Exploiting inhalable microparticles incorporating hybrid polymer-lipid nanoparticles loaded with Iloprost manages lung hyper-inflammation.

This study focuses on developing of a novel inhalation therapy for managing lung hyper-inflammation, producing hybrid polymer-lipid nanoparticles loaded with Iloprost (Ilo). These nanoparticles showed a size of approximately 100 nm with a core-shell structure and provided prolonged drug release, reaching 28 wt% after 6 h of incubation. The phospholipid composition and quantity (64 wt% on the total sample weight) result in minimal interaction with mucin and a significant effect on the rheology of a cystic fibrosis mucus model, in terms of reducing complex viscosity. To obtain an inhalable microparticulate matrix suitable for incorporating Ilo@PEG-LPHNPs, the qualitative and quantitative composition of the feed fluid for the spray drying (SD) process was optimized. The selected composition (10 % wt/vol of mannitol and 10 % wt of ammonium bicarbonate relative to the weight of mannitol) was used to produce Nano-into Microparticles (NiM). The characterization of NiM revealed excellent aerodynamic properties, with a Mass Median Aerodynamic Diameter (MMAD) of 4.34 µm and a Fine Particle Fraction (FPF) of approximately 57 %. Biological characterization revealed that the particles are non-toxic to 16-HBE cells and can effectively evade macrophage uptake, likely due to the presence of PEG in their composition. Moreover, the delivered Iloprost significantly downregulates the production of the pro-inflammatory cytokine IL-6, showing the therapeutic potential of this drug delivery system.

Bacterial-like inflammatory response in children with adenovirus leads to inappropriate antibiotic use: a multicenter cohort study.

PURPOSE: To compare the clinical severity of Human Adenovirus (HAdV) infection with other viral diseases in a cohort of children, evaluating presentation, therapy, and outcome. METHODS: We conducted a retrospective multicenter cohort study in Italian children hospitalized from January to December 2023 for respiratory symptoms. The study included children with HAdV infection presenting primarily with respiratory symptoms. Patients with isolated gastrointestinal involvement or coinfection with bacteria were excluded. RESULTS: A total of 171 children were enrolled: 98 with HAdV infection (age 44.3 ± 37.9 months) and 73 with other viruses (age 20.4 ± 27.2 months). In the first group, 57.1% had a coinfection with one or more additional viruses. The most common symptoms were fever (89.8%), cough (73.5%) and sore throat (52%). Respiratory distress and hypoxemia were more frequent in the non-HAdV group. Children with HAdV infection demonstrated significantly higher C-reactive protein levels (50.8 ± 54.2 vs. 16.5 ± 33.8 mg/L, p < 0.001), experienced a longer duration of fever (4.9 ± 3.6 vs. 3.4 ± 2.3 days, p = 0.009) and were more likely to receive antibiotic treatment (77.6% vs. 27.4%, p < 0.001). No differences were observed in hospitalization stay, rate of complications, and ICU admission. CONCLUSIONS: Interestingly, our data suggests that HAdV-infected children exhibit a more pronounced inflammatory response despite experiencing less severe respiratory symptoms compared to other viruses. The presence of prolonged fever and a strong inflammatory response often leads to antibiotic overuse during the initial phase, when the viral etiology is yet to be confirmed. Early and accurate identification of HAdV infection is crucial to optimize treatment strategies and minimize unnecessary antibiotic use.

Prospective Variation of Cytokine Trends during COVID-19: A Progressive Approach from Disease Onset until Outcome.

