A two-way street - cellular metabolism and myofibroblast contraction.

Tissue fibrosis is characterised by the high-energy consumption associated with myofibroblast contraction. Although myofibroblast contraction relies on ATP production, the role of cellular metabolism in myofibroblast contraction has not yet been elucidated. Studies have so far only focused on myofibroblast contraction regulators, such as integrin receptors, TGF-ß and their shared transcription factor YAP/TAZ, in a fibroblast-myofibroblast transition setting. Additionally, the influence of the regulators on metabolism and vice versa have been described in this context. However, this has so far not yet been connected to myofibroblast contraction. This review focuses on the known and unknown of how cellular metabolism influences the processes leading to myofibroblast contraction and vice versa. We elucidate the signalling cascades responsible for myofibroblast contraction by looking at FMT regulators, mechanical cues, biochemical signalling, ECM properties and how they can influence and be influenced by cellular metabolism. By reviewing the existing knowledge on the link between cellular metabolism and the regulation of myofibroblast contraction, we aim to pinpoint gaps of knowledge and eventually help identify potential research targets to identify strategies that would allow switching tissue fibrosis towards tissue regeneration.

Associations of myeloid cells with cellular and humoral responses following vaccinations in patients with neuroimmunological diseases.

Disease-modifying therapies (DMTs) are widely used in neuroimmunological diseases such as multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD). Although these treatments are known to predispose patients to infections and affect their responses to vaccination, little is known about the impact of DMTs on the myeloid cell compartment. In this study, we use mass cytometry to examine DMT-associated changes in the innate immune system in untreated and treated patients with MS (n = 39) or NMOSD (n = 23). We also investigated the association between changes in myeloid cell phenotypes and longitudinal responsiveness to homologous primary, secondary, and tertiary SARS-CoV-2 mRNA vaccinations. Multiple DMT-associated myeloid cell clusters, in particular CD64+HLADRlow granulocytes, showed significant correlations with B and T cell responses induced by vaccination. Our findings suggest the potential role of myeloid cells in cellular and humoral responses following vaccination in DMT-treated patients with neuroimmunological diseases.

Understanding, diagnosing, and treating Myalgic encephalomyelitis/chronic fatigue syndrome - State of the art: Report of the 2nd international meeting at the Charité Fatigue Center.

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a devastating disease affecting millions of people worldwide. Due to the 2019 pandemic of coronavirus disease (COVID-19), we are facing a significant increase of ME/CFS prevalence. On May 11th to 12th, 2023, the second international ME/CFS conference of the Charité Fatigue Center was held in Berlin, Germany, focusing on pathomechanisms, diagnosis, and treatment. During the two-day conference, more than 100 researchers from various research fields met on-site and over 700 attendees participated online to discuss the state of the art and novel findings in this field. Key topics from the conference included: the role of the immune system, dysfunction of endothelial and autonomic nervous system, and viral reactivation. Furthermore, there were presentations on innovative diagnostic measures and assessments for this complex disease, cutting-edge treatment approaches, and clinical studies. Despite the increased public attention due to the COVID-19 pandemic, the subsequent rise of Long COVID-19 cases, and the rise of funding opportunities to unravel the pathomechanisms underlying ME/CFS, this severe disease remains highly underresearched. Future adequately funded research efforts are needed to further explore the disease etiology and to identify diagnostic markers and targeted therapies.

Haunting innate immune memories of COVID-19.

In addition to acute hyperinflammatory responses, SARS-CoV-2 infections can have long-term effects on our immune system leading to, for example, post-acute sequelae of COVID-19 (PASC). In this issue of Cell, Cheong et al. show that severe infections via IL-6 induce persistent epigenetic signatures in hemopoietic stem cells and their myeloid progenitors associated with increased inflammatory potential.

Fighting Post-COVID and ME/CFS - development of curative therapies.

