The Innate Immune Response to Mycobacterium tuberculosis Infection.

Infection with Mycobacterium tuberculosis causes >1.5 million deaths worldwide annually. Innate immune cells are the first to encounter M. tuberculosis, and their response dictates the course of infection. Dendritic cells (DCs) activate the adaptive response and determine its characteristics. Macrophages are responsible both for exerting cell-intrinsic antimicrobial control and for initiating and maintaining inflammation. The inflammatory response to M. tuberculosis infection is a double-edged sword. While cytokines such as TNF-α and IL-1 are important for protection, either excessive or insufficient cytokine production results in progressive disease. Furthermore, neutrophils-cells normally associated with control of bacterial infection-are emerging as key drivers of a hyperinflammatory response that results in host mortality. The roles of other innate cells, including natural killer cells and innate-like T cells, remain enigmatic. Understanding the nuances of both cell-intrinsic control of infection and regulation of inflammation will be crucial for the successful development of host-targeted therapeutics and vaccines.

The Transcription Factor STAT6 Mediates Direct Repression of Inflammatory Enhancers and Limits Activation of Alternatively Polarized Macrophages.

The molecular basis of signal-dependent transcriptional activation has been extensively studied in macrophage polarization, but our understanding remains limited regarding the molecular determinants of repression. Here we show that IL-4-activated STAT6 transcription factor is required for the direct transcriptional repression of a large number of genes during in vitro and in vivo alternative macrophage polarization. Repression results in decreased lineage-determining transcription factor, p300, and RNA polymerase II binding followed by reduced enhancer RNA expression, H3K27 acetylation, and chromatin accessibility. The repressor function of STAT6 is HDAC3 dependent on a subset of IL-4-repressed genes. In addition, STAT6-repressed enhancers show extensive overlap with the NF-κB p65 cistrome and exhibit decreased responsiveness to lipopolysaccharide after IL-4 stimulus on a subset of genes. As a consequence, macrophages exhibit diminished inflammasome activation, decreased IL-1ß production, and pyroptosis. Thus, the IL-4-STAT6 signaling pathway establishes an alternative polarization-specific epigenenomic signature resulting in dampened macrophage responsiveness to inflammatory stimuli.

Protection of Quiescence and Longevity of IgG Memory B Cells by Mitochondrial Autophagy.

The development of long-lived immune memory cells against pathogens is critical for the success of vaccines to establish protection against future infections. However, the mechanisms governing the long-term survival of immune memory cells remain to be elucidated. In this article, we show that the maintenance mitochondrial homeostasis by autophagy is critical for restricting metabolic functions to protect IgG memory B cell survival. Knockout of mitochondrial autophagy genes, Nix and Bnip3, leads to mitochondrial accumulation and increases in oxidative phosphorylation and fatty acid synthesis, resulting in the loss of IgG+ memory B cells in mice. Inhibiting fatty acid synthesis or silencing necroptosis gene Ripk3 rescued Nix-/-Bnip3-/- IgG memory B cells, indicating that mitochondrial autophagy is important for limiting metabolic functions to prevent cell death. Our results suggest a critical role for mitochondrial autophagy in the maintenance of immunological memory by protecting the metabolic quiescence and longevity of memory B cells.

Toll-like receptor and inflammasome signals converge to amplify the innate bactericidal capacity of T helper 1 cells.

T cell effector functions can be elicited by noncognate stimuli, but the mechanism and contribution of this pathway to the resolution of intracellular macrophage infections have not been defined. Here, we show that CD4(+) T helper 1 (Th1) cells could be rapidly stimulated by microbe-associated molecular patterns during active infection with Salmonella or Chlamydia. Further, maximal stimulation of Th1 cells by lipopolysaccharide (LPS) did not require T-cell-intrinsic expression of toll-like receptor 4 (TLR4), interleukin-1 receptor (IL-1R), or interferon-γ receptor (IFN-γR) but instead required IL-18R, IL-33R, and adaptor protein MyD88. Innate stimulation of Th1 cells also required host expression of TLR4 and inflammasome components that together increased serum concentrations of IL-18. Finally, the elimination of noncognate Th1 cell stimulation hindered the resolution of primary Salmonella infection. Thus, the in vivo bactericidal capacity of Th1 cells is regulated by the response to noncognate stimuli elicited by multiple innate immune receptors.

