Lack of a prompt normalization of immunological parameters is associated with long-term care and poor prognosis in COVID-19 affected patients receiving convalescent plasma: a single center experience.

OBJECTIVES: Being COVID-19 convalescent plasma (CCP) a therapeutic option that can have a potential impact on the normalization of immunological parameters of COVID-19 affected patients, a detailed analysis of post-infusion immunological changes was conducted in CCP treated patients, aiming to identify possible predictive hallmarks of disease prognosis. METHODS: This prospective observational study describes a cohort of 28 patients who received CCP shortly after being hospitalized for COVID-19 and diagnosed for Acute Respiratory Distress Syndrome. All patients were subjected to a detailed flow cytometry based evaluation of immunological markers at baseline and on days +3 and +7 after transfusion. RESULTS: At baseline almost all patients suffered from lymphopenia (25/28 on T-cells and 16/28 on B-cells) coupled with neutrophil-lymphocyte ratio exceeding normal values (26/28). Lymphocyte subsets were generally characterized by increased percentages of CD19+CD20-CD38hiCD27+ plasmablasts and reduction of CD4+CD45RA+CCR7+CD31+ recent thymic emigrants, while monocytes presented a limited expression of CD4 and HLA-DR molecules. Amelioration of immunological parameters began to be evident from day +3 and became more significant at day +7 post-CCP transfusion in 18 patients who recovered within 30 days from hospitalization. Conversely, baseline immunological characteristics generally persisted in ten critical patients who eventually progressed to death (6) or long-term care (4). CONCLUSIONS: This study demonstrates that proper immunophenotyping panels can be potentially useful for monitoring CCP treated patients from the first days after infusion in order to presume higher risk of medical complications.

The unreversible reduced but persistent activated NK and CD8+ T cells in severe/critical COVID-19 during omicron pandemic in China.

As a hallmark of COVID-19 progression, lymphopenia alongside its subtle immune disturbance has been widely reported, but yet to be thoroughly elucidated. Aiming at exploring clinical immune biomarkers with accessibility in the current and acute omicron epidemic abrupted in China post-control era, we design a real-world prospective observation cohort in Peking Union Medical College Hospital to describe immunological, haematological profiles inducing lymphocyte subsets related to SARS-CoV-2 infection. In this COVID-19 cohort, we enrolled 17 mild/moderate (M/M), 24 severe (S) and 25 critical (C) patients. The dynamics of lymphocytes of COVID-19 demonstrated that the sharp decline of NK, CD8+, and CD4+ T cell counts was the main contributor to lymphopenia in the S/C group, compared to the M/M group. Expressions of activation marker CD38 and proliferation marker Ki-67 both in CD8+ T and NK cells were significantly higher in all COVID-19 patients than that in healthy donors, independent of disease severity. The subsequent analysis showed in contrast to the M/M group, NK and CD8+ T cell counts remained low-level after therapy in the S/C group. CD38 and Ki-67 expressions in NK and CD8+ T cells still stay at a high level, despite active treatment. Targeting relatively elderly patients with SARS-CoV-2 infection, severe COVID-19 features the unreversible reduction of NK and CD8+ T cells with persistent activation and proliferation, which assist clinicians in early recognizing and saving severe or critical COVID-19 patients. Given that immunophenotype, the new immunotherapy improving NK and CD8+ T lymphocyte antiviral efficiency should be considered.

Immunophenotype of Measurable Residual Blast Cells as an Additional Prognostic Factor in Adults with B-Cell Acute Lymphoblastic Leukemia.

Measurable residual disease (MRD) is a well-known independent prognostic factor in acute leukemias, and multicolor flow cytometry (MFC) is widely used to detect MRD. MFC is able not only to enumerate MRD accurately but also to describe an antigen expression profile of residual blast cells. However, the relationship between MRD immunophenotype and patient survival probability has not yet been studied. We determined the prognostic impact of MRD immunophenotype in adults with B-cell acute lymphoblastic leukemia (B-ALL). In a multicenter study RALL-2016 (NCT03462095), 267 patients were enrolled from 2016 to 2022. MRD was assessed at the end of induction (day 70) in 94 patients with B-ALL by six- or 10-color flow cytometry in the bone marrow specimens. The 4 year relapse-free survival (RFS) was lower in MRD-positive B-ALL patients [37% vs. 78% (p < 0.0001)]. The absence of CD10, positive expression of CD38, and high expression of CD58 on MRD cells worsened the 4 year RFS [19% vs. 51% (p = 0.004), 0% vs. 51% (p < 0.0001), and 21% vs. 40% (p = 0.02), respectively]. The MRD immunophenotype is associated with RFS and could be an additional prognostic factor for B-ALL patients.

