Understanding innate immune responses in COVID-19 is important to decipher mechanisms of host responses and interpret disease pathogenesis. Natural killer (NK) cells are innate effector lymphocytes ...that respond to acute viral infections but might also contribute to immunopathology. Using 28-color flow cytometry, we here reveal strong NK cell activation across distinct subsets in peripheral blood of COVID-19 patients. This pattern was mirrored in scRNA-seq signatures of NK cells in bronchoalveolar lavage from COVID-19 patients. Unsupervised high-dimensional analysis of peripheral blood NK cells furthermore identified distinct NK cell immunotypes that were linked to disease severity. Hallmarks of these immunotypes were high expression of perforin, NKG2C, and Ksp37, reflecting increased presence of adaptive NK cells in circulation of patients with severe disease. Finally, arming of CD56
NK cells was observed across COVID-19 disease states, driven by a defined protein-protein interaction network of inflammatory soluble factors. This study provides a detailed map of the NK cell activation landscape in COVID-19 disease.
Proteomics is an emerging field in the study of joint disease. Our two aims with this pilot analysis were to compare healthy human knee articular cartilage with meniscus, two tissues both known to ...become affected in the osteoarthritic disease process, and to compare two mass spectrometry (MS)-based methods: data-dependent acquisition (DDA) and data-independent acquisition (DIA).
Healthy knee articular cartilage taken from the medial tibial condyle and medial meniscus samples taken from the body region were obtained from three adult forensic medicine cases. Proteins were extracted from tissue pieces and prepared for MS analysis. Each sample was subjected to liquid chromatography (LC)-MS/MS analysis using an Orbitrap mass spectrometer, and run in both DDA and DIA mode. Linear mixed effects models were used for statistical analysis.
A total of 653 proteins were identified in the DDA analysis, of which the majority was present in both tissue types. Only proteins with quantitation information in both tissues (n = 90) were selected for more detailed analysis, of which the majority did not statistically significantly differ in abundance between the two tissue types, in either of the MS analyses. However, 21 proteins were statistically significantly different (p < 0.05) between meniscus and cartilage in the DIA analysis. Out of these, 11 proteins were also significantly different in the DDA analysis. Aggrecan core protein was the most abundant protein in articular cartilage and significantly differed between the two tissues in both methods. The corresponding protein in meniscus was serum albumin. Dermatopontin exhibited the highest meniscus vs articular cartilage ratio among the statistically significant proteins. The DIA method led to narrower confidence intervals for the abundance differences between the two tissue types than DDA.
Although articular cartilage and meniscus had similar proteomic composition, we detected several differences by MS. Between the two analyses, DIA yielded more precise estimates and more statistically significant different proteins than DDA, and had no missing values, which makes it preferable for future LC-MS/MS analyses.
Upon infection with
(Mtb) the host immune response might clear the bacteria, control its growth leading to latent tuberculosis (LTB), or fail to control its growth resulting in active TB (ATB). There ...is however no clear understanding of the features underlying a more or less effective response. Mtb glycolipids are abundant in the bacterial cell envelope and modulate the immune response to Mtb, but the patterns of response to glycolipids are still underexplored. To identify the CD45
leukocyte activation landscape induced by Mtb glycolipids in peripheral blood of ATB and LTB, we performed a detailed assessment of the immune response of PBMCs to the Mtb glycolipids lipoarabinomannan (LAM) and its biosynthetic precursor phosphatidyl-inositol mannoside (PIM), and purified-protein derivate (PPD). At 24 h of stimulation, cell profiling and secretome analysis was done using mass cytometry and high-multiplex immunoassay. PIM induced a diverse cytokine response, mainly affecting antigen-presenting cells to produce both pro-inflammatory and anti-inflammatory cytokines, but not IFN-γ, contrasting with PPD that was a strong inducer of IFN-γ. The effect of PIM on the antigen-presenting cells was partly TLR2-dependent. Expansion of monocyte subsets in response to PIM or LAM was reduced primarily in LTB as compared to healthy controls, suggesting a hyporesponsive/tolerance pattern derived from Mtb infection.
