Issue published October 1, 2020 Previous issue | Next issue
- Volume 130, Issue 10
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On the cover: Hemolysis-transformed macrophages mitigate sterile inflammation
Liver macrophages that phagocytose lysed red blood cells (RBCs) play an important role in clearing hemoglobin and heme, which trigger inflammation, oxidative stress, and nitric oxide depletion when released into plasma and tissue. In this issue, Pfefferlé, Ingoglia, et al. identify a specialized erythrophagocytic and anti-inflammatory phenotype in liver macrophages that is driven by increased hemolysis in a mouse model of spherocytosis. In models of sterile inflammatory disease, erythrocytosis-skewed macrophages attenuated disease severity, suggesting that hemolysis is driving an adaptive transformation in liver macrophages. The cover image is an impressionistic rendition of erythrophagocytes lining sinusoidal capillaries in the liver. These specialized macrophages (brown) limit the potential adverse effects of excessive RBC lysis (red). Image credit: Rok Humar.
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Review Series
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Abstract
Intermittent hypoxia (IH) is a hallmark manifestation of obstructive sleep apnea (OSA), a widespread disorder of breathing. This Review focuses on the role of hypoxia-inducible factors (HIFs) in hypertension, type 2 diabetes (T2D), and cognitive decline in experimental models of IH patterned after O2 profiles seen in OSA. IH increases HIF-1α and decreases HIF-2α protein levels. Dysregulated HIFs increase reactive oxygen species (ROS) through HIF-1–dependent activation of pro-oxidant enzyme genes in addition to reduced transcription of antioxidant genes by HIF-2. ROS in turn activate chemoreflex and suppress baroreflex, thereby stimulating the sympathetic nervous system and causing hypertension. We also discuss how increased ROS generation by HIF-1 contributes to IH-induced insulin resistance and T2D as well as disrupted NMDA receptor signaling in the hippocampus, resulting in cognitive decline.
Authors
Nanduri R. Prabhakar, Ying-Jie Peng, Jayasri Nanduri
×Abstract
The liver has strong innate immunity to counteract pathogens from the gastrointestinal tract. During the development of liver cancer, which is typically driven by chronic inflammation, the composition and biological roles of the innate immune cells are extensively altered. Hypoxia is a common finding in all stages of liver cancer development. Hypoxia drives the stabilization of hypoxia-inducible factors (HIFs), which act as central regulators to dampen the innate immunity of liver cancer. HIF signaling in innate immune cells and liver cancer cells together favors the recruitment and maintenance of pro-tumorigenic immune cells and the inhibition of anti-tumorigenic immune cells, promoting immune evasion. HIFs represent attractive therapeutic targets to inhibit the formation of an immunosuppressive microenvironment and growth of liver cancer.
Authors
Vincent Wai-Hin Yuen, Carmen Chak-Lui Wong
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Hypoxia can be defined as a relative deficiency in the amount of oxygen reaching the tissues. Hypoxia-inducible factors (HIFs) are critical regulators of the mammalian response to hypoxia. In normal circumstances, HIF-1α protein turnover is rapid, and hyperglycemia further destabilizes the protein. In addition to their role in diabetes pathogenesis, HIFs are implicated in development of the microvascular and macrovascular complications of diabetes. Improving glucose control in people with diabetes increases HIF-1α protein and has wide-ranging benefits, some of which are at least partially mediated by HIF-1α. Nevertheless, most strategies to improve diabetes or its complications via regulation of HIF-1α have not currently proven to be clinically useful. The intersection of HIF biology with diabetes is a complex area in which many further questions remain, especially regarding the well-conducted studies clearly describing discrepant effects of different methods of increasing HIF-1α, even within the same tissues. This Review presents a brief overview of HIFs; discusses the range of evidence implicating HIFs in β cell dysfunction, diabetes pathogenesis, and diabetes complications; and examines the differing outcomes of HIF-targeting approaches in these conditions.
Authors
Jenny E. Gunton
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Hypoxia-inducible factors (HIFs) and the HIF-dependent cancer hallmarks angiogenesis and metabolic rewiring are well-established drivers of breast cancer aggressiveness, therapy resistance, and poor prognosis. Targeting of HIF and its downstream targets in angiogenesis and metabolism has been unsuccessful so far in the breast cancer clinical setting, with major unresolved challenges residing in target selection, development of robust biomarkers for response prediction, and understanding and harnessing of escape mechanisms. This Review discusses the pathophysiological role of HIFs, angiogenesis, and metabolism in breast cancer and the challenges of targeting these features in patients with breast cancer. Rational therapeutic combinations, especially with immunotherapy and endocrine therapy, seem most promising in the clinical exploitation of the intricate interplay of HIFs, angiogenesis, and metabolism in breast cancer cells and the tumor microenvironment.