Heterozygous Truncating Variants in POMP Escape Nonsense-Mediated Decay and Cause a Unique Immune Dysregulatory Syndrome.

The proteasome processes proteins to facilitate immune recognition and host defense. When inherently defective, it can lead to aberrant immunity resulting in a dysregulated response that can cause autoimmunity and/or autoinflammation. Biallelic or digenic loss-of-function variants in some of the proteasome subunits have been described as causing a primary immunodeficiency disease that manifests as a severe dysregulatory syndrome: chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE). Proteasome maturation protein (POMP) is a chaperone for proteasome assembly and is critical for the incorporation of catalytic subunits into the proteasome. Here, we characterize and describe POMP-related autoinflammation and immune dysregulation disease (PRAID) discovered in two unrelated individuals with a unique constellation of early-onset combined immunodeficiency, inflammatory neutrophilic dermatosis, and autoimmunity. We also begin to delineate a complex genetic mechanism whereby de novo heterozygous frameshift variants in the penultimate exon of POMP escape nonsense-mediated mRNA decay (NMD) and result in a truncated protein that perturbs proteasome assembly by a dominant-negative mechanism. To our knowledge, this mechanism has not been reported in any primary immunodeficiencies, autoinflammatory syndromes, or autoimmune diseases. Here, we define a unique hypo- and hyper-immune phenotype and report an immune dysregulation syndrome caused by frameshift mutations that escape NMD.

Successful treatment of chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) syndrome with tofacitinib.

Chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperature (CANDLE) syndrome is a rare autoinflammatory disorder. Cutaneous manifestations of CANDLE syndrome include characteristic recurring violaceous annular plaques comprised of an immature dermal mononuclear cell infiltrate. In CANDLE syndrome, deleterious genetic mutations inhibit proteasome-immunoproteasome function, resulting in cellular accumulation of ubiquitinated waste proteins that activate type I interferon signaling to drive inflammation. We describe a report of successful treatment of a 12-year-old girl with CANDLE syndrome with tofacitinib.

Modeling hospital energy and economic costs for COVID-19 infection control interventions.

The study objective assessed the energy demand and economic cost of two hospital-based COVID-19 infection control interventions: negative pressure (NP) treatment rooms and xenon pulsed ultraviolet (XP-UV) equipment. After projecting COVID-19 hospitalizations, a Hospital Energy Model and Infection De-escalation Models quantified increases in energy demand and reductions in infections. The NP intervention was applied to 11, 22, and 44 rooms for small, medium, and large hospitals, while the XP-UV equipment was used eight, nine, and ten hours a day. For small, medium, and large hospitals, the annum kWh for NP rooms were 116,700 kWh, 332,530 kWh, 795,675 kWh, which correspond to annum energy costs of $11,845 ($1,077/room), $33,752 ($1,534/room), and $80,761 ($1,836/room). For XP-UV, the annum-kilowatt-hours (and costs) were 438 ($45), 493 ($50), and 548 ($56) for small, medium, and large hospitals. While energy efficiencies may be expected for the large hospital, the hospital contained more energy-intensive use rooms (ICUs) which resulted in higher operational and energy costs. XP-UV had a greater reduction in secondary COVID-19 infections in large and medium hospitals. NP rooms had a greater reduction in secondary SARS-CoV-2 transmission in small hospitals. Early implementation of interventions can result in realized cost savings through reduced hospital-acquired infections.

Manipulation of small Rho GTPases is a pathogen-induced process detected by NOD1.