Immune Response after COVID-19 mRNA Vaccination in Multiple Sclerosis Patients Treated with DMTs.

The impact of disease-modifying therapies (DMTs) on the immune response to coronavirus disease-2019 (COVID-19) vaccines in persons with multiple sclerosis (pwMS) needs further elucidation. We investigated BNT162b2 mRNA COVID-19 vaccine effects concerning antibody seroconversion, inflammatory mediators' level and immunophenotype assessment in pwMS treated with cladribine (c-pwMS, n = 29), fingolimod (f-pwMS, n = 15) and ocrelizumab (o-pwMS, n = 54). Anti-spike immunoglobulin (Ig)-G detection was performed by an enzyme immunoassay; molecular mediators (GrB, IFN-γ and TNF-α) were quantified using the ELLA platform, and immunophenotype was assessed by flow cytometry. ANCOVA, Student's t-test and Pearson correlation analyses were applied. Only one o-pwMS showed a mild COVID-19 infection despite most o-pwMS lacking seroconversion and showing lower anti-spike IgG titers than c-pwMS and f-pwMS. No significant difference in cytokine production and lymphocyte count was observed in c-pwMS and f-pwMS. In contrast, in o-pwMS, a significant increase in GrB levels was detected after vaccination. Considering non-seroconverted o-pwMS, a significant increase in GrB serum levels and CD4+ T lymphocyte count was found after vaccination, and a negative correlation was observed between anti-spike IgG production and CD4+ T cells count. Differences in inflammatory mediators' production after BNT162b2 vaccination in o-pwMS, specifically in those lacking anti-spike IgG, suggest a protective cellular immune response.

Differences in B-Cell Immunophenotypes and Neutralizing Antibodies Against SARS-CoV-2 After Administration of BNT162b2 (Pfizer-BioNTech) Vaccine in Individuals with and without Prior COVID-19 - A Prospective Cohort Study.

Purpose: Understanding the humoral immune response dynamics carried out by B cells in COVID-19 vaccination is little explored; therefore, we analyze the changes induced in the different cellular subpopulations of B cells after vaccination with BNT162b2 (Pfizer-BioNTech). Methods: This prospective cohort study evaluated thirty-nine immunized health workers (22 with prior COVID-19 and 17 without prior COVID-19) and ten subjects not vaccinated against SARS-CoV-2 (control group). B cell subpopulations (transitional, mature, naïve, memory, plasmablasts, early plasmablast, and double-negative B cells) and neutralizing antibody levels were analyzed and quantified by flow cytometry and ELISA, respectively. Results: The dynamics of the B cells subpopulations after vaccination showed the following pattern: the percentage of transitional B cells was higher in the prior COVID-19 group (p < 0.05), whereas virgin B cells were more prevalent in the group without prior COVID-19 (p < 0.05), mature B cells predominated in both vaccinated groups (p < 0.01), and memory B cells, plasmablasts, early plasmablasts, and double-negative B cells were higher in the not vaccinated group (p < 0.05). Conclusion: BNT162b2 vaccine induces changes in B cell subpopulations, especially generating plasma cells and producing neutralizing antibodies against SARS-CoV-2. However, the previous infection with SARS-CoV-2 does not significantly alter the dynamics of these subpopulations but induces more rapid and optimal antibody production.

Programmed Cell Death-1/Programmed Cell Death-1 Ligand as Prognostic Markers of Coronavirus Disease 2019 Severity.

Current research proves that immune dysregulation is a common feature of coronavirus disease 2019 (COVID-19), and immune exhaustion is associated with increased disease mortality. Immune checkpoint molecules, including the programmed cell death-1 (PD-1)/PD-1 ligand (PD-L1) axis, may serve as markers of disease severity. Accordingly, in this study, we evaluated the expression of PD-1/PD-L1 in patients with COVID-19. Blood immunophenotypes of hospitalized patients with moderate (n = 17, requiring oxygen support) and severe (n = 35, requiring mechanical ventilation in the intensive care setting) COVID-19 were compared and associated with clinical, laboratory, and survival data. The associations between severity and lymphocyte profiles were analysed at baseline and after 7 and 14 days of in-hospital treatment. Forty patients without COVID-19 infection were used as controls. For PD-1-positive T and B lymphocyte subsets, notable increases were observed between controls and patients with moderate or severe COVID-19 for CD4+PD-1+ T cells, CD8+PD-1+ T and CD19+PD-1+ B cells. Similar trends were observed for PD-L1-positive lymphocytes, namely, CD4+PD-L1+ T cells, CD8+PD-L1+ T cells and CD19+PD-L1+ B cells. Importantly, all markers associated with PD-1 and PD-L1 were stable over time for the analysed time points in the moderate and severe COVID-19 groups. Increased abundances of PD-1+ and PD-L1+ lymphocytes were associated with disease severity and mortality and were stable over time in patients with moderate to severe COVID-19. These immune exhaustion parameters may be attractive biomarkers of COVID-19 severity.