Authors
Ellen C. de Heer, Mathilde Jalving, Adrian L. Harris
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Review
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Abstract
Epithelial cell dysfunction has emerged as a central component of the pathophysiology of diffuse parenchymal diseases including idiopathic pulmonary fibrosis (IPF). Alveolar type 2 (AT2) cells represent a metabolically active lung cell population important for surfactant biosynthesis and alveolar homeostasis. AT2 cells and other distal lung epithelia, like all eukaryotic cells, contain an elegant quality control network to respond to intrinsic metabolic and biosynthetic challenges imparted by mutant protein conformers, dysfunctional subcellular organelles, and dysregulated telomeres. Failed AT2 quality control components (the ubiquitin-proteasome system, unfolded protein response, macroautophagy, mitophagy, and telomere maintenance) result in diverse cellular endophenotypes and molecular signatures including ER stress, defective autophagy, mitochondrial dysfunction, apoptosis, inflammatory cell recruitment, profibrotic signaling, and altered progenitor function that ultimately converge to drive downstream fibrotic remodeling in the IPF lung. As this complex network becomes increasingly better understood, opportunities will emerge to identify targets and therapeutic strategies for IPF.
Authors
Jeremy Katzen, Michael F. Beers
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Commentaries
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Abstract
The tight junction protein claudin-2 is upregulated in inflammatory bowel disease, and yet its deficit worsens infectious and chemical colitis. In this issue of the JCI, Raju and Shashikanth et al. examined the contribution of claudin-2 to immune-mediated colitis. The authors used transgenic mouse models to show that claudin-2 deficiency attenuated colitis progression as well as a leak barrier defect, albeit at the risk of intestinal obstruction. Further, inhibition of claudin-2 by targeting casein kinase 2 (CK2) also ameliorated colitis. The findings reveal unsuspected links between the pore and leak pathways of intestinal permeability and immune responses leading to colitis. They additionally suggest potential targets for therapeutic intervention in inflammatory bowel disease.
Authors
Kim E. Barrett
×Abstract
Even with treatment of HIV with antiretroviral therapy (ART), the risk of tuberculosis (TB) reactivation remains higher in HIV-infected than HIV-uninfected persons. In this issue of the JCI, Ganatra et al. explored TB reactivation in the context of ART using TB and simian immunodeficiency virus–coinfected (TB/SIV-coinfected) nonhuman primates. The authors found that treating rhesus macaques with ART restored CD4+ T cells in whole blood, spleen, and bronchoalveolar lavage (BAL) fluid, but not in the lung interstitium. TB risk was not decreased in the coinfected macaques treated with ART for 14–63 days, suggesting that ART does not decrease the short-term risk of reactivation. Reactivation occurred as CD4+ T cells were increasing, which is consistent with observations made in humans. This study provides a potential model for systematic evaluation of TB/SIV coinfection and different treatment regimens and strategies to prevent TB reactivation.
Authors
Timothy R. Sterling, Philana Ling Lin
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Idiopathic CD4+ T cell lymphocytopenia (ICL) is a heterogeneous syndrome presenting with persistent CD4+ T cell lymphopenia of unknown origin, and opportunistic infections in some patients. The underlying pathogenesis and appropriate management remain understudied. In this issue of the JCI, Perez-Diez and Wong et al. assessed the prevalence of autoantibodies from the sera of 51 adult ICL patients (out of a cohort of 72). Some patients showed high levels of IgG and IgM autoantibodies against numerous autoantigens, and some autoantibodies were specific for lymphocytes. The researchers implicate these autoantibodies as a possible pathogenic mechanism responsible for the reduction in circulating CD4+ T cells. This study goes beyond defining a mechanism in a complex, poorly defined disease; it also brings a renewed focus on ICL that will likely result in improved diagnostic evaluation and treatment.
Authors
Jose S. Campos, Sarah E. Henrickson, Roshini S. Abraham
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The right ventricle (RV) is involved in systemic circulation in the fetal mammalian heart but quickly transitions to being solely responsible for pulmonary circulation after birth when the left ventricle (LV) becomes the systemic ventricle. To handle the increased workload, LV growth greatly outpaces that of the RV during postnatal stages. However, the molecular basis for this differential growth pattern between the 2 chambers is largely unknown. In this issue of the JCI, Yokota et al. reveal that the p38 mitogen-activated protein kinase (MAPK)/IRE1α/XBP1 axis specifically controls postnatal RV growth by suppressing cell cycle regulatory genes.