COVID-19 is characterized by pronounced hypercytokinemia. The cytokine switch, marked by an imbalance between pro-inflammatory and anti-inflammatory cytokines, emerged as a focal point of investigation throughout the COVID-19 pandemic. However, the kinetics and temporal dynamics of cytokine release remain contradictory, making the development of new therapeutics difficult, especially in severe cases. This study collected serum samples from SARS-CoV-2 infected patients at 72 h intervals and monitored them for various cytokines at each timepoint until hospital discharge or death. Cytokine levels were analyzed based on time since symptom onset and patient outcomes. All cytokines studied prospectively were strong predictors of mortality, particularly IL-4 (AUC = 0.98) and IL-1ß (AUC = 0.96). First-timepoint evaluations showed elevated cytokine levels in the mortality group (p < 0.001). Interestingly, IFN-γ levels decreased over time in the death group but increased in the survival group. Patients who died exhibited sustained levels of IL-1ß and IL-4 and increased IL-6 levels over time. These findings suggest cytokine elevation is crucial in predicting COVID-19 mortality. The dynamic interplay between IFN-γ and IL-4 highlights the balance between Th1/Th2 immune responses and underscores IFN-γ as a powerful indicator of immune dysregulation throughout the infection.

Kawasaki Disease-Associated Cytokine Storm Syndrome.

Kawasaki disease (KD) is a hyperinflammatory syndrome manifesting as an acute systemic vasculitis characterized by fever, nonsuppurative conjunctival injection, rash, oral mucositis, extremity changes, and cervical lymphadenopathy. KD predominantly affects young children and shares clinical features and immunobiology with other hyperinflammation syndromes including systemic juvenile idiopathic arthritis (sJIA) and multisystem inflammatory syndrome in children (MIS-C). Cytokine storm syndrome (CSS) is an acute complication in ~2% of KD patients; however, the incidence is likely underestimated as many clinical and laboratory features of both diseases overlap. CSS should be entertained when a child with KD is unresponsive to IVIG therapy with recalcitrant fever. Early recognition and prompt institution of immunomodulatory treatment can substantially reduce the mortality and morbidity of CSS in KD. Given the known pathogenetic role of IL-1ß in both syndromes, the early use of IL-1 blockers in refractory KD with CSS deserves consideration.

Extracorporeal Elimination of Pro- and Anti-inflammatory Modulators by the Cytokine Adsorber CytoSorb® in Patients with Hyperinflammation: A Prospective Study.

INTRODUCTION: The release of pro-inflammatory cytokines in critically ill patients with sepsis leads to endothelial dysfunction resulting in cardiocirculatory insufficiency. Their extracorporeal elimination using the cytokine adsorber CytoSorb® (CS) (adsorption of especially hydrophobic molecules < 60 kDa) might be promising, but data about the adsorption capacity as well as a potential harmful adsorption of anti-inflammatory cytokines are missing so far. METHODS: The prospective Cyto-SOLVE-study included 15 patients with sepsis or other hyperinflammatory conditions (interleukin 6 > 500 pg/ml), continuous kidney replacement therapy, and the application of CS. Various cytokines and chemokines were measured pre- and post-CS as well as in patients' blood at predefined timepoints. Significant changes in the concentrations were detected with the Wilcoxon test with associated samples. Clearance of the adsorber (ml/min) was calculated with: b l o o d f l o w ∗ c o n c e n t r a t i o n p r e - p o s t c o n c e n t r a t i o n pre . RESULTS: Most of the inflammatory mediators showed a high initial extracorporeal clearance of 70-100 ml/min after CS installation, which dropped quickly to 10-30 ml/min after 6 h of treatment. No difference in clearance was observed between pro- and anti-inflammatory cytokines. Despite extracorporeal adsorption, a significant (p < 0.05) decrease in the blood concentration after 6 h was only observed for the pro-inflammatory cytokines tumor necrosis factorα (TNF-α) (median 284 vs. 230 pg/ml), vascular endothelial growth factor (VEGF) (median 294 vs. 252 pg/ml), macrophage inflammatory protein 1a (MIP-1a) (median 11.1 vs. 9.0 pg/ml), and regulated upon activation, normal T cell expressed and secreted (RANTES) (median 811 vs. 487 pg/ml) as well as the anti-inflammatory cytokines interleukin 4 (median 9.3 vs. 6.4 pg/ml), interleukin 10 (median 88 vs. 56 pg/ml), and platelet-derived growth factor (PDGF) (median 177 vs. 104 pg/ml). A significant (p < 0.05) decrease in patients' blood after 12 h was only detected for interleukin 10. CONCLUSIONS: CS can adsorb pro- as well as anti-inflammatory mediators with no relevant difference regarding the adsorption rate. A fast saturation of the adsorber resulted in a rapid decrease of the clearance. The potential clinical benefit or harm of this unspecific cytokine adsorption needs to be evaluated in the future. TRIAL REGISTRATION: ClinicalTrials.gov NCT04913298, registration date June 4, 2021.