The sequela of COVID-19 include a broad spectrum of symptoms that fall under the umbrella term post-COVID-19 condition or syndrome (PCS). Immune dysregulation, autoimmunity, endothelial dysfunction, viral persistence, and viral reactivation have been identified as potential mechanisms. However, there is heterogeneity in expression of biomarkers, and it is unknown yet whether these distinguish different clinical subgroups of PCS. There is an overlap of symptoms and pathomechanisms of PCS with postinfectious myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). No curative therapies are available for ME/CFS or PCS. The mechanisms identified so far provide targets for therapeutic interventions. To accelerate the development of therapies, we propose evaluating drugs targeting different mechanisms in clinical trial networks using harmonized diagnostic and outcome criteria and subgrouping patients based on a thorough clinical profiling including a comprehensive diagnostic and biomarker phenotyping.

Donor antigen-specific regulatory T cell administration to recipients of live donor kidneys: A ONE Study consortium pilot trial.

Regulatory T cells (Tregs) can inhibit cellular immunity in diverse experimental models and have entered early phase clinical trials in autoimmunity and transplantation to assess safety and efficacy. As part of the ONE Study consortium, we conducted a phase I-II clinical trial in which purified donor antigen reactive (dar)-Tregs (CD4+CD25+CD127lo) were administered to 3 patients, 7 to 11 days after live donor renal transplant. Recipients received a modified immunosuppression regimen, without induction therapy, consisting of maintenance tacrolimus, mycophenolate mofetil, and steroids. Steroids were weaned off over 14 weeks. No rejection was seen on any protocol biopsy. Therefore, all patients discontinued mycophenolate mofetil 11 to 13 months posttransplant, per protocol. An early for-cause biopsy in 1 patient, 5 days after dar-Treg infusion, revealed absence of rejection and accumulation of Tregs in the kidney allograft. All patients had Treg-containing lymphoid aggregates evident on protocol biopsies performed 8 months posttransplant. The patients are now all >6 years posttransplant on tacrolimus monotherapy with excellent graft function. None experienced rejection episodes. No serious adverse events were attributable to Treg administration. These results support a favorable safety profile of dar-Tregs administered early after renal transplant, suggest early biopsy might be an instructive research endpoint and provide preliminary evidence of potential immunomodulatory activity.

CD4+ T cell calibration of antigen-presenting cells optimizes antiviral CD8+ T cell immunity.

Antiviral CD8+ T cell immunity depends on the integration of various contextual cues, but how antigen-presenting cells (APCs) consolidate these signals for decoding by T cells remains unclear. Here, we describe gradual interferon-α/interferon-ß (IFNα/ß)-induced transcriptional adaptations that endow APCs with the capacity to rapidly activate the transcriptional regulators p65, IRF1 and FOS after CD4+ T cell-mediated CD40 stimulation. While these responses operate through broadly used signaling components, they induce a unique set of co-stimulatory molecules and soluble mediators that cannot be elicited by IFNα/ß or CD40 alone. These responses are critical for the acquisition of antiviral CD8+ T cell effector function, and their activity in APCs from individuals infected with severe acute respiratory syndrome coronavirus 2 correlates with milder disease. These observations uncover a sequential integration process whereby APCs rely on CD4+ T cells to select the innate circuits that guide antiviral CD8+ T cell responses.

RECAST: Study protocol for an observational study for the understanding of the increased REsilience of Children compared to Adults in SARS-CoV-2 infecTion.

INTRODUCTION: The SARS-CoV-2 pandemic remains a threat to public health. Soon after its outbreak, it became apparent that children are less severely affected. Indeed, opposing clinical manifestations between children and adults are observed for other infections. The SARS-CoV-2 outbreak provides the unique opportunity to study the underlying mechanisms. This protocol describes the methods of an observational study that aims to characterise age dependent differences in immune responses to primary respiratory infections using SARS-CoV-2 as a model virus and to assess age differences in clinical outcomes including lung function. METHODS AND ANALYSIS: The study aims to recruit at least 120 children and 60 adults that are infected with SARS-CoV-2 and collect specimen for a multiomics analysis, including single cell RNA sequencing of nasal epithelial cells and peripheral blood mononuclear cells, mass cytometry of whole blood samples and nasal cells, mass spectrometry-based serum and plasma proteomics, nasal epithelial cultures with functional in vitro analyses, SARS-CoV-2 antibody testing, sequencing of the viral genome and lung function testing. Data obtained from this multiomics approach are correlated with medical history and clinical data. Recruitment started in October 2020 and is ongoing. ETHICS AND DISSEMINATION: The study was reviewed and approved by the Ethics Committee of Charité - Universitätsmedizin Berlin (EA2/066/20). All collected specimens are stored in the central biobank of Charité - Universitätsmedizin Berlin and are made available to all participating researchers and on request. TRIAL REGISTRATION NUMBER: DRKS00025715, pre-results publication.