Authors
Tongbin Wu, Ju Chen
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Most patients with COVID-19 lack antibody to SARS-CoV-2 in the first 10 days of illness while the virus drives disease pathogenesis. SARS-CoV-2 antibody deficiency in the setting of a tissue viral burden suggests that using an antibody as a therapeutic agent would augment the antiviral immune response. In this issue of the JCI, Wang and collaborators describe the kinetics of viral load and the antibody responses of 23 individuals with COVID-19 experiencing mild and severe disease. The researchers found that (a) individuals with mild and severe disease produced neutralizing IgG to SARS-CoV-2 10 days after disease onset, (b) SARS-CoV-2 persisted longer in those with severe disease, and (c) there was cross-reactivity between antibodies to SARS-CoV-1 and SARS-CoV-2, but only antibodies from patients with COVID-19 neutralized SARS-CoV-2. These observations provide important information on the serological response to SARS-CoV-2 of hospitalized patients with COVID-19 that can inform the use of convalescent plasma therapy.
Authors
Arturo Casadevall, Michael J. Joyner, Liise-anne Pirofski
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NK cells are responsible for defense against viral infections and cancer. Although activated NK cells are armed to combat tumors, the tumor microenvironment (TME) contains ROS, which suppress NK cell antitumor activity. In this issue of the JCI, Yang, Neo, and colleagues explored NK cell resistance to oxidative stress in vitro and in human non–small-cell lung cancer (NSCLC). High surface thiol density and elevated expression of the ROS scavenger thioredoxin (Trx1) protected NK cells from ROS. Trx1 and thiol levels were higher in IL-15– than in IL-2–primed NK cells. Tumor-infiltrating Trx1+ NK cells were present in patients with NSCLC with elevated ROS levels in the tumor. Smokers scored higher for the ROS signature, which predicted poor prognosis, compared with nonsmokers. This study explains how activated NK cells survive in the ROS-rich TME and suggests that smokers with lung cancer may benefit from therapies using IL-15–primed NK cells.
Authors
Theresa L. Whiteside
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Allergic disorders include food allergy, allergic rhinitis, and certain forms of asthma resulting from the inappropriate development of immune responses to otherwise innocuous aeroallergens and foods. In this issue of the JCI, Thouvenot and Roitel et al. explore transcription infidelity as a mechanism that underlies the ability of these benign proteins to become allergens. Some foods and bioaerosols that produce allergies have RNA polymerase with a propensity to generate RNA gaps, thereby causing translational frameshifts. These frameshifts often create cationic carboxy-terminus residues that replace hydrophobic amino acids and have enhanced MHC binding, resulting in the tendency to provoke immune responses. IgE antibody responses initiated by these variant transcripts can later lead to IgE against the native molecule and also explain how anaphylaxis may occur in individuals who lacked specific IgE when tested using native protein reagents. This study has the potential to transform the diagnosis and treatment of allergic disorders.
Authors
Larry Borish
×Abstract
Bacillus Calmette-Guérin (BCG) vaccination induces variable protection against pulmonary tuberculosis (TB), and a more effective TB vaccine is needed. The potential for BCG to provide protection against heterologous infections, by induction of innate immune memory, is increasingly recognized. These nonspecific responses may substantially benefit public health, but are also variable. In this issue of the JCI, Koeken and de Bree et al. report that BCG reduces circulating inflammatory markers in males but not in females, while de Bree and Mouritis et al. describe how diurnal rhythms affect the degree of BCG-induced innate memory. These studies further delineate factors that influence the magnitude of responses to BCG and may be crucial to harnessing its potential benefits.
Authors
S. Prentice, H.M. Dockrell
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Retinoic acid (RA) signaling is involved in various physiological and pathological conditions, including development, tumorigenesis, inflammation, and tissue damage and repair. In kidneys, the beneficial effect of RA has been reported in multiple disease models, such as glomerulosclerosis, renal fibrosis, and acute kidney injury. In this issue of the JCI, Chen et al. report a pathway activated by RA signaling that is mediated by the retinoic acid receptor responder protein 1 (RARRES1). Specifically, RARRES1, which is proteolytically cleaved to release the extracellular domain, was endocytosed by podocytes to induce apoptosis and glomerular dysfunction kidney disease. These findings unveil the contrasting aspects, a Janus face, of RA signaling that may guide its therapeutic use.
Authors
Qingqing Wei, Zheng Dong
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Research Articles
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Abstract
Proteins created from recurrent fusion genes like CBFB-MYH11 are prevalent in acute myeloid leukemia (AML), often necessary for leukemogenesis, persistent throughout the disease course, and highly leukemia specific, making them attractive neoantigen targets for immunotherapy. A nonameric peptide derived from a prevalent CBFB-MYH11 fusion protein was found to be immunogenic in HLA-B*40:01+ donors. High-avidity CD8+ T cell clones isolated from healthy donors killed CBFB-MYH11+ HLA-B*40:01+ AML cell lines and primary human AML samples in vitro. CBFB-MYH11–specific T cells also controlled CBFB-MYH11+ HLA-B*40:01+ AML in vivo in a patient-derived murine xenograft model. High-avidity CBFB-MYH11 epitope–specific T cell receptors (TCRs) transduced into CD8+ T cells conferred antileukemic activity in vitro. Our data indicate that the CBFB-MYH11 fusion neoantigen is naturally presented on AML blasts and enables T cell recognition and killing of AML. We provide proof of principle for immunologically targeting AML-initiating fusions and demonstrate that targeting neoantigens has clinical relevance even in low–mutational frequency cancers like fusion-driven AML. This work also represents a first critical step toward the development of TCR T cell immunotherapy targeting fusion gene–driven AML.