The Evolution of Blood Cell Phenotypes, Intracellular and Plasma Cytokines and Morphological Changes in Critically Ill COVID-19 Patients.

BACKGROUND: Severe coronavirus disease 2019 (COVID-19) causes a strong inflammatory response. To obtain an overview of inflammatory mediators and effector cells, we studied 25 intensive-care-unit patients during the timeframe after off-label chloroquine treatment and before an introduction of immunomodulatory drugs. MATERIAL AND METHODS: Blood samples were weekly examined with flow cytometry (FCM) for surface and intracytoplasmic markers, cytokine assays were analyzed for circulating interleukins (ILs), and blood smears were evaluated for morphological changes. Samples from healthy volunteers were used for comparison. Organ function data and 30-day mortality were obtained from medical records. RESULTS: Compared to that of the healthy control group, the expression levels of leukocyte surface markers, i.e., the cluster of differentiation (CD) markers CD2, CD4, CD8, CD158d, CD25, CD127, and CD19, were lower (p < 0.001), while those of leukocytes expressing CD33 were increased (p < 0.05). An aberrant expression of CD158d on granulocytes was found on parts of the granulocyte population. The expression levels of intracellular tumor necrosis factor alpha (TNFα) and IL-1 receptor type 2 in leukocytes were higher (p < 0.001) as well as plasma levels of TNFα, IL-2, IL-6, IL-8, IL-10 (p < 0.001), interferon gamma (IFNγ) (p < 0.01), and granulocyte-macrophage colony-stimulating factor (GM-CSF) (p < 0.05). The expression levels of CD33+ leukocytes and circulating IL-6 were higher (p < 0.05) among patients with arterial oxygen partial pressure-to-fractional inspired oxygen (PaO2/FiO2) ratios below 13.3 kPa compared to in the remaining patients. The expression levels of TNFα, IL-2, IL-4, IL-6, IL-8, and IL-10 were higher in patients treated with continuous renal replacement therapy (CRRT) (p < 0.05), and the levels of the maximum plasma creatinine and TNFα Spearman's rank-order correlation coefficient (rho = 0.51, p < 0.05) and IL-8 (rho = 0.44, p < 0.05) correlated. Blood smears revealed neutrophil dysplasia with pseudo-Pelger forms being most common. CONCLUSION: These findings suggest that patients with severe COVID-19, in addition to augmented ILs, lymphopenia, and increased granulocytes, also had effects on the bone marrow.

Regulatory T Cells as Predictors of Clinical Course in Hospitalised COVID-19 Patients.

Background: The host immune response has a prominent role in the progression and outcome of SARS-CoV-2 infection. Lymphopenia has been described as an important feature of SARS-CoV-2 infection and has been associated with severe disease manifestation. Lymphocyte dysregulation and hyper-inflammation have been shown to be associated with a more severe clinical course; however, a T cell subpopulation whose dysfunction correlate with disease progression has yet to be identify. Methods: We performed an immuno-phenotypic analysis of T cell sub-populations in peripheral blood from patients affected by different severity of COVID-19 (n=60) and undergoing a different clinical evolution. Clinical severity was established based on a modified WHO score considering both ventilation support and respiratory capacity (PaO2/FiO2 ratio). The ability of circulating cells at baseline to predict the probability of clinical aggravation was explored through multivariate regression analyses. Results: The immuno-phenotypic analysis performed by multi-colour flow cytometry confirmed that patients suffering from severe COVID-19 harboured significantly reduced circulating T cell subsets, especially for CD4+ T, Th1, and regulatory T cells. Peripheral T cells also correlated with parameters associated with disease severity, i.e., PaO2/FiO2 ratio and inflammation markers. CD4+ T cell subsets showed an important significant association with clinical evolution, with patients presenting markedly decreased regulatory T cells at baseline having a significantly higher risk of aggravation. Importantly, the combination of gender and regulatory T cells allowed distinguishing between improved and worsened patients with an area under the ROC curve (AUC) of 82%. Conclusions: The present study demonstrates the association between CD4+ T cell dysregulation and COVID-19 severity and progression. Our results support the importance of analysing baseline regulatory T cell levels, since they were revealed able to predict the clinical worsening during hospitalization. Regulatory T cells assessment soon after hospital admission could thus allow a better clinical stratification and patient management.