The role of hemoadsorption in cardiac surgery - a systematic review.

BACKGROUND: Extracorporeal blood purification has been widely used in intensive care medicine, nephrology, toxicology, and other fields. During the last decade, with the emergence of new adsorptive blood purification devices, hemoadsorption has been increasingly applied during CPB in cardiac surgery, for patients at different inflammatory risks, or for postoperative complications. Clinical evidence so far has not provided definite answers concerning this adjunctive treatment. The current systematic review aimed to critically assess the role of perioperative hemoadsorption in cardiac surgery, by summarizing the current knowledge in this clinical setting. METHODS: A literature search of PubMed, Cochrane library, and the database provided by CytoSorbents was conducted on June 1st, 2023. The search terms were chosen by applying neutral search keywords to perform a non-biased systematic search, including language variations of terms "cardiac surgery" and "hemoadsorption". The screening and selection process followed scientific principles (PRISMA statement). Abstracts were considered for inclusion if they were written in English and published within the last ten years. Publications were eligible for assessment if reporting on original data from any type of study (excluding case reports) in which a hemoadsorption device was investigated during or after cardiac surgery. Results were summarized according to sub-fields and presented in a tabular view. RESULTS: The search resulted in 29 publications with a total of 1,057 patients who were treated with hemoadsorption and 988 control patients. Articles were grouped and descriptively analyzed due to the remarkable variability in study designs, however, all reported exclusively on CytoSorb® therapy. A total of 62% (18/29) of the included articles reported on safety and no unanticipated adverse events have been observed. The most frequently reported clinical outcome associated with hemoadsorption was reduced vasopressor demand resulting in better hemodynamic stability. CONCLUSIONS: The role of hemoadsorption in cardiac surgery seems to be justified in selected high-risk cases in infective endocarditis, aortic surgery, heart transplantation, and emergency surgery in patients under antithrombotic therapy, as well as in those who develop a dysregulated inflammatory response, vasoplegia, or septic shock postoperatively. Future large randomized controlled trials are needed to better define proper patient selection, dosing, and timing of the therapy.

One gene to rule them all - clinical perspectives of a potent suppressor of cytokine signaling - SOCS1.

The discovery of Suppressor of Cytokine Signaling 1 (SOCS1) in 1997 marked a significant milestone in understanding the regulation of Janus kinase/Signal transducer and activator of transcription (JAK/STAT) signaling pathways. Subsequent research deciphered its cellular functions, and recent insights into SOCS1 deficiencies in humans underscored its critical role in immune regulation. In humans, SOCS-haploinsufficiency (SOCS1-HI) presents a diverse clinical spectrum, encompassing autoimmune diseases, infection susceptibility, and cancer. Variability in disease manifestation, even within families sharing the same genetic variant, raises questions about clinical penetrance and the need for individualized treatments. Current therapeutic strategies include JAK inhibition, with promising results in controlling inflammation in SOCS1-HI patients. Hematopoietic stem cell transplantation and gene therapy emerge as promising avenues for curative treatments. The evolving landscape of SOCS1 research, emphasizes the need for a nuanced understanding of genetic variants and their functional consequences.

Inhaled Macrophage Apoptotic Bodies-Engineered Microparticle Enabling Construction of Pro-Regenerative Microenvironment to Fight Hypoxic Lung Injury in Mice.