A Phase I/IIa study of autologous tolerogenic dendritic cells immunotherapy in kidney transplant recipients.

Kidney transplant survival is shortened by chronic rejection and side effects of standard immunosuppressive drugs. Cell-based immunotherapy with tolerogenic dendritic cells has long been recognized as a promising approach to reduce general immunosuppression. Published trials report the safety and the absence of therapy-related adverse reactions in patients treated with tolerogenic dendritic cells suffering from several inflammatory diseases. Here, we present the first phase I clinical trial results using human autologous tolerogenic dendritic cells (ATDC) in kidney transplantation. Eight patients received ATDC the day before transplantation in conjunction with standard steroids, mycophenolate mofetil and tacrolimus immunosuppression with an option to taper mycophenolate mofetil. ATDC preparations were manufactured in a Good Manufacturing Practice-compliant facility and fulfilled cell count, viability, purity and identity criteria for release. A control group of nine patients received the same standard immunosuppression, except basiliximab induction replaced ATDC therapy and mycophenolate tapering was not allowed. During the three-year follow-up, no deaths occurred and there was 100% graft survival. No significant increase of adverse events was associated with ATDC infusion. Episodes of rejection were observed in two patients from the ATDC group and one patient from the control group. However, all rejections were successfully treated by glucocorticoids. Mycophenolate was successfully reduced/stopped in five patients from the ATDC group, allowing tacrolimus monotherapy for two of them. Regarding immune monitoring, reduced CD8 T cell activation markers and increased Foxp3 expression were observed in the ATDC group. Thus, our results demonstrate ATDC administration safety in kidney-transplant recipients.

Pro-Resolving Mediators in Rotator Cuff Disease: How Is the Bursa Involved?

So far, tendon regeneration has mainly been analyzed independent from its adjacent tissues. However, the subacromial bursa in particular appears to influence the local inflammatory milieu in the shoulder. The resolution of local inflammation in the shoulder tissues is essential for tendon regeneration, and specialized pro-resolving mediators (SPMs) play a key role in regulating the resolution of inflammation. Here, we aimed to understand the influence of the bursa on disease-associated processes in neighboring tendon healing. Bursa tissue and bursa-derived cells from patients with intact, moderate and severe rotator cuff disease were investigated for the presence of pro-resolving and inflammatory mediators, as well as their effect on tenocytes and sensitivity to mechanical loading by altering SPM signaling mediators in bursa cells. SPM signal mediators were present in the bursae and altered depending on the severity of rotator cuff disease. SPMs were particularly released from the bursal tissue of patients with rotator cuff disease, and the addition of bursa-released factors to IL-1ß-challenged tenocytes improved tenocyte characteristics. In addition, mechanical loading modulated pro-resolving processes in bursa cells. In particular, pathological high loading (8% strain) increased the expression and secretion of SPM signaling mediators. Overall, this study confirms the importance of bursae in regulating inflammatory processes in adjacent rotator cuff tendons.

Distinct immunological and molecular signatures underpinning influenza vaccine responsiveness in the elderly.

Seasonal influenza outbreaks, especially in high-risk groups such as the elderly, represent an important public health problem. Prevailing inadequate efficacy of seasonal vaccines is a crucial bottleneck. Understanding the immunological and molecular mechanisms underpinning differential influenza vaccine responsiveness is essential to improve vaccination strategies. Here we show comprehensive characterization of the immune response of randomly selected elderly participants (≥ 65 years), immunized with the adjuvanted influenza vaccine Fluad. In-depth analyses by serology, multi-parametric flow cytometry, multiplex and transcriptome analysis, coupled to bioinformatics and mathematical modelling, reveal distinguishing immunological and molecular features between responders and non-responders defined by vaccine-induced seroconversion. Non-responders are specifically characterized by multiple suppressive immune mechanisms. The generated comprehensive high dimensional dataset enables the identification of putative mechanisms and nodes responsible for vaccine non-responsiveness independently of confounding age-related effects, with the potential to facilitate development of tailored vaccination strategies for the elderly.