Annually, approximately 10 million people are diagnosed with active tuberculosis (TB), and 1.4 million die of the disease. If left untreated, each person with active TB will infect 10–15 new ...individuals. The lack of non-sputum-based diagnostic tests leads to delayed diagnoses of active pulmonary TB cases, contributing to continued disease transmission. In this exploratory study, we aimed to identify biomarkers associated with active TB. We assessed the plasma levels of 92 proteins associated with inflammation in individuals with active TB (n = 20), latent TB (n = 14), or healthy controls (n = 10). Using co-expression network analysis, we identified one module of proteins with strong association with active TB. We removed proteins from the module that had low abundance or were associated with non-TB diseases in published transcriptomic datasets, resulting in a 12-protein plasma signature that was highly enriched in individuals with pulmonary and extrapulmonary TB and was further associated with disease severity.
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•We identified a 12-protein signature highly associated with active tuberculosis (TB)•The signature was correlated with TB disease severity and inflammation•The signature was enriched in active TB compared with other diseases•IFNG, IL-6, and VEGFA of the signature have previously been associated with active TB
Clinical finding; Disease; Proteomics; Infectious disease
Objectives
The role of innate lymphoid cells (ILCs) in coronavirus disease 2019 (COVID‐19), caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), is unknown. Understanding the ...immune response in COVID‐19 could contribute to unravel the pathogenesis and identification of treatment targets. Here, we describe the phenotypic landscape of circulating ILCs in COVID‐19 patients and identified ILC phenotypes correlated to serum biomarkers, clinical markers and laboratory parameters relevant in COVID‐19.
Methods
Blood samples collected from moderately (n = 11) and severely ill (n = 12) COVID‐19 patients, as well as healthy control donors (n = 16), were analysed with 18‐parameter flow cytometry. Using supervised and unsupervised approaches, we examined the ILC activation status and homing profile. Clinical and laboratory parameters were obtained from all COVID‐19 patients, and serum biomarkers were analysed with multiplex immunoassays.
Results
Innate lymphoid cells were largely depleted from the circulation of COVID‐19 patients compared with healthy controls. Remaining circulating ILCs revealed decreased frequencies of ILC2 in severe COVID‐19, with a concomitant decrease of ILC precursors (ILCp) in all patients, compared with controls. ILC2 and ILCp showed an activated phenotype with increased CD69 expression, whereas expression levels of the chemokine receptors CXCR3 and CCR4 were significantly altered in ILC2 and ILCp, and ILC1, respectively. The activated ILC profile of COVID‐19 patients was associated with soluble inflammatory markers, while frequencies of ILC subsets were correlated with laboratory parameters that reflect the disease severity.
Conclusion
This study provides insights into the potential role of ILCs in immune responses against SARS‐CoV‐2, particularly linked to the severity of COVID‐19.
In this study, we found that circulating innate lymphoid cells (ILCs) are reduced in COVID‐19 patients. Severely diseased patients display decreased frequencies of ILC2 compared to moderate COVID‐19 patients and healthy donors, whereas ILC precursors are found decreased in all patients. The remaining circulating ILCs in COVID‐19 patients show a dysregulated expression of activation and migration markers, with an activated profile that associates with soluble inflammatory markers. Moreover, frequencies of ILC subsets correlate with laboratory parameters that reflect the disease severity.
During the large Ebola outbreak that affected West Africa in 2014 and 2015, studies were launched to evaluate potential treatments for the disease. A clinical trial to evaluate the effectiveness of ...the antiviral drug favipiravir was conducted in Guinea. This paper describes the main challenges of the implementation of the trial in the Ebola treatment center of Guéckédou. Following the principles of the Good Clinical Research Practices, we explored the aspects of the community's communication and engagement, ethical conduct, trial protocol compliance, informed consent of participants, ongoing benefit/risk assessment, record keeping, confidentiality of patients and study data, and roles and responsibilities of the actors involved. We concluded that several challenges have to be addressed to successfully implement a clinical trial during an international medical emergency but that the potential for collaboration between research teams and humanitarian organizations needs to be highlighted.