Oxygen therapy cannot rescue local lung hypoxia in patients with severe respiratory failure. Here, an inhalable platform is reported for overcoming the aberrant hypoxia-induced immune changes and alveolar damage using camouflaged poly(lactic-co-glycolic) acid (PLGA) microparticles with macrophage apoptotic body membrane (cMAB). cMABs are preloaded with mitochondria-targeting superoxide dismutase/catalase nanocomplexes (NCs) and modified with pathology-responsive macrophage growth factor colony-stimulating factor (CSF) chains, which form a core-shell platform called C-cMAB/NC with efficient deposition in deeper alveoli and high affinity to alveolar epithelial cells (AECs) after CSF chains are cleaved by matrix metalloproteinase 9. Therefore, NCs can be effectively transported into mitochondria to inhibit inflammasome-mediated AECs damage in mouse models of hypoxic acute lung injury. Additionally, the at-site CSF release is sufficient to rescue circulating monocytes and macrophages and alter their phenotypes, maximizing synergetic effects of NCs on creating a pro-regenerative microenvironment that enables resolution of lung injury and inflammation. This inhalable platform may have applications to numerous inflammatory lung diseases.

Proteomic mapping identifies serum marker signatures associated with MIS-C specific hyperinflammation and cardiovascular manifestation.

Multisystem inflammatory syndrome in children (MIS-C) shares several clinical and immunological features with Kawasaki Disease (KD) and pediatric hyperinflammation, but the immuno-phenotypic overlap among these clinical mimics is still incompletely understood. Here we analyzed serum samples from treatment-naïve patients with MIS-C (n = 31) and KD (n = 11), pediatric hyperinflammation (n = 13) and healthy controls (HC, n = 10) by proximity extension assay (PEA) to profile 184 blood biomarkers. Collectively, immunophenotypic overlap between MIS-C and hyperinflammation exceeds overlap with KD. Overexpression of IL-17A in MIS-C and KD could best separate these conditions from hyperinflammatory conditions, while those were hallmarked by overabundance of adenosin deaminase and IL-18. Depletion in serum TNF-related subfamily member 9 (TNFRSF9) and apoptosis inducing ligand (TRAIL) linked with cardiovascular manifestations and myocarditis in MIS-C. Altogether, our analysis highlights important differences in molecular marker signatures also across different MIS-C and KD cohorts and suggests several previously unidentified molecular associations in context of cardiovascular inflammation.

The Effects of Convalescent Plasma Transfusion on Serum Levels of Macrophage-Associated Inflammatory Biomarkers in Patients with Severe COVID-19.

As an antibody-based therapy, plasma therapy has been used as an emergency therapeutic strategy against severe acute respiratory syndrome coronavirus type 2 infection. Due to the critical role of macrophages in coronavirus disease-19 (COVID-19)-associated hyperinflammation, the main objective of this study was to assess the effect of plasma transfusion on the expression levels of the inflammatory biomarkers involved in activation and pulmonary infiltration of macrophages. The target population included 50 severe hospitalized COVID-19 patients who were randomly assigned into 2 groups, including intervention and control. Serum levels of chemokine (C-C motif) ligand (CCL)-2, CCL-3, tumor necrosis factor (TNF)-α, and interleukin (IL)-6 were measured by enzyme-linked immunosorbent assay. Moreover, quantitative real-time polymerase chain reaction (PCR) was carried out to assess the relative expression of nuclear factor (NF)-κB1, NF-κB2, nuclear factor erythroid 2 p45-related factor 2 (NRF-2), and thioredoxin-interacting protein genes. Sampling was done at baseline and 72 h after receiving plasma. The intervention group demonstrated significantly lower serum levels of IL-6, TNF-α, and CCL-3. In addition, real-time PCR data analyses showed that the relative expression of NF-κB2 was significantly declined in the patients who received plasma. The use of convalescent plasma probably has a significant inhibitory effect on the cytokines, chemokines, and inflammatory genes related to macrophage activation, which are closely associated with the worsening of clinical outcomes in severe COVID-19.

Individualized High Dose Intravenous Anakinra Treatment in Cancer Patients with COVID-19 Associated Cytokine Storm: A Retrospective Controlled Study.