Authors
Melinda A. Biernacki, Kimberly A. Foster, Kyle B. Woodward, Michael E. Coon, Carrie Cummings, Tanya M. Cunningham, Robson G. Dossa, Michelle Brault, Jamie Stokke, Tayla M. Olsen, Kelda Gardner, Elihu Estey, Soheil Meshinchi, Anthony Rongvaux, Marie Bleakley
×Abstract
Essential tremor is a common brain disorder affecting millions of people, yet the neuronal mechanisms underlying this prevalent disease remain elusive. Here, we showed that conditional deletion of synaptotagmin-2, the fastest Ca2+ sensor for synaptic neurotransmitter release, from parvalbumin neurons in mice caused an action tremor syndrome resembling the core symptom of essential tremor patients. Combining brain region–specific and cell type–specific genetic manipulation methods, we found that deletion of synaptotagmin-2 from excitatory parvalbumin-positive neurons in cerebellar nuclei was sufficient to generate an action tremor. The synaptotagmin-2 deletion converted synchronous into asynchronous neurotransmitter release in projections from cerebellar nuclei neurons onto gigantocellular reticular nucleus neurons, which might produce an action tremor by causing signal oscillations during movement. The tremor was rescued by completely blocking synaptic transmission with tetanus toxin in cerebellar nuclei, which also reversed the tremor phenotype in the traditional harmaline-induced essential tremor model. Using a promising animal model for action tremor, our results thus characterized a synaptic circuit mechanism that may underlie the prevalent essential tremor disorder.
Authors
Mu Zhou, Maxwell D. Melin, Wei Xu, Thomas C. Südhof
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The correlation of HIV-specific antibody-dependent cellular cytotoxicity (ADCC) responses with protection from and delayed progression of HIV-1 infection provides a rationale to leverage ADCC-mediating antibodies for treatment purposes. We evaluated ADCC mediated by different combinations of 2 to 6 neutralizing and non-neutralizing anti–HIV-1 Envelope (Env) mAbs, using concentrations ≤ 1 μg/mL, to identify combinations effective at targeting latent reservoir HIV-1 viruses from 10 individuals. We found that within 2 hours, combinations of 3 mAbs mediated more than 30% killing of HIV-infected primary CD4+ T cells in the presence of autologous NK cells, with the combination of A32 (C1C2), DH511.2K3 (MPER), and PGT121 (V3) mAbs being the most effective. Increasing the incubation of target and effector cells in the presence of mAb combinations from 2 to 24 hours resulted in increased specific killing of infected cells, even with neutralization-resistant viruses. The same combination eliminated reactivated latently HIV-1–infected cells in an ex vivo quantitative viral outgrowth assay. Therefore, administration of a combination of 3 mAbs should be considered in planning in vivo studies seeking to eliminate persistently HIV-1–infected cells.
Authors
Marina Tuyishime, Carolina Garrido, Shalini Jha, Matt Moeser, Dieter Mielke, Celia LaBranche, David Montefiori, Barton F. Haynes, Sarah Joseph, David M. Margolis, Guido Ferrari
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While the advent of combination antiretroviral therapy (ART) has significantly improved survival, tuberculosis (TB) remains the leading cause of death in the HIV-infected population. We used Mycobacterium tuberculosis/simian immunodeficiency virus–coinfected (M. tuberculosis/SIV–coinfected) macaques to model M. tuberculosis/HIV coinfection and study the impact of ART on TB reactivation due to HIV infection. Although ART significantly reduced viral loads and increased CD4+ T cell counts in blood and bronchoalveolar lavage (BAL) samples, it did not reduce the relative risk of SIV-induced TB reactivation in ART-treated macaques in the early phase of treatment. CD4+ T cells were poorly restored specifically in the lung interstitium, despite their significant restoration in the alveolar compartment of the lung as well as in the periphery. IDO1 induction in myeloid cells in the inducible bronchus-associated lymphoid tissue (iBALT) likely contributed to dysregulated T cell homing and impaired lung immunity. Thus, although ART was indispensable for controlling viral replication, restoring CD4+ T cells, and preventing opportunistic infection, it appeared inadequate in reversing the clinical signs of TB reactivation during the relatively short duration of ART administered in this study. This finding warrants the modeling of concurrent treatment of TB and HIV to potentially reduce the risk of reactivation of TB due to HIV to inform treatment strategies in patients with M. tuberculosis/HIV coinfection.