Our innate immune system distinguishes microbes from self by detecting conserved pathogen-associated molecular patterns. However, these are produced by all microbes, regardless of their pathogenic potential. To distinguish virulent microbes from those with lower disease-causing potential the innate immune system detects conserved pathogen-induced processes, such as the presence of microbial products in the host cytosol, by mechanisms that are not fully resolved. Here we show that NOD1 senses cytosolic microbial products by monitoring the activation state of small Rho GTPases. Activation of RAC1 and CDC42 by bacterial delivery or ectopic expression of SopE, a virulence factor of the enteric pathogen Salmonella, triggered the NOD1 signalling pathway, with consequent RIP2 (also known as RIPK2)-mediated induction of NF-κB-dependent inflammatory responses. Similarly, activation of the NOD1 signalling pathway by peptidoglycan required RAC1 activity. Furthermore, constitutively active forms of RAC1, CDC42 and RHOA activated the NOD1 signalling pathway. Our data identify the activation of small Rho GTPases as a pathogen-induced process sensed through the NOD1 signalling pathway.

Rapid infusion of rituximab is well tolerated in children with hematologic, oncologic, and rheumatologic disorders.

Traditional administration of rituximab requires careful titration and may involve many hours to minimize the risk of reactions. The objective of this study was to evaluate the safety of rapid infusions of rituximab in a pilot group of children with hematologic, oncologic, and rheumatologic disorders, and to determine the incidence of rate-related infusion reactions. Twenty patients enrolled in the study. All patients tolerated the rapid infusion of rituximab and no patient had an infusion-related reaction. We conclude that rapid infusions of rituximab are well tolerated and safe in our pilot group of patients.

Diagnosis and management of 365 ureteric injuries following obstetric and gynecologic surgery in resource-limited settings.

INTRODUCTION: Ureteric injuries are among the most serious complications of pelvic surgery. The incidence in low-resource settings is not well documented. METHODS: This retrospective review analyzes a cohort of 365 ureteric injuries with ureterovaginal fistulas in 353 women following obstetric and gynecologic operations in 11 countries in Africa and Asia, all low-resource settings. The patients with ureteric injury were stratified into three groups according to the initial surgery: (a) obstetric operations, (b) gynecologic operations, and (c) vesicovaginal fistula (VVF) repairs. RESULTS: The 365 ureteric injuries in this series comprise 246 (67.4%) after obstetric procedures, 65 (17.8%) after gynecologic procedures, and 54 (14.8%) after repair of obstetric fistulas. Demographic characteristics show clear differences between women with iatrogenic injuries and women with obstetric fistulas. The study describes abdominal ureter reimplantation and other treatment procedures. Overall surgical results were good: 92.9% of women were cured (326/351), 5.4% were healed with some residual incontinence (19/351), and six failed (1.7%). CONCLUSIONS: Ureteric injuries after obstetric and gynecologic operations are not uncommon. Unlike in high-resource contexts, in low-resource settings obstetric procedures are most often associated with urogenital fistula. Despite resource limitations, diagnosis and treatment of ureteric injuries is possible, with good success rates. Training must emphasize optimal surgical techniques and different approaches to assisted vaginal delivery.

Iatrogenic genitourinary fistula: an 18-year retrospective review of 805 injuries.

INTRODUCTION AND HYPOTHESIS: Genitourinary fistula poses a public health challenge in areas where women have inadequate access to quality emergency obstetric care. Fistulas typically develop during prolonged, obstructed labor, but providers can also inadvertently cause a fistula when performing obstetric or gynecological surgery. METHODS: This retrospective study analyzes 805 iatrogenic fistulas from a series of 5,959 women undergoing genitourinary fistula repair in 11 countries between 1994 and 2012. Injuries fall into three categories: ureteric, vault, and vesico-[utero]/-cervico-vaginal. This analysis considers the frequency and characteristics of each type of fistula and the risk factors associated with iatrogenic fistula development. RESULTS: In this large series, 13.2 % of genitourinary fistula repairs were for injuries caused by provider error. A range of cadres conducted procedures resulting in iatrogenic fistula. Four out of five iatrogenic fistulas developed following surgery for obstetric complications: cesarean section, ruptured uterus repair, or hysterectomy for ruptured uterus. Others developed during gynecological procedures, most commonly hysterectomy. Vesico-[utero]/-cervico-vaginal fistulas were the most common (43.6 %), followed by ureteric injuries (33.9 %) and vault fistulas (22.5 %). One quarter of women with iatrogenic fistulas had previously undergone a laparotomy, nearly always a cesarean section. Among these women, one quarter had undergone more than one previous cesarean section. CONCLUSIONS: Women with previous cesarean sections are at an increased risk of iatrogenic injury. Work environments must be adequate to reduce surgical error. Training must emphasize the importance of optimal surgical techniques, obstetric decision-making, and alternative ways to deliver dead babies. Iatrogenic fistulas should be recognized as a distinct genitourinary fistula category.