Objective: Patients with COVID-19 accompanying cancer have been reported to have higher morbidity and mortality. In this study, we aimed to evaluate the high-dose high intravenous anakinra treatment response and outcome in patients with COVID-19-associated cytokine storm accompanying cancer. Materials and Methods: This retrospective observational study was carried out at a tertiary referral center between September 01, 2021, and February 01, 2022, in Turkey. The study population consisted of two groups: patients receiving high-dose intravenous anakinra and patients treated with standard care. Results: Data from 146 patients in the anakinra group and 114 patients in the control group were analyzed. Malignancy frequency was 11% (n=16) in the anakinra group and 7% (n=8) in the control group. In survival analysis, a significantly lower survival rate was observed in patients with malignancy than those without in the control group (log-rank: p=0.002) and patients with malignancy in the control group compared to the anakinra group (log-rank: p=0.013). However, it did not differ between patients with and without malignancy in the anakinra group (log-rank: p=0.9). Conclusion: In the control group, mortality was higher in patients with malignancy compared to those without malignancy, but not in the anakinra group. Also, mortality was higher in patients receiving SoC compared to anakinra. Intravenous high-dose anakinra treatment is safe and effective in patients with COVID-19 accompanying cancer.

Early hematopoietic cell transplantation for familial hemophagocytic lymphohistiocytosis in a regional treatment network in Japan.

Familial hemophagocytic lymphohistiocytosis (FHLH) is a fatal hyperinflammation syndrome arising from the genetic defect of perforin-mediated cytolysis. Curative hematopoietic cell transplantation (HCT) is needed before development of central nervous system (CNS) disease. We studied treatment outcomes of 13 patients (FHLH2 n = 11, FHLH3 n = 2) consecutively diagnosed from 2011 to 2022 by flow cytometric screening for non-myeloablative HCT in a regional treatment network in Kyushu, Japan. One patient with a novel PRF1 variant escaped screening, but all patients with FHLH2 reached diagnosis and 8 of them received HCT until 3 and 9 months of age, respectively. The earliest HCT was conducted 65 days after birth. Three pretransplant deaths occurred in newborns with liver failure at diagnosis. Ten posttransplant patients have remained disease-free, 7 of whom had no neurological involvement. Time from first etoposide infusion to HCT was shorter in patients without CNS disease or bleeding than in patients with those factors (median [range] days: 62 [50-81] vs. 122 [89-209], p = 0.016). Six of 9 unrelated patients had a PRF1 c.1090_1091delCT variant. These results suggest that the critical times to start etoposide and HCT are within 3 months after birth and during etoposide control, respectively. Newborn screening may increase the percentage of disease-free survivors without complications.

Monocyte state 1 (MS1) cells in critically ill patients with sepsis or non-infectious conditions: association with disease course and host response.

BACKGROUND: Sepsis is a life-threatening condition arising from an aberrant host response to infection. Recent single-cell RNA sequencing investigations identified an immature bone-marrow-derived CD14+ monocyte phenotype with immune suppressive properties termed "monocyte state 1" (MS1) in patients with sepsis. Our objective was to determine the association of MS1 cell profiles with disease presentation, outcomes, and host response characteristics. METHODS: We used the transcriptome deconvolution method (CIBERSORTx) to estimate the percentage of MS1 cells from blood RNA profiles of patients with sepsis admitted to the intensive care unit (ICU). We compared these profiles to ICU patients without infection and to healthy controls. Host response dysregulation was further studied by gene co-expression network and gene set enrichment analyses of blood leukocytes, and measurement of 15 plasma biomarkers indicative of pathways implicated in sepsis pathogenesis. RESULTS: Sepsis patients (n = 332) were divided into three equally-sized groups based on their MS1 cell levels (low, intermediate, and high). MS1 groups did not differ in demographics or comorbidities. The intermediate and high MS1 groups presented with higher disease severity and more often had shock. MS1 cell abundance did not differ between survivors and non-survivors, or between patients who did or did not acquire a secondary infection. Higher MS1 cell percentages were associated with downregulation of lymphocyte-related and interferon response genes in blood leukocytes, with concurrent upregulation of inflammatory response pathways, including tumor necrosis factor signaling via nuclear factor-κB. Previously described sepsis host response transcriptomic subtypes showed different MS1 cell abundances, and MS1 cell percentages positively correlated with the "quantitative sepsis response signature" and "molecular degree of perturbation" scores. Plasma biomarker levels, indicative of inflammation, endothelial cell activation, and coagulation activation, were largely similar between MS1 groups. In ICU patients without infection (n = 215), MS1 cell percentages and their relation with disease severity, shock, and host response dysregulation were highly similar to those in sepsis patients. CONCLUSIONS: High MS1 cell percentages are associated with increased disease severity and shock in critically ill patients with sepsis or a non-infectious condition. High MS1 cell abundance likely indicates broad immune dysregulation, entailing not only immunosuppression but also anomalies reflecting exaggerated inflammatory responses.

