecular structures that closely mimic the native conformation of viruses without carrying viral genetic material. Particularly, HIV-1 Gag-based VLPs are a suitable platform for the presentation of the SARS-CoV-2 Spike (S) protein on their surface. In this context, this work studies the effect of different rationally engineered mutations of the S protein to improve some of its characteristics. The studied variants harbored mutations such as proline substitutions for S stabilization, D614G from the early dominant pandemic form, the elimination of the S1/S2 furin cleavage site to improve S homogeneity, the suppression of a retention motif to favor its membrane localization, and cysteine substitutions to increase its immunogenicity and avoid potential undesired antibody-dependent enhancement (ADE) effects. The influence of the mutations on VLP expression was studied, as well as their immunogenic potential, by testing the recognition of the generated VLP variants by COVID-19 convalescent patients' sera. The results of this work are conceived to give insights on the selection of S protein candidates for their use as immunogens and to showcase the potential of VLPs as carriers for antigen presentation.

Presence of Elevated Interleukin – 6 (IL-6) in the Blood of COVID-19 Convalescent Patients (40 – 93 Days) Post Onset of Symptoms Could be an Indicator of Ongoing Activation of the Immune System

IL-6 concentrations rise with the onset of COVID-19 infection and is detected in 68% of patients on admission but is expected to reduce after the acute phase. IL-6 concentrations at time points: (2-7 days), (6-11 days), (11-15 days) and (13-20 days) after intensive care unit (ICU) admission showed the highest level of IL-6 concentrations at time point 2-7 days, we decided to characterize IL-6 concentrations in serum samples collected in the sera of COVID-19 convalescent patients (40 – 93 days) post onset of symptoms. The estimated IL- 6 concentrations detected ranged between 2.65 pg/ml and 29.01 pg/ml, with a median of 9.18 pg/ml. Nine out of the 28 (32.14%) patients had normal IL-6 levels (<7 pg/ml) and 19 of the 28 patients (68%) had IL-6 elevated concentrations (>7 pg/ml). The IL-6 concentrations were grouped normal (<7 pg.ml, n= 9), mildly elevated (7 – 10 pg/ml, n = 10) and elevated (>10 pg/ml n = 9). There was no relationship between IL-6 concentrations with gender, days from onset of symptoms, days from subsiding symptoms and COVID-19 IgG antibodies. There was some relationship with age, as the younger patient cohort produced much higher IL-6 concentrations than the older patient cohort. This study describes an increase in blood IL-6 concentrations in the sera of COVID-19 convalescent patients (40 – 93 days) post onset of symptoms like that seen in COVID patients on admission and higher levels seen in the younger patient cohort indicating that “cytokine storms” still occur in the recovery phase.

PB0065 Targeting the Endothelium when Treating Convalescent COVID-19 Patients, the Effect on Serum Biomarkers, and Post-COVID Functional Status. Midterm Results of a Randomized, Placebo-Controlled Trial

PB0065 Targeting the Endothelium when Treating Convalescent COVID-19 Patients, the Effect on Serum Biomarkers, and Post-COVID Functional Status. Midterm Results of a Randomized, Placebo-Controlled Trial

Interpretable machine-learning model for Predicting the Convalescent COVID-19 patients with pulmonary diffusing capacity impairment

IntroductionThe COVID-19 patients in the convalescent stage noticeably have pulmonary diffusing capacity impairment (PDCI). The pulmonary diffusing capacity is a frequently-used indicator of the COVID-19 survivors’ prognosis of pulmonary function, but the current studies focusing on prediction of the pulmonary diffusing capacity of these people are limited. The aim of this study was to develop and validate a machine learning (ML) model for predicting PDCI in the COVID-19 patients using routinely available clinical data, thus assisting the clinical diagnosis.MethodsCollected from a follow-up study from August to September 2021 of 221 hospitalized survivors of COVID-19 18 months after discharge from Wuhan, including the demographic characteristics and clinical examination, the data in this study were randomly separated into a training (80%) data set and a validation (20%) data set. Six popular machine learning models were developed to predict the pulmonary diffusing capacity of patients infected with COVID-19 in the recovery stage. The performance indicators of the model included area under the curve (AUC), Accuracy, Recall, Precision, Positive Predictive Value(PPV), Negative Predictive Value (NPV) and F1. The model with the optimum performance was defined as the optimal model, which was further employed in the interpretability analysis. The MAHAKIL method was utilized to balance the data and optimize the balance of sample distribution, while the RFECV method for feature selection was utilized to select combined features more favorable to machine learning.ResultsA total of 221 COVID-19 survivors were recruited in this study after discharge from hospitals in Wuhan. Of these participants, 117 (52.94%) were female, with a median age of 58.2 years (standard deviation (SD) = 12). After feature selection, 31 of the 37 clinical factors were finally selected for use in constructing the model. Among the six tested ML models, the best performance was accomplished in the XGBoost model, with an AUC of 0.755 and an accuracy of 78.01% after experimental verification. The SHAPELY Additive explanations (SHAP) summary analysis exhibited that hemoglobin (Hb), maximal voluntary ventilation (MVV), severity of illness, platelet (PLT), Uric Acid (UA) and blood urea nitrogen (BUN) were the top six most important factors affecting the XGBoost model decision-making.ConclusionThe XGBoost model reported here showed a good prognostic prediction ability for PDCI of COVID-19 survivors during the recovery period. Among the interpretation methods based on the importance of SHAP values, Hb and MVV contributed the most to the prediction of PDCI outcomes of COVID-19 survivors in the recovery period.