Th17/1 and ex-Th17 cells are detected in patients with polyarticular juvenile arthritis and increase following treatment.

BACKGROUND: A better understanding of the pathogenesis of polyarticular juvenile idiopathic arthritis (polyJIA) is needed to aide in the development of data-driven approaches to guide selection between therapeutic options. One inflammatory pathway of interest is JAK-STAT signaling. STAT3 is a transcription factor critical to the differentiation of inflammatory T helper 17 cells (Th17s). Previous studies have demonstrated increased STAT3 activation in adult patients with rheumatoid arthritis, but less is known about STAT3 activation in polyJIA. We hypothesized that Th17 cells and STAT3 activation would be increased in treatment-naïve polyJIA patients compared to pediatric controls. METHODS: Blood from 17 patients with polyJIA was collected at initial diagnosis and again if remission was achieved (post-treatment). Pediatric healthy controls were also collected. Peripheral blood mononuclear cells were isolated and CD4 + T cell subsets and STAT activation (phosphorylation) were evaluated using flow cytometry. Data were analyzed using Mann-Whitney U and Wilcoxon matched-pairs signed rank tests. RESULTS: Treatment-naïve polyJIA patients had increased Th17 cells (CD3 + CD4 + interleukin(IL)-17 +) compared to controls (0.15% v 0.44%, p < 0.05), but Tregs (CD3 + CD4 + CD25 + FOXP3 +) from patients did not differ from controls. Changes in STAT3 phosphorylation in CD4 + T cells following ex vivo stimulation were not significantly different in patients compared to controls. We identified dual IL-17 + and interferon (IFN)γ + expressing CD4 + T cells in patients, but not controls. Further, both Th17/1 s (CCR6 + CD161 + IFNγ + IL-17 +) and ex-Th17s (CCR6 + CD161 + IFNγ + IL-17neg) were increased in patients' post-treatment (Th17/1: 0.3% v 0.07%, p < 0.05 and ex-Th17s: 2.3% v 1.4%, p < 0.05). The patients with the highest IL-17 expressing cells post-treatment remained therapy-bound. CONCLUSIONS: Patients with polyJIA have increased baseline Th17 cells, potentially reflecting higher tonic STAT3 activation in vivo. These quantifiable immune markers may identify patients that would benefit upfront from pathway-focused biologic therapies. Our data also suggest that inflammatory CD4 + T cell subsets not detected in controls but increased in post-treatment samples should be further evaluated as a tool to stratify patients in remission on medication. Future work will explore these proposed diagnostic and prognostic biomarkers.

Childhood Arthritis and Rheumatology Research Alliance Biologic DMARD Consensus Treatment Plans for Refractory Moderately Severe Juvenile Dermatomyositis.

OBJECTIVE: The objective was to develop consensus treatment plans (CTPs) for patients with refractory moderately severe juvenile dermatomyositis (JDM) treated with biologic disease-modifying antirheumatic drugs (bDMARDs). METHODS: The Biologics Workgroup of the Childhood Arthritis and Rheumatology Research Alliance (CARRA) JDM Research Committee used case-based surveys, consensus framework, and nominal group technique to produce bDMARD CTPs for patients with refractory moderately severe JDM. RESULTS: Four bDMARD CTPs were proposed: TNF-alpha inhibitor (adalimumab or infliximab), abatacept, rituximab, and tocilizumab. Each CTP has different options for dosing and/or route. Among 76 respondents, consensus was achieved for the proposed CTPs (93% [67/72]) as well as for patient characteristics, assessments, outcome measures, and follow up. By weighted average, respondents indicated that they would most likely use rituximab followed by abatacept, TNF-alpha inhibitor, and tocilizumab. CONCLUSION: CTPs for the use of bDMARDs in refractory moderately severe JDM were developed using consensus methodology. The implementation of the bDMARD CTPs will lay the groundwork for registry-based prospective comparative effectiveness studies.