Postoperative hyper-inflammation as a predictor of poor outcomes in patients with acute type A aortic dissection (ATAAD) undergoing surgical repair.

BACKGROUND: Postoperative hyper-inflammation is a frequent event in patients with acute Stanford type A aortic dissection (ATAAD) after surgical repair. This study's objective was to determine which inflammatory biomarkers could be used to make a better formula for identifying postoperative hyper-inflammation, and which risk factors were associated with hyper-inflammation. METHODS: A total of 405 patients were enrolled in this study from October 1, 2020 to April 1, 2023. Of these patients, 124 exhibited poor outcomes. In order to investigate the optimal cut-off values for poor outcomes, logistic and receiver operating characteristic analyses were performed on the following parameters on the first postoperative day: procalcitonin (PCT), C-reactive protein (CRP), interleukin-6 (IL-6), and systemic immune-inflammation index (SII). These cut-off points were used to separate the patients into hyper-inflammatory (n = 52) and control (n = 353) groups. Finally, the logistic were used to find the risk factors of hyper-inflammatory. RESULTS: PCT, CRP, IL-6, and SII were independent risk factors of poor outcomes in the multivariate logistic model. Cut-off points of these biomarkers were 2.18 ng/ml, 49.76 mg/L, 301.88 pg/ml, 2509.96 × 109/L respectively. These points were used to define postoperative hyper-inflammation (OR 2.97, 95% CI 1.35-6.53, P < 0.01). Cardiopulmonary bypass (CPB) > 180 min, and deep hypothermia circulatory arrest (DHCA) > 40 min were the independent risk factors for hyper-inflammation. CONCLUSIONS: PCT > 2.18, CRP > 49.76, IL-6 > 301.88, and SII < 2509.96 could be used to define postoperative hyper-inflammation which increased mortality and morbidity in patients after ATAAD surgery. Based on these findings, we found that CPB > 180 min and DHCA > 40 min were separate risk factors for postoperative hyper-inflammation.

Ruxolitinib, IV Immunoglobulin, and High-Dose Glucocorticoids for Critically Ill Adults With Secondary Hemophagocytic Lymphohistiocytosis: A Single-Center Observational Pilot Study.