SARS-CoV-2 mRNA vaccinations fail to elicit humoral and cellular immune responses in patients with multiple sclerosis receiving fingolimod.

BACKGROUND: SARS-CoV-2 mRNA vaccination of healthy individuals is highly immunogenic and protective against severe COVID-19. However, there are limited data on how disease-modifying therapies (DMTs) alter SARS-CoV-2 mRNA vaccine immunogenicity in patients with autoimmune diseases. METHODS: As part of a prospective cohort study, we investigated the induction, stability and boosting of vaccine-specific antibodies, B cells and T cells in patients with multiple sclerosis (MS) on different DMTs after homologous primary, secondary and booster SARS-CoV-2 mRNA vaccinations. Of 126 patients with MS analysed, 105 received either anti-CD20-based B cell depletion (aCD20-BCD), fingolimod, interferon-ß, dimethyl fumarate, glatiramer acetate, teriflunomide or natalizumab, and 21 were untreated MS patients for comparison. RESULTS: In contrast to all other MS patients, and even after booster, most aCD20-BCD- and fingolimod-treated patients showed no to markedly reduced anti-S1 IgG, serum neutralising activity and a lack of receptor binding domain-specific and S2-specific B cells. Patients receiving fingolimod additionally lacked spike-reactive CD4+ T cell responses. The duration of fingolimod treatment, rather than peripheral blood B and T cell counts prior to vaccination, determined whether a humoral immune response was elicited. CONCLUSIONS: The lack of immunogenicity under long-term fingolimod treatment demonstrates that functional immune responses require not only immune cells themselves, but also access of these cells to the site of inoculation and their unimpeded movement. The absence of humoral and T cell responses suggests that fingolimod-treated patients with MS are at risk for severe SARS-CoV-2 infections despite booster vaccinations, which is highly relevant for clinical decision-making and adapted protective measures, particularly considering additional recently approved sphingosine-1-phosphate receptor antagonists for MS treatment.

The Host Peritoneal Cavity Harbors Prominent Memory Th2 and Early Recall Responses to an Intestinal Nematode.

Intestinal parasitic nematodes affect a quarter of the world's population, typically eliciting prominent effector Th2-driven host immune responses. As not all infected hosts develop protection against reinfection, our current understanding of nematode-induced memory Th2 responses remains limited. Here, we investigated the activation of memory Th2 cells and the mechanisms driving early recall responses to the enteric nematode Heligmosomoides polygyrus in mice. We show that nematode-cured mice harbor memory Th2 cells in lymphoid and non-lymphoid organs with distinct transcriptional profiles, expressing recirculation markers like CCR7 and CD62-L in the mesenteric lymph nodes (mLN), and costimulatory markers like Ox40, as well as tissue homing and activation markers like CCR2, CD69 and CD40L in the gut and peritoneal cavity (PEC). While memory Th2 cells persist systemically in both lymphoid and non-lymphoid tissues following cure of infection, peritoneal memory Th2 cells in particular displayed an initial prominent expansion and strong parasite-specific Th2 responses during early recall responses to a challenge nematode infection. This effect was paralleled by a significant influx of dendritic cells (DC) and eosinophils, both also appearing exclusively in the peritoneal cavity of reinfected mice. In addition, we show that within the peritoneal membrane lined by peritoneal mesothelial cells (PeM), the gene expression levels of cell adhesion markers VCAM-1 and ICAM-1 decrease significantly in response to a secondary infection. Overall, our findings indicate that the host peritoneal cavity in particular harbors prominent memory Th2 cells and appears to respond directly to H. polygyrus by an early recall response via differential regulation of cell adhesion markers, marking the peritoneal cavity an important site for host immune responses to an enteric pathogen.

Age-Related Differences in Structure and Function of Nasal Epithelial Cultures From Healthy Children and Elderly People.