Recovery of Endothelium-dependent vascular relaxation impairment in convalescent COVID-19 patients: Insight from a pilot study

BackgroundEndothelial dysfunction is a key-feature in acute COVID-19. However, follow-up data regarding endothelial dysfunction and injury after COVID-19 infection are lacking. We aimed to investigate the changes in endothelium-dependent vasorelaxation at baseline and four months after hospital discharge in COVID-19 patients. MethodsTwenty COVID-19 patients were compared to 24 healthy controls. Clinical and morphological data were collected after hospital admission for SARS-CoV-2 infection and reactive hyperaemia index (RHI) measurement was performed with a delay between 24 and 48 h after hospital admission and four months after hospital discharge in the outpatient clinics. Blood tests including inflammatory markers and measurement of post-occlusive vasorelaxation by digital peripheral arterial tonometry were performed at both visits. ResultsAt baseline, COVID-19 patients exhibited reduced RHI compared to controls (p < 0.001), in line with an endothelial dysfunction. At four months follow-up, there was a 51% increase in the RHI (1.69 ± 0.32 to 2.51 ± 0.91; p < 0.01) in favor of endothelium-dependent vascular relaxation recovery. RHI changes were positively correlated with baseline C-reactive protein (r = 0.68; p = 0.02). Compared to COVID-19 patients with a decrease in RHI, COVID-19 patients with an increase in RHI beyond the day-to-day variability (i.e. >11%) had less severe systemic inflammation at baseline. ConclusionConvalescent COVID-19 patients showed a recovery of systemic artery endothelial dysfunction, in particular patients with lower inflammation at baseline. Further studies are needed to decipher the interplay between inflammation and endothelial dysfunction in COVID-19 patients.

HLA-E-restricted SARS-CoV-2-specific T cells from convalescent COVID-19 patients suppress virus replication despite HLA class Ia down-regulation.

Pathogen-specific CD8+ T cell responses restricted by the nonpolymorphic nonclassical class Ib molecule human leukocyte antigen E (HLA-E) are rarely reported in viral infections. The natural HLA-E ligand is a signal peptide derived from classical class Ia HLA molecules that interact with the NKG2/CD94 receptors to regulate natural killer cell functions, but pathogen-derived peptides can also be presented by HLA-E. Here, we describe five peptides from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that elicited HLA-E-restricted CD8+ T cell responses in convalescent patients with coronavirus disease 2019. These T cell responses were identified in the blood at frequencies similar to those reported for classical HLA-Ia-restricted anti-SARS-CoV-2 CD8+ T cells. HLA-E peptide-specific CD8+ T cell clones, which expressed diverse T cell receptors, suppressed SARS-CoV-2 replication in Calu-3 human lung epithelial cells. SARS-CoV-2 infection markedly down-regulated classical HLA class I expression in Calu-3 cells and primary reconstituted human airway epithelial cells, whereas HLA-E expression was not affected, enabling T cell recognition. Thus, HLA-E-restricted T cells could contribute to the control of SARS-CoV-2 infection alongside classical T cells.

Antioxidant Genetic Variants Modify Echocardiography Indices in Long COVID.

Although disturbance of redox homeostasis might be responsible for COVID-19 cardiac complications, this molecular mechanism has not been addressed yet. We have proposed modifying the effects of antioxidant proteins polymorphisms (superoxide dismutase 2 (SOD2), glutathione peroxidase 1 (GPX1), glutathione peroxidase 3 (GPX3) and nuclear factor erythroid 2-related factor 2, (Nrf2)) in individual susceptibility towards the development of cardiac manifestations of long COVID-19. The presence of subclinical cardiac dysfunction was assessed via echocardiography and cardiac magnetic resonance imaging in 174 convalescent COVID-19 patients. SOD2, GPX1, GPX3 and Nrf2 polymorphisms were determined via the appropriate PCR methods. No significant association of the investigated polymorphisms with the risk of arrhythmia development was found. However, the carriers of variant GPX1*T, GPX3*C or Nrf2*A alleles were more than twice less prone for dyspnea development in comparison with the carriers of the referent ones. These findings were even more potentiated in the carriers of any two variant alleles of these genes (OR = 0.273, and p = 0.016). The variant GPX alleles were significantly associated with left atrial and right ventricular echocardiographic parameters, specifically LAVI, RFAC and RV-EF (p = 0.025, p = 0.009, and p = 0.007, respectively). Based on the relation between the variant SOD2*T allele and higher levels of LV echocardiographic parameters, EDD, LVMI and GLS, as well as troponin T (p = 0.038), it can be proposed that recovered COVID-19 patients, who are the carriers of this genetic variant, might have subtle left ventricular systolic dysfunction. No significant association between the investigated polymorphisms and cardiac disfunction was observed when cardiac magnetic resonance imaging was performed. Our results on the association between antioxidant genetic variants and long COVID cardiological manifestations highlight the involvement of genetic propensity in both acute and long COVID clinical manifestations.