OBJECTIVES: Secondary hemophagocytic lymphohistiocytosis (sHLH) is a cytokine-driven inflammatory syndrome that is associated with substantial morbidity and mortality and frequently leads to ICU admission. Overall survival in adults with sHLH remains poor, especially in those requiring intensive care. Classical chemotherapeutic treatment exhibits myelosuppression and toxicity. Recently, inhibition of Janus kinase signaling by ruxolitinib has shown efficacy in pediatric HLH. We therefore aimed to determine the activity and safety of a ruxolitinib-based regimen, in critically ill adults with sHLH. DESIGN: Observational pilot study. SETTING: Single-center tertiary academic ICU. PATIENTS: Nine adults (≥ 18 yr) who fulfilled at least five of the eight HLH-2004 criteria. INTERVENTION: Triplet regimen combining: 1) ruxolitinib, 2) polyvalent human IV immunoglobulins (IVIG) at a dose of 1 g/kg bodyweight for 5 days, and 3) high-dose corticosteroids (CSs, dexamethasone 10 mg/m² body surface area, or methylprednisolone equivalent) with subsequent tapering according to the HLH-2004 protocol. MEASUREMENT AND MAIN RESULTS: Nine patients (median age: 42 yr [25th-75th percentile: 32-54]; male: n = 6 males, median H-score: 299 [255-304]) were treated with the triplet regimen. The median Sequential Organ Failure Assessment score at HLH diagnosis was 9 (median; 25th-75th percentile: 7-12), indicating multiple-organ dysfunction in all patients. Within 10 days a significant decrease of the inflammatory parameters soluble interleukin-2 receptor and ferritin as well as a stabilization of the blood count could be shown. All patients were alive at ICU discharge (100% ICU survival), 1 patient died after ICU discharge because of traumatic intracerebral hemorrhage that might be related to HLH or treatment, corresponding to an overall survival of 86% in a 6 months follow-up period. CONCLUSION: In this small case series, a triplet regimen of ruxolitinib in combination with IVIG and CS was highly effective and save for treating critically ill adults with sHLH.

Novel Presentation of Major Histocompatibility Complex Class II Deficiency with Hemophagocytic Lymphohistiocytosis.

PURPOSE: Major histocompatibility complex (MHC) class II deficiency is one of the combined immune deficiency disorders caused by defects in the MHC class II regulatory genes leading to abnormal T cells development and function. Therefore, patients mainly present with increased susceptibility to infections, diarrhea, and failure to thrive. In this report, we present one MHC class II deficient patient with a novel presentation with Hemophagocytic Lymphohistiocytosis (HLH). METHODS: Immunophenotyping of lymphocyte subpopulations and HLA-DR expression was assess by flow cytometry. Gene mutational analysis was performed by whole exome and Sanger sequencing. RESULTS: We reported a 7-year-old girl, who was diagnosed at age of 2 years with MHC class II deficiency by genetic testing and flow cytometry. Two years later, she developed disseminated BCGitis which was treated with proper antimicrobial agents. At the age of 7 years, she presented with clinical features fulfilling 6 diagnostic criteria of HLH including evidence of hemophagocytic activity in bone marrow aspiration. Accordingly, the diagnosis of HLH was established and the patient was started on IV Dexamethasone, Anakinra and IVIG. Eventually, patient started to improve and was discharged in good condition. Few months later, the patient was readmitted with severe pneumonia and sepsis leading to death. CONCLUSION: Patients with MHC class II deficiency might present with disseminated BCGitis especially if the patient has severe T cell lymphopenia. Additionally, this immune defect might be added to the list of inborn errors of immunity that can be complicated with HLH.

TNFα: TNFR1 signaling inhibits maturation and maintains the pro-inflammatory programming of monocyte-derived macrophages in murine chronic granulomatous disease.

Introduction: Loss of NADPH oxidase activity results in proinflammatory macrophages that contribute to hyperinflammation in Chronic Granulomatous Disease (CGD). Previously, it was shown in a zymosan-induced peritonitis model that gp91phox-/- (CGD) monocyte-derived macrophages (MoMacs) fail to phenotypically mature into pro-resolving MoMacs characteristic of wild type (WT) but retain the ability to do so when placed in the WT milieu. Accordingly, it was hypothesized that soluble factor(s) in the CGD milieu thwart appropriate programming. Methods: We sought to identify key constituents using ex vivo culture of peritoneal inflammatory leukocytes and their conditioned media. MoMac phenotyping was performed via flow cytometry, measurement of efferocytic capacity and multiplex analysis of secreted cytokines. Addition of exogenous TNFα, TNFα neutralizing antibody and TNFR1-/- MoMacs were used to study the role of TNFα: TNFR1 signaling in MoMac maturation. Results: More extensive phenotyping defined normal MoMac maturation and demonstrated failure of maturation of CGD MoMacs both ex vivo and in vivo. Protein components, and specifically TNFα, produced and released by CGD neutrophils and MoMacs into conditioned media was identified as critical to preventing maturation. Exogenous addition of TNFα inhibited WT MoMac maturation, and its neutralization allowed maturation of cultured CGD MoMacs. TNFα neutralization also reduced production of IL-1ß, IL-6 and CXCL1 by CGD cells though these cytokines played no role in MoMac programming. MoMacs lacking TNFR1 matured more normally in the CGD milieu both ex vivo and following adoptive transfer in vivo. Discussion: These data lend mechanistic insights into the utility of TNFα blockade in CGD and to other diseases where such therapy has been shown to be beneficial.