Authors
Shashank R. Ganatra, Allison N. Bucşan, Xavier Alvarez, Shyamesh Kumar, Ayan Chatterjee, Melanie Quezada, Abigail Fish, Dhiraj K. Singh, Bindu Singh, Riti Sharan, Tae-Hyung Lee, Uma Shanmugasundaram, Vijayakumar Velu, Shabaana A. Khader, Smriti Mehra, Jyothi Rengarajan, Deepak Kaushal
×Abstract
Psoriasis is a severe disease associated with the disturbance of metabolism and inflammation, but the molecular mechanisms underlying these aspects of psoriasis pathology are poorly understood. Here, we report that glutaminase 1–mediated (GLS1-mediated) glutaminolysis was aberrantly activated in patients with psoriasis and in psoriasis-like mouse models, which promoted Th17 and γδ T17 (IL-17A–producing γδ T) cell differentiation through enhancement of histone H3 acetylation of the Il17a promoter, thereby contributing to the immune imbalance and development of psoriasis. We further demonstrate that mucosa-associated lymphoid tissue lymphoma translocation protein 1 (MALT1) protease was constitutively active in psoriatic CD4+ and γδ T cells, thereby supporting GLS1 expression by stabilizing c-Jun, which directly binds to the GLS1 promoter region. Blocking the activity of either GLS1 or MALT1 protease resolved Th17 and γδ T17 cell differentiation and epidermal hyperplasia in the psoriasis-like mouse models. Finally, IL-17A enhanced GLS1 expression via the MALT1/cJun pathway in keratinocytes, resulting in hyperproliferation of and chemokine production by keratinocytes. Our findings identify the role of the MALT1/cJun/GLS1/glutaminolysis/H3 acetylation/T17 axis in psoriasis pathogenesis and reveal potential therapeutic targets for this disease.
Authors
Xichun Xia, Guangchao Cao, Guodong Sun, Leqing Zhu, Yixia Tian, Yueqi Song, Chengbin Guo, Xiao Wang, Jingxiang Zhong, Wei Zhou, Peng Li, Hua Zhang, Jianlei Hao, Zhizhong Li, Liehua Deng, Zhinan Yin, Yunfei Gao
×Abstract
The tight junction protein claudin-2 is upregulated in disease. Although many studies have linked intestinal barrier loss to local and systemic disease, these have relied on macromolecular probes. In vitro analyses show, however, that these probes cannot be accommodated by size- and charge-selective claudin-2 channels. We sought to define the impact of claudin-2 channels on disease. Transgenic claudin-2 overexpression or IL-13–induced claudin-2 upregulation increased intestinal small cation permeability in vivo. IL-13 did not, however, affect permeability in claudin-2–knockout mice. Claudin-2 is therefore necessary and sufficient to effect size- and charge-selective permeability increases in vivo. In chronic disease, T cell transfer colitis severity was augmented or diminished in claudin-2–transgenic or –knockout mice, respectively. We translated the in vitro observation that casein kinase-2 (CK2) inhibition blocks claudin-2 channel function to prevent acute, IL-13–induced, claudin-2–mediated permeability increases in vivo. In chronic immune-mediated colitis, CK2 inhibition attenuated progression in claudin-2–sufficient, but not claudin-2–knockout, mice, i.e., the effect was claudin-2 dependent. Paracellular flux mediated by claudin-2 channels can therefore promote immune-mediated colitis progression. Although the mechanisms by which claudin-2 channels intensify disease remain to be defined, these data suggest that claudin-2 may be an accessible target in immune-mediated disorders, including inflammatory bowel disease.
Authors
Preeti Raju, Nitesh Shashikanth, Pei-Yun Tsai, Pawin Pongkorpsakol, Sandra Chanez-Paredes, Peter R. Steinhagen, Wei-Ting Kuo, Gurminder Singh, Sachiko Tsukita, Jerrold R. Turner
×Abstract
BACKGROUND Although mania is characteristic of bipolar disorder, it can also occur following focal brain damage. Such cases may provide unique insight into brain regions responsible for mania symptoms and identify therapeutic targets.METHODS Lesion locations associated with mania were identified using a systematic literature search (n = 41) and mapped onto a common brain atlas. The network of brain regions functionally connected to each lesion location was computed using normative human connectome data (resting-state functional MRI, n = 1000) and contrasted with those obtained from lesion locations not associated with mania (n = 79). Reproducibility was assessed using independent cohorts of mania lesions derived from clinical chart review (n = 15) and of control lesions (n = 490). Results were compared with brain stimulation sites previously reported to induce or relieve mania symptoms.RESULTS Lesion locations associated with mania were heterogeneous and no single brain region was lesioned in all, or even most, cases. However, these lesion locations showed a unique pattern of functional connectivity to the right orbitofrontal cortex, right inferior temporal gyrus, and right frontal pole. This connectivity profile was reproducible across independent lesion cohorts and aligned with the effects of therapeutic brain stimulation on mania symptoms.CONCLUSION Brain lesions associated with mania are characterized by a specific pattern of brain connectivity that lends insight into localization of mania symptoms and potential therapeutic targets.FUNDING Fundação para a Ciência e Tecnologia (FCT), Harvard Medical School DuPont-Warren Fellowship, Portuguese national funds from FCT and Fundo Europeu de Desenvolvimento Regional, Child Neurology Foundation Shields Research, Sidney R. Baer, Jr. Foundation, Nancy Lurie Marks Foundation, Mather’s Foundation, and the NIH.