The nasal epithelium represents the first line of defense against inhaled pathogens, allergens, and irritants and plays a key role in the pathogenesis of a spectrum of acute and chronic airways diseases. Despite age-dependent clinical phenotypes triggered by these noxious stimuli, little is known about how aging affects the structure and function of the airway epithelium that is crucial for lung homeostasis and host defense. The aim of this study was therefore to determine age-related differences in structural and functional properties of primary nasal epithelial cultures from healthy children and non-smoking elderly people. To achieve this goal, highly differentiated nasal epithelial cultures were established from nasal brushes at air-liquid interface and used to study epithelial cell type composition, mucin (MUC5AC and MUC5B) expression, and ion transport properties. Furthermore, we determined age-dependent molecular signatures using global proteomic analysis. We found lower numeric densities of ciliated cells and higher levels of MUC5AC expression in cultures from children vs. elderly people. Bioelectric studies showed no differences in basal ion transport properties, ENaC-mediated sodium absorption, or CFTR-mediated chloride transport, but detected decreased calcium-activated TMEM16A-mediated chloride secretory responses in cultures from children vs. elderly people. Proteome analysis identified distinct age-dependent molecular signatures associated with ciliation and mucin biosynthesis, as well as other pathways implicated in aging. Our data identified intrinsic, age-related differences in structure and function of the nasal epithelium and provide a basis for further studies on the role of these findings in age-dependent airways disease phenotypes observed with a spectrum of respiratory infections and other noxious stimuli.

Early prediction of renal graft function: Analysis of a multi-center, multi-level data set.

PURPOSE OF THE STUDY: Long-term graft survival rates after renal transplantation are still poor. We aimed to build an early predictor of an established long-term outcomes marker, the estimated glomerular filtration rate (eGFR) one year post-transplant (eGFR-1y). MATERIALS AND METHODS: A large cohort of 376 patients was characterized for a multi-level bio-marker panel including gene expression, cytokines, metabolomics and antibody reactivity profiles. Almost one thousand samples from the pre-transplant and early post-transplant period were analysed and employed for machine learning-assisted prediction. RESULTS: Pre-transplant data led to a prediction achieving a Pearson's correlation coefficient of r=0.38 between measured and predicted eGFR-1y. Two weeks post-transplant, the correlation was improved to r=0.63, and at the third month, to r=0.76. eGFR values were stable throughout the first post-transplant year. Several characteristics were predictive for eGFR, including age of donor and recipient, body mass index, HLA mismatch, cytomegalovirus mismatch and valganciclovir prophylaxis. Additionally, a subset of 19 nuclear magnetic resonance bins of the urine metabolome data was shown to have potential applications in non-invasive eGFR monitoring. Importantly, we identified the expression of the genes TMEM176B and HMMR as potential prognostic markers for changes in the eGFR after the second post-transplantation week. CONCLUSIONS: Our multi-center, multi-level data set represents a milestone in the efforts to predict transplant outcome. While an acceptable predictive capacity was achieved, we are still far from predicting changes in the eGFR precisely. Additional studies employing further marker panels are needed to establish predictors of eGFR-1y for clinical application; herein, gene expression markers seem to hold the most promise.

SARS-CoV-2 infection triggers profibrotic macrophage responses and lung fibrosis.

COVID-19-induced "acute respiratory distress syndrome" (ARDS) is associated with prolonged respiratory failure and high mortality, but the mechanistic basis of lung injury remains incompletely understood. Here, we analyze pulmonary immune responses and lung pathology in two cohorts of patients with COVID-19 ARDS using functional single-cell genomics, immunohistology, and electron microscopy. We describe an accumulation of CD163-expressing monocyte-derived macrophages that acquired a profibrotic transcriptional phenotype during COVID-19 ARDS. Gene set enrichment and computational data integration revealed a significant similarity between COVID-19-associated macrophages and profibrotic macrophage populations identified in idiopathic pulmonary fibrosis. COVID-19 ARDS was associated with clinical, radiographic, histopathological, and ultrastructural hallmarks of pulmonary fibrosis. Exposure of human monocytes to SARS-CoV-2, but not influenza A virus or viral RNA analogs, was sufficient to induce a similar profibrotic phenotype in vitro. In conclusion, we demonstrate that SARS-CoV-2 triggers profibrotic macrophage responses and pronounced fibroproliferative ARDS.