Lasting Changes to Circulating Leukocytes in People with Mild SARS-CoV-2 Infections.

Survivors of severe SARS-CoV-2 infections frequently suffer from a range of post-infection sequelae. Whether survivors of mild or asymptomatic infections can expect any long-term health consequences is not yet known. Herein we investigated lasting changes to soluble inflammatory factors and cellular immune phenotype and function in individuals who had recovered from mild SARS-CoV-2 infections (n = 22), compared to those that had recovered from other mild respiratory infections (n = 11). Individuals who had experienced mild SARS-CoV-2 infections had elevated levels of C-reactive protein 1-3 months after symptom onset, and changes in phenotype and function of circulating T-cells that were not apparent in individuals 6-9 months post-symptom onset. Markers of monocyte activation, and expression of adherence and chemokine receptors indicative of altered migratory capacity, were also higher at 1-3 months post-infection in individuals who had mild SARS-CoV-2, but these were no longer elevated by 6-9 months post-infection. Perhaps most surprisingly, significantly more T-cells could be activated by polyclonal stimulation in individuals who had recently experienced a mild SARS-CoV-2, infection compared to individuals with other recent respiratory infections. These data are indicative of prolonged immune activation and systemic inflammation that persists for at least three months after mild or asymptomatic SARS-CoV-2 infections.

Transfer transcriptomic signatures for infectious diseases.

The modulation of the transcriptome is among the earliest responses to infection. However, defining the transcriptomic signatures of disease is challenging because logistic, technical, and cost factors limit the size and representativeness of samples in clinical studies. These limitations lead to a poor performance of signatures when applied to new datasets. Although the study focuses on infection, the central hypothesis of the work is the generalization of sets of signatures across diseases. We use a machine learning approach to identify common elements in datasets and then test empirically whether they are informative about a second dataset from a disease or process distinct from the original dataset. We identify sets of genes, which we name transfer signatures, that are predictive across diverse datasets and/or species (e.g., rhesus to humans). We demonstrate the usefulness of transfer signatures in two use cases: the progression of latent to active tuberculosis and the severity of COVID-19 and influenza A H1N1 infection. This indicates that transfer signatures can be deployed in settings that lack disease-specific biomarkers. The broad significance of our work lies in the concept that a small set of archetypal human immunophenotypes, captured by transfer signatures, can explain a larger set of responses to diverse diseases.

Transcriptional Differences for COVID-19 Disease Map Genes between Males and Females Indicate a Different Basal Immunophenotype Relevant to the Disease.

Worldwide COVID-19 epidemiology data indicate differences in disease incidence amongst sex and gender demographic groups. Specifically, male patients are at a higher death risk than female patients, and the older population is significantly more affected than young individuals. Whether this difference is a consequence of a pre-existing differential response to the virus, has not been studied in detail. We created DeCovid, an R shiny app that combines gene expression (GE) data of different human tissue from the Genotype-Tissue Expression (GTEx) project along with the COVID-19 Disease Map and COVID-19 related pathways gene collections to explore basal GE differences across healthy demographic groups. We used this app to study differential gene expression of COVID-19 associated genes in different age and sex groups. We identified that healthy women show higher expression-levels of interferon genes. Conversely, healthy men exhibit higher levels of proinflammatory cytokines. Additionally, young people present a stronger complement system and maintain a high level of matrix metalloproteases than older adults. Our data suggest the existence of different basal immunophenotypes amongst different demographic groups, which are relevant to COVID-19 progression and may contribute to explaining sex and age biases in disease severity. The DeCovid app is an effective and easy to use tool for exploring the GE levels relevant to COVID-19 across demographic groups and tissues.

Definition and Risks of Cytokine Release Syndrome in 11 Critically Ill COVID-19 Patients With Pneumonia: Analysis of Disease Characteristics.

Corona virus disease 2019 (COVID-19) patients with severe immune abnormalities are at risk of cytokine release syndrome (CRS). The definition, prevention, and treatment of symptoms of CRS in critically ill patients with COVID-19 are important problems. We report a single-center case series of 11 COVID-19 patients with acute respiratory distress syndrome from The First Affiliated Hospital of Guangzhou Medical University in China from 26 January 2020 to 18 February 2020. The termination date of follow-up was 19 February 2020. Eight patients were determined to have characteristics of CRS, including pulmonary inflammation, fever, and dysfunction of nonpulmonary organs. An increase in interleukin-6 in peripheral blood was the highest risk factor and an early indicator of CRS in COVID-19.