Authors
Gonçalo Cotovio, Daniel Talmasov, J. Bernardo Barahona-Corrêa, Joey Hsu, Suhan Senova, Ricardo Ribeiro, Louis Soussand, Ana Velosa, Vera Cruz e Silva, Natalia Rost, Ona Wu, Alexander L. Cohen, Albino J. Oliveira-Maia, Michael D. Fox
×Abstract
Dengue virus (DENV) infection requires cholesterol as a proviral factor, although statin treatment did not show antiviral efficacy in patients with dengue. Here, we show that DENV infection manipulated cholesterol metabolism in cells residing in low-oxygen microenvironments (hypoxia) such as in the liver, spleen, and lymph nodes. DENV infection induced expression of proprotein convertase subtilisin/kexin type 9 (PCSK9), which reduces low-density lipoprotein receptor (LDLR) recycling and hence cholesterol uptake. We found that, whereas LDLR uptake would have distributed cholesterol throughout the various cell compartments, de novo cholesterol synthesis enriched this lipid in the endoplasmic reticulum (ER). With cholesterol enrichment in the ER, ER-resident STING and type I IFN (IFN) activation was repressed during DENV infection. Our in vitro findings were further supported by the detection of elevated plasma PCSK9 levels in patients with dengue with high viremia and increased severity of plasma leakage. Our findings therefore suggest that PCSK9 plays a hitherto unrecognized role in dengue pathogenesis and that PCSK9 inhibitors could be a suitable host-directed treatment for patients with dengue.
Authors
Esther Shuyi Gan, Hwee Cheng Tan, Duyen Huynh Thi Le, Trieu Trung Huynh, Bridget Wills, Nabil G. Seidah, Eng Eong Ooi, Sophie Yacoub
×Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent for coronavirus 2019 (COVID-19) pneumonia. Little is known about the kinetics, tissue distribution, cross-reactivity, and neutralization antibody response in patients with COVID-19. Two groups of patients with RT-PCR–confirmed COVID-19 were enrolled in this study: 12 severely ill patients in intensive care units who needed mechanical ventilation and 11 mildly ill patients in isolation wards. Serial clinical samples were collected for laboratory detection. Results showed that most of the severely ill patients had viral shedding in a variety of tissues for 20–40 days after onset of disease (8/12, 66.7%), while the majority of mildly ill patients had viral shedding restricted to the respiratory tract and had no detectable virus RNA 10 days after onset (9/11, 81.8%). Mildly ill patients showed significantly lower IgM response compared with that of the severe group. IgG responses were detected in most patients in both the severe and mild groups at 9 days after onset, and remained at a high level throughout the study. Antibodies cross-reactive to SARS-CoV and SARS-CoV-2 were detected in patients with COVID-19 but not in patients with MERS. High levels of neutralizing antibodies were induced after about 10 days after onset in both severely and mildly ill patients which were higher in the severe group. SARS-CoV-2 pseudotype neutralization test and focus reduction neutralization test with authentic virus showed consistent results. Sera from patients with COVID-19 inhibited SARS-CoV-2 entry. Sera from convalescent patients with SARS or Middle East respiratory syndrome (MERS) did not. Anti–SARS-CoV-2 S and N IgG levels exhibited a moderate correlation with neutralization titers in patients’ plasma. This study improves our understanding of immune response in humans after SARS-CoV-2 infection.