Ragweed pollen extract intensifies lipopolysaccharide-induced priming of NLRP3 inflammasome in human macrophages.

Ragweed pollen extract (RWE) possesses intrinsic NADPH oxidase activity that induces oxidative stress by initiating the production of intracellular reactive oxygen species (ROS). The ROS are important contributors to the manifestation of allergic inflammation; furthermore, concomitant exposure to an allergen and an endotoxin trigger a stronger inflammatory response. One of the main pro-inflammatory cytokines produced in inflammatory responses is interleukin-1ß (IL-1ß), and its production is associated with caspase-1-containing inflammasome complexes. Intracellular ROS have been implicated in NLRP3 inflammasome-mediated IL-1ß production, therefore, we aimed to study whether RWE influences the function of NLRP3 inflammasome. Here we describe that, in the presence of NADPH, RWE significantly elevates lipopolysaccharide-induced IL-1ß production of THP-1 cells as well as human primary macrophages and dendritic cells. We also demonstrate that increased IL-1ß production is mediated through NLRP3 inflammasome in THP-1 macrophages. We provide evidence that RWE elevates cytosolic ROS level in these cells, and ROS inhibitors abolish IL-1ß production. Furthermore, we show that RWE enhances lipopolysaccharide-induced gene transcription/expression of pro-IL-1ß and key components of the inflammasome via a ROS-dependent mechanism.

Solvent detergent vs. fresh frozen plasma in cirrhotic patients undergoing liver transplant surgery: a prospective randomized control study.

BACKGROUND: Although orthotopic liver transplantation (OLT) is nowadays considered standard practice at experienced centres, it can still be affected by a significant risk of massive bleeding and its related complications. Solvent/detergent plasma (S/D Plasma) has been proposed as an alternative to fresh frozen plasma (FFP) to curtail such complications. This study aimed at evaluating the efficacy of S/D Plasma in OLT patients by comparing it to FFP. MATERIALS AND METHODS: Sixty-three OLT patients were randomized into two groups depending on whether they were transfused with FFP or S/D plasma. A thromboelastography-based protocol aimed at achieving and maintaining predetermined coagulation goals was used to guide plasma transfusions. At the beginning and the end of surgery, standard laboratory coagulation tests were performed together with the assessment of the VII, VIII, V, XII factors and S protein blood levels. RESULTS: The two study groups equally achieved the thromboelastography goals but with a reduced amount of transfusions in the S/D plasma group (P < 0.0001). At the end of surgery, factors V and XII and S protein blood levels were lower in the S/D plasma patients who also showed lower INR, aPTT and antithrombin III levels. CONCLUSION: In cirrhotic patients undergoing OLT, the use of S\D plasma associated with thromboelastography allows the same clinical results but with a significant reduction in the amount of plasma transfusions.

Targeting HIV-1 reservoirs in T cell subsets.

The HIV-1 reservoirs harbor the latent proviruses that are integrated into the host genome. It is a challenging task to eradicate the proviruses in order to achieve an HIV cure. We have described a strategy for the clearance of HIV-1 infection through selective elimination of host cells harboring replication-competent HIV (SECH), by inhibition of autophagy and promotion of apoptosis during viral re-activation. HIV-1 can infect various CD4+ T cell subsets, but it is not known whether the SECH approach is equally effective in targeting HIV-1 reservoirs in these different subsets in vivo. In a humanized mouse model, we found that treatments of HIV-1 infection by suppressive antiretroviral therapy (ART) led to the establishment of latent HIV reservoirs in naïve, central memory and effector memory T cells. Moreover, SECH treatments could clear latent HIV-1 reservoirs in these different T cell subsets of humanized mice. Co-culture studies showed that T cell subsets latently infected by HIV-1, but not uninfected bystander cells, were susceptible to cell death induced by SECH treatments. Our study suggests that the SECH strategy is effective for specific targeting of latent HIV-1 reservoirs in different T cell subsets.