Quantitative magnetic resonance imaging (MRI), e.g. relaxation parameter mapping, may be sensitive to structural and compositional tissue changes, and could potentially be used to non-invasively ...detect and monitor early meniscus degeneration related to knee osteoarthritis.
To investigate MR relaxation times as potential biomarkers for meniscus degeneration through comparisons with histopathology.
We measured MR relaxation parameters in the posterior horn of 40 menisci (medial and lateral) at a wide range of degenerative stages. T1, T2 and T2∗ were mapped using standard and ultrashort echo time sequences at 9.4 T and compared to gold standard histology using Pauli's histopathological scoring system, including assessment of surface integrity, collagen organization, cellularity and Safranin-O staining.
All three relaxation times increased with total Pauli score (mean difference per score (95% CI) for T2∗: 0.62 (0.37, 0.86), T2: 0.83 (0.53, 1.1) and T1: 24.7 (16.5, 32.8) ms/score). Clear associations were seen with scores of surface integrity (mean difference per score for T2∗: 3.0 (1.8, 4.2), T2: 4.0 (2.5, 5.5) and T1: 116 (75.6, 156) ms/score) and collagen organization (mean difference between highest and lowest score for T2∗: 5.3 (1.6, 8.9), T2: 6.1 (1.7, 11) and T1: 204 (75.9, 332) ms). The results were less clear for the remaining histopathological measures.
MR relaxation times T1, T2 and T2∗ of ex vivo human menisci are associated with histologically verified degenerative processes, in particular related to surface integrity and collagen organization. If confirmed in vivo, MR relaxation times may thus be potential biomarkers for meniscus degeneration.
The underlying molecular mechanisms in osteoarthritis (OA) development are largely unknown. This study explores the proteome and the pairwise interplay of proteins in synovial fluid from patients ...with late-stage knee OA (arthroplasty), early knee OA (arthroscopy due to degenerative meniscal tear), and from deceased controls without knee OA. Synovial fluid samples were analyzed using state-of-the-art mass spectrometry with data-independent acquisition. The differential expression of the proteins detected was clustered and evaluated with data mining strategies and a multilevel model. Group-specific slopes of associations were estimated between expressions of each pair of identified proteins to assess the co-expression (i.e., interplay) between the proteins in each group. More proteins were increased in early-OA versus controls than late-stage OA versus controls. For most of these proteins, the fold changes between late-stage OA versus controls and early-stage OA versus controls were remarkably similar suggesting potential involvement in the OA process. Further, for the first time, this study illustrated distinct patterns in protein co-expression suggesting that the interplay between the protein machinery is increased in early-OA and lost in late-stage OA. Further efforts should focus on earlier stages of the disease than previously considered.
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•Synovial fluid proteomics study of different stages of osteoarthritis (OA).•Higher catabolic activity is found in both early- and late-stage OA.•Imbalance of the metabolic homeostasis in late-stage OA.•Understanding early-stage OA may lead to finding better effective therapies.
This study presents data of the proteome in human synovial fluid from three groups representing early-stage knee OA, end-stage knee OA, and controls without knee OA. The early stage had an increased protein activity, while in end-stage OA, there was a loss of interplay between the proteins. These results highlight the importance of studying the early stage of OA progression and that the interplay between the proteins may be an additional key element in disentangling the complex OA pathogenesis.