Authors
Yanqun Wang, Lu Zhang, Ling Sang, Feng Ye, Shicong Ruan, Bei Zhong, Tie Song, Abeer N. Alshukairi, Rongchang Chen, Zhaoyong Zhang, Mian Gan, Airu Zhu, Yongbo Huang, Ling Luo, Chris Ka Pun Mok, Manal M. Al Gethamy, Haitao Tan, Zhengtu Li, Xiaofang Huang, Fang Li, Jing Sun, Yanjun Zhang, Liyan Wen, Yuming Li, Zhao Chen, Zhen Zhuang, Jianfen Zhuo, Chunke Chen, Lijun Kuang, Junxiang Wang, Huibin Lv, Yongliang Jiang, Min Li, Yimin Lin, Ying Deng, Lan Tang, Jieling Liang, Jicheng Huang, Stanley Perlman, Nanshan Zhong, Jingxian Zhao, J.S. Malik Peiris, Yimin Li, Jincun Zhao
×Abstract
The congenital sideroblastic anemias (CSAs) can be caused by primary defects in mitochondrial iron-sulfur (Fe-S) cluster biogenesis. HSCB (heat shock cognate B), which encodes a mitochondrial cochaperone, also known as HSC20 (heat shock cognate protein 20), is the partner of mitochondrial heat shock protein A9 (HSPA9). Together with glutaredoxin 5 (GLRX5), HSCB and HSPA9 facilitate the transfer of nascent 2-iron, 2-sulfur clusters to recipient mitochondrial proteins. Mutations in both HSPA9 and GLRX5 have previously been associated with CSA. Therefore, we hypothesized that mutations in HSCB could also cause CSA. We screened patients with genetically undefined CSA and identified a frameshift mutation and a rare promoter variant in HSCB in a female patient with non-syndromic CSA. We found that HSCB expression was decreased in patient-derived fibroblasts and K562 erythroleukemia cells engineered to have the patient-specific promoter variant. Furthermore, gene knockdown and deletion experiments performed in K562 cells, zebrafish, and mice demonstrate that loss of HSCB results in impaired Fe-S cluster biogenesis, a defect in RBC hemoglobinization, and the development of siderocytes and more broadly perturbs hematopoiesis in vivo. These results further affirm the involvement of Fe-S cluster biogenesis in erythropoiesis and hematopoiesis and define HSCB as a CSA gene.
Authors
Andrew Crispin, Chaoshe Guo, Caiyong Chen, Dean R. Campagna, Paul J. Schmidt, Daniel Lichtenstein, Chang Cao, Anoop K. Sendamarai, Gordon J. Hildick-Smith, Nicholas C. Huston, Jeanne Boudreaux, Sylvia S. Bottomley, Matthew M. Heeney, Barry H. Paw, Mark D. Fleming, Sarah Ducamp
×Abstract
Immunotherapeutic strategies are increasingly important in neuro-oncology, and the elucidation of escape mechanisms that lead to treatment resistance is crucial. We investigated the impact of immune pressure on the clonal dynamics and immune escape signature by comparing glioma growth in immunocompetent versus immunodeficient mice. Glioma-bearing WT and Pd-1–/– mice survived significantly longer than immunodeficient Pfp–/– Rag2–/– mice. While tumors in Pfp–/– Rag2–/– mice were highly polyclonal, immunoedited tumors in WT and Pd-1–/– mice displayed reduced clonality with emergence of immune escape clones. Tumor cells in WT mice were distinguished by an IFN-γ–mediated response signature with upregulation of genes involved in immunosuppression. Tumor-infiltrating stromal cells, which include macrophages/microglia, contributed even more strongly to the immunosuppressive signature than the actual tumor cells. The identified murine immune escape signature was reflected in human patients and correlated with poor survival. In conclusion, immune pressure profoundly shapes the clonal composition and gene regulation in malignant gliomas.
Authors
Cecile L. Maire, Malte Mohme, Michael Bockmayr, Krystian D. Fita, Kristoffer Riecken, Daniela Börnigen, Malik Alawi, Antonio Failla, Katharina Kolbe, Svenja Zapf, Mareike Holz, Katrin Neumann, Lasse Dührsen, Tobias Lange, Boris Fehse, Manfred Westphal, Katrin Lamszus
×Abstract
Human natural killer cell deficiency (NKD) arises from inborn errors of immunity that lead to impaired NK cell development, function, or both. Through the understanding of the biological perturbations in individuals with NKD, requirements for the generation of terminally mature functional innate effector cells can be elucidated. Here, we report a cause of NKD resulting from compound heterozygous mutations in minichromosomal maintenance complex member 10 (MCM10) that impaired NK cell maturation in a child with fatal susceptibility to CMV. MCM10 has not been previously associated with monogenic disease and plays a critical role in the activation and function of the eukaryotic DNA replisome. Through evaluation of patient primary fibroblasts, modeling patient mutations in fibroblast cell lines, and MCM10 knockdown in human NK cell lines, we have shown that loss of MCM10 function leads to impaired cell cycle progression and induction of DNA damage–response pathways. By modeling MCM10 deficiency in primary NK cell precursors, including patient-derived induced pluripotent stem cells, we further demonstrated that MCM10 is required for NK cell terminal maturation and acquisition of immunological system function. Together, these data define MCM10 as an NKD gene and provide biological insight into the requirement for the DNA replisome in human NK cell maturation and function.