[Hemophagocytic lymphohistiocytosis and macrophage activation syndrome : A multidisciplinary challenge]. / Hämophagozytische Lymphohistiozytose und Makrophagenaktivierungssyndrom : Eine multidisziplinäre Herausforderung.

Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening hyperinflammatory syndrome that is characterized by hyperferritinemia, cytopenia, disseminated intravascular coagulopathy and functional disorders of the liver and the central nervous system. The term macrophage activation syndrome is predominantly used for secondary HLH in the context of autoimmune diseases (e.g., systemic juvenile idiopathic arthritis). In addition, malignancies and genetic inborn errors of immunity can predispose to the development of HLH. Infections (e.g., Epstein-Barr virus) in turn represent possible triggers of an acute episode. Due to the unspecific manifestation of the disease, a systematic evaluation of the organ systems is recommended in the clinical and laboratory analytical clarification of hyperinflammatory syndromes. In general, the treatment should be carried out by a multidisciplinary team with expertise in rheumatology, hematological oncology, infectious diseases and intensive care medicine. The primary treatment of HLH usually consists of glucocorticoids and in cases of a rapid deterioration of the condition anakinra (interleukin 1 block) and intravenous immunoglobulins can be employed. Treatment of the underlying disease should be consequently carried out in parallel, together with antimicrobial treatment.

Gene editing of hematopoietic stem cells restores T-cell response in familial hemophagocytic lymphohistiocytosis.

BACKGROUND: Hemophagocytic lymphohistiocytosis (HLH) is a hyperinflammatory disorder characterized by a life-threatening cytokine storm and immunopathology. Familial HLH type 3 (FHL3) accounts for approximately 30% of all inborn HLH cases worldwide. It is caused by mutations in the UNC13D gene that result in impaired degranulation of cytotoxic vesicles and hence compromised T-cell- and natural killer-cell-mediated killing. Current treatment protocols, including allogeneic hematopoietic stem cell (HSC) transplantation, still show high mortality. OBJECTIVE: We sought to develop and evaluate a curative genome editing strategy in the preclinical FHL3 Jinx mouse model. Jinx mice harbor a cryptic splice donor site in Unc13d intron 26 and develop clinical symptoms of human FHL3 upon infection with lymphocytic choriomeningitis virus (LCMV). METHODS: We employed clustered regularly interspaced short palindromic repeats (CRISPR)-Cas technology to delete the disease-causing mutation in HSCs and transplanted Unc13d-edited stem cells into busulfan-conditioned Jinx recipient mice. Safety studies included extensive genotyping and chromosomal aberrations analysis by single targeted linker-mediated PCR sequencing (CAST-Seq)-based off-target analyses. Cure from HLH predisposition was assessed by LCMV infection. RESULTS: Hematopoietic cells isolated from transplanted mice revealed efficient gene editing (>95%), polyclonality of the T-cell receptor repertoire, and neither signs of off-target effects nor leukemogenesis. Unc13d transcription levels of edited and wild-type cells were comparable. While LCMV challenge resulted in acute HLH in Jinx mice transplanted with mock-edited HSCs, Jinx mice grafted with Unc13d-edited cells showed rapid virus clearance and protection from HLH. CONCLUSIONS: Our study demonstrates that transplantation of CRISPR-Cas edited HSCs supports the development of a functional polyclonal T-cell response in the absence of genotoxicity-associated clonal outgrowth.