Identification of eight novel proteasome variants in five unrelated cases of proteasome-associated autoinflammatory syndromes (PRAAS).

Mutations in genes coding for proteasome subunits and/or proteasome assembly helpers typically cause recurring autoinflammation referred to as chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperatures (CANDLE) or proteasome-associated autoinflammatory syndrome (PRAAS). Patients with CANDLE/PRAAS present with mostly chronically elevated type I interferon scores that emerge as a consequence of increased proteotoxic stress by mechanisms that are not fully understood. Here, we report on five unrelated patients with CANDLE/PRAAS carrying novel inherited proteasome missense and/or nonsense variants. Four patients were compound heterozygous for novel pathogenic variants in the known CANDLE/PRAAS associated genes, PSMB8 and PSMB10, whereas one patient showed additive loss-of-function mutations in PSMB8. Variants in two previously not associated proteasome genes, PSMA5 and PSMC5, were found in a patient who also carried the PSMB8 founder mutation, p.T75M. All newly identified mutations substantially impact the steady-state expression of the affected proteasome subunits and/or their incorporation into mature 26S proteasomes. Our observations expand the spectrum of PRAAS-associated genetic variants and improve a molecular diagnosis and genetic counseling of patients with sterile autoinflammation.

Distal extremity necrosis as a manifestation of cutaneous polyarteritis nodosa: case report and review of the acute management of a pediatric patient.

We present the case of an 8-year-old girl who presented with distal extremity necrosis of the hands, feet, nose, and ears as an acute manifestation of cutaneous polyarteritis nodosa (CPAN). She was emergently managed with intravenous steroids, nifedipine, sildenafil, pentoxifylline, nitroglycerin paste, aspirin, low-molecular-weight heparin, and intravenous gamma globulin. The necrosis was controlled, and reperfusion was attained to salvage the extremities. It is important for clinicians to be aware that acute distal extremity necrosis can be a manifestation of CPAN in children. Etiology is often not clear on presentation, but once infection is excluded, acute management with systemic steroids and systemic vasodilators is indicated regardless of the cause. Iloprost and bosentan may represent options for adjunctive vasodilators. More studies are needed to create guidelines for the acute and long-term management of these children. Close follow-up of children with CPAN, especially with a history of vaso-occlusive symptoms, is important to allow prompt intervention in the event of distal extremity infarction.

Mitochondrion-Mediated Cell Death through Erk1-Alox5 Independent of Caspase-9 Signaling.

Mitochondrial disruption leads to the release of cytochrome c to activate caspase-9 and the downstream caspase cascade for the execution of apoptosis. However, cell death can proceed efficiently in the absence of caspase-9 following mitochondrial disruption, suggesting the existence of caspase-9-independent cell death mechanisms. Through a genome-wide siRNA library screening, we identified a network of genes that mediate caspase-9-independent cell death, through ROS production and Alox5-dependent membrane lipid peroxidation. Erk1-dependent phosphorylation of Alox5 is critical for targeting Alox5 to the nuclear membrane to mediate lipid peroxidation, resulting in nuclear translocation of cytolytic molecules to induce DNA damage and cell death. Consistently, double knockouts of caspase-9 and Alox5 in mice, but not deletion of either gene alone, led to significant T cell expansion with inhibited cell death, indicating that caspase-9- and Alox5-dependent pathways function in parallel to regulate T cell death in vivo. This unbiased whole-genome screening reveals an Erk1-Alox5-mediated pathway that promotes membrane lipid peroxidation and nuclear translocation of cytolytic molecules, leading to the execution of cell death in parallel to the caspase-9 signaling cascade.