Ebola virus disease (EVD) is a highly lethal condition for which no specific treatment has proven efficacy. In September 2014, while the Ebola outbreak was at its peak, the World Health Organization ...released a short list of drugs suitable for EVD research. Favipiravir, an antiviral developed for the treatment of severe influenza, was one of these. In late 2014, the conditions for starting a randomized Ebola trial were not fulfilled for two reasons. One was the perception that, given the high number of patients presenting simultaneously and the very high mortality rate of the disease, it was ethically unacceptable to allocate patients from within the same family or village to receive or not receive an experimental drug, using a randomization process impossible to understand by very sick patients. The other was that, in the context of rumors and distrust of Ebola treatment centers, using a randomized design at the outset might lead even more patients to refuse to seek care. Therefore, we chose to conduct a multicenter non-randomized trial, in which all patients would receive favipiravir along with standardized care. The objectives of the trial were to test the feasibility and acceptability of an emergency trial in the context of a large Ebola outbreak, and to collect data on the safety and effectiveness of favipiravir in reducing mortality and viral load in patients with EVD. The trial was not aimed at directly informing future guidelines on Ebola treatment but at quickly gathering standardized preliminary data to optimize the design of future studies.
Inclusion criteria were positive Ebola virus reverse transcription PCR (RT-PCR) test, age ≥ 1 y, weight ≥ 10 kg, ability to take oral drugs, and informed consent. All participants received oral favipiravir (day 0: 6,000 mg; day 1 to day 9: 2,400 mg/d). Semi-quantitative Ebola virus RT-PCR (results expressed in "cycle threshold" Ct) and biochemistry tests were performed at day 0, day 2, day 4, end of symptoms, day 14, and day 30. Frozen samples were shipped to a reference biosafety level 4 laboratory for RNA viral load measurement using a quantitative reference technique (genome copies/milliliter). Outcomes were mortality, viral load evolution, and adverse events. The analysis was stratified by age and Ct value. A "target value" of mortality was defined a priori for each stratum, to guide the interpretation of interim and final analysis. Between 17 December 2014 and 8 April 2015, 126 patients were included, of whom 111 were analyzed (adults and adolescents, ≥13 y, n = 99; young children, ≤6 y, n = 12). Here we present the results obtained in the 99 adults and adolescents. Of these, 55 had a baseline Ct value ≥ 20 (Group A Ct ≥ 20), and 44 had a baseline Ct value < 20 (Group A Ct < 20). Ct values and RNA viral loads were well correlated, with Ct = 20 corresponding to RNA viral load = 7.7 log10 genome copies/ml. Mortality was 20% (95% CI 11.6%-32.4%) in Group A Ct ≥ 20 and 91% (95% CI 78.8%-91.1%) in Group A Ct < 20. Both mortality 95% CIs included the predefined target value (30% and 85%, respectively). Baseline serum creatinine was ≥110 μmol/l in 48% of patients in Group A Ct ≥ 20 (≥300 μmol/l in 14%) and in 90% of patients in Group A Ct < 20 (≥300 μmol/l in 44%). In Group A Ct ≥ 20, 17% of patients with baseline creatinine ≥110 μmol/l died, versus 97% in Group A Ct < 20. In patients who survived, the mean decrease in viral load was 0.33 log10 copies/ml per day of follow-up. RNA viral load values and mortality were not significantly different between adults starting favipiravir within <72 h of symptoms compared to others. Favipiravir was well tolerated.
In the context of an outbreak at its peak, with crowded care centers, randomizing patients to receive either standard care or standard care plus an experimental drug was not felt to be appropriate. We did a non-randomized trial. This trial reaches nuanced conclusions. On the one hand, we do not conclude on the efficacy of the drug, and our conclusions on tolerance, although encouraging, are not as firm as they could have been if we had used randomization. On the other hand, we learned about how to quickly set up and run an Ebola trial, in close relationship with the community and non-governmental organizations; we integrated research into care so that it improved care; and we generated knowledge on EVD that is useful to further research. Our data illustrate the frequency of renal dysfunction and the powerful prognostic value of low Ct values. They suggest that drug trials in EVD should systematically stratify analyses by baseline Ct value, as a surrogate of viral load. They also suggest that favipiravir monotherapy merits further study in patients with medium to high viremia, but not in those with very high viremia.
ClinicalTrials.gov NCT02329054.
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