Deciphering the Role of Humoral and Cellular Immune Responses in Different COVID-19 Vaccines-A Comparison of Vaccine Candidate Genes in Roborovski Dwarf Hamsters.

With the exception of inactivated vaccines, all SARS-CoV-2 vaccines currently used for clinical application focus on the spike envelope glycoprotein as a virus-specific antigen. Compared to other SARS-CoV-2 genes, mutations in the spike protein gene are more rapidly selected and spread within the population, which carries the risk of impairing the efficacy of spike-based vaccines. It is unclear to what extent the loss of neutralizing antibody epitopes can be compensated by cellular immune responses, and whether the use of other SARS-CoV-2 antigens might cause a more diverse immune response and better long-term protection, particularly in light of the continued evolution towards new SARS-CoV-2 variants. To address this question, we explored immunogenicity and protective effects of adenoviral vectors encoding either the full-length spike protein (S), the nucleocapsid protein (N), the receptor binding domain (RBD) or a hybrid construct of RBD and the membrane protein (M) in a highly susceptible COVID-19 hamster model. All adenoviral vaccines provided life-saving protection against SARS-CoV-2-infection. The most efficient protection was achieved after exposure to full-length spike. However, the nucleocapsid protein, which triggered a robust T-cell response but did not facilitate the formation of neutralizing antibodies, controlled early virus replication efficiently and prevented severe pneumonia. Although the full-length spike protein is an excellent target for vaccines, it does not appear to be the only option for future vaccine design.

Transient mTOR inhibition rescues 4-1BB CAR-Tregs from tonic signal-induced dysfunction.

The use of chimeric antigen receptor (CAR)-engineered regulatory T cells (Tregs) has emerged as a promising strategy to promote immune tolerance. However, in conventional T cells (Tconvs), CAR expression is often associated with tonic signaling, which can induce CAR-T cell dysfunction. The extent and effects of CAR tonic signaling vary greatly according to the expression intensity and intrinsic properties of the CAR. Here, we show that the 4-1BB CSD-associated tonic signal yields a more dramatic effect in CAR-Tregs than in CAR-Tconvs with respect to activation and proliferation. Compared to CD28 CAR-Tregs, 4-1BB CAR-Tregs exhibit decreased lineage stability and reduced in vivo suppressive capacities. Transient exposure of 4-1BB CAR-Tregs to a Treg stabilizing cocktail, including an mTOR inhibitor and vitamin C, during ex vivo expansion sharply improves their in vivo function and expansion after adoptive transfer. This study demonstrates that the negative effects of 4-1BB tonic signaling in Tregs can be mitigated by transient mTOR inhibition.

Untimely TGFß responses in COVID-19 limit antiviral functions of NK cells.

SARS-CoV-2 is a single-stranded RNA virus that causes COVID-19. Given its acute and often self-limiting course, it is likely that components of the innate immune system play a central part in controlling virus replication and determining clinical outcome. Natural killer (NK) cells are innate lymphocytes with notable activity against a broad range of viruses, including RNA viruses1,2. NK cell function may be altered during COVID-19 despite increased representation of NK cells with an activated and adaptive phenotype3,4. Here we show that a decline in viral load in COVID-19 correlates with NK cell status and that NK cells can control SARS-CoV-2 replication by recognizing infected target cells. In severe COVID-19, NK cells show defects in virus control, cytokine production and cell-mediated cytotoxicity despite high expression of cytotoxic effector molecules. Single-cell RNA sequencing of NK cells over the time course of the COVID-19 disease spectrum reveals a distinct gene expression signature. Transcriptional networks of interferon-driven NK cell activation are superimposed by a dominant transforming growth factor-ß (TGFß) response signature, with reduced expression of genes related to cell-cell adhesion, granule exocytosis and cell-mediated cytotoxicity. In severe COVID-19, serum levels of TGFß peak during the first two weeks of infection, and serum obtained from these patients severely inhibits NK cell function in a TGFß-dependent manner. Our data reveal that an untimely production of TGFß is a hallmark of severe COVID-19 and may inhibit NK cell function and early control of the virus.