Authors
Emily M. Mace, Silke Paust, Matilde I. Conte, Ryan M. Baxley, Megan M. Schmit, Sagar L. Patil, Nicole C. Guilz, Malini Mukherjee, Ashley E. Pezzi, Jolanta Chmielowiec, Swetha Tatineni, Ivan K. Chinn, Zeynep Coban Akdemir, Shalini N. Jhangiani, Donna M. Muzny, Asbjørg Stray-Pedersen, Rachel E. Bradley, Mo Moody, Philip P. Connor, Adrian G. Heaps, Colin Steward, Pinaki P. Banerjee, Richard A. Gibbs, Malgorzata Borowiak, James R. Lupski, Stephen Jolles, Anja K. Bielinsky, Jordan S. Orange
×Abstract
In the mammalian heart, the left ventricle (LV) rapidly becomes more dominant in size and function over the right ventricle (RV) after birth. The molecular regulators responsible for this chamber-specific differential growth are largely unknown. We found that cardiomyocytes in the neonatal mouse RV had lower proliferation, more apoptosis, and a smaller average size compared with the LV. This chamber-specific growth pattern was associated with a selective activation of p38 mitogen-activated protein kinase (MAPK) activity in the RV and simultaneous inactivation in the LV. Cardiomyocyte-specific deletion of both the Mapk14 and Mapk11 genes in mice resulted in loss of p38 MAPK expression and activity in the neonatal heart. Inactivation of p38 activity led to a marked increase in cardiomyocyte proliferation and hypertrophy but diminished cardiomyocyte apoptosis, specifically in the RV. Consequently, the p38-inactivated hearts showed RV-specific enlargement postnatally, progressing to pulmonary hypertension and right heart failure at the adult stage. Chamber-specific p38 activity was associated with differential expression of dual-specific phosphatases (DUSPs) in neonatal hearts, including DUSP26. Unbiased transcriptome analysis revealed that IRE1α/XBP1–mediated gene regulation contributed to p38 MAPK–dependent regulation of neonatal cardiomyocyte proliferation and binucleation. These findings establish an obligatory role of DUSP/p38/IRE1α signaling in cardiomyocytes for chamber-specific growth in the postnatal heart.
Authors
Tomohiro Yokota, Jin Li, Jijun Huang, Zhaojun Xiong, Qing Zhang, Tracey Chan, Yichen Ding, Christoph Rau, Kevin Sung, Shuxun Ren, Rajan Kulkarni, Tzung Hsiai, Xinshu Xiao, Marlin Touma, Susumu Minamisawa, Yibin Wang
×Abstract
Tissue factor (TF) is the primary initiator of blood coagulation in vivo and the only blood coagulation factor for which a human genetic defect has not been described. As there are no routine clinical assays that capture the contribution of endogenous TF to coagulation initiation, the extent to which reduced TF activity contributes to unexplained bleeding is unknown. Using whole genome sequencing, we identified a heterozygous frameshift variant (p.Ser117HisfsTer10) in F3, the gene encoding TF, causing premature termination of TF (TFshort) in a woman with unexplained bleeding. Routine hematological laboratory evaluation of the proposita was normal. CRISPR-edited human induced pluripotent stem cells recapitulating the variant were differentiated into vascular smooth muscle and endothelial cells that demonstrated haploinsufficiency of TF. The variant F3 transcript is eliminated by nonsense-mediated decay. Neither overexpression nor addition of exogenous recombinant TFshort inhibited factor Xa or thrombin generation, excluding a dominant-negative mechanism. F3+/– mice provide an animal model of TF haploinsufficiency and exhibited prolonged bleeding times, impaired thrombus formation, and reduced survival following major injury. Heterozygous TF deficiency is present in at least 1 in 25,000 individuals and could limit coagulation initiation in undiagnosed individuals with abnormal bleeding but a normal routine laboratory evaluation.
Authors
Sol Schulman, Emale El-Darzi, Mary H.C. Florido, Max Friesen, Glenn Merrill-Skoloff, Marisa A. Brake, Calvin R. Schuster, Lin Lin, Randal J. Westrick, Chad A. Cowan, Robert Flaumenhaft, NIHR BioResource, Willem H. Ouwehand, Kathelijne Peerlinck, Kathleen Freson, Ernest Turro, Bruce Furie
×Abstract
Pediatric and adult high-grade gliomas (HGGs) frequently harbor PDGFRA alterations. We hypothesized that cotreatment with everolimus may improve the efficacy of dasatinib in PDGFRα-driven glioma through combinatorial synergism and increased tumor accumulation of dasatinib. We performed dose-response, synergism, P-glycoprotein inhibition, and pharmacokinetic studies in in vitro and in vivo human and mouse models of HGG. Six patients with recurrent PDGFRα-driven glioma were treated with dasatinib and everolimus. We found that dasatinib effectively inhibited the proliferation of mouse and human primary HGG cells with a variety of PDGFRA alterations. Dasatinib exhibited synergy with everolimus in the treatment of HGG cells at low nanomolar concentrations of both agents, with a reduction in mTOR signaling that persisted after dasatinib treatment alone. Prolonged exposure to everolimus significantly improved the CNS retention of dasatinib and extended the survival of PPK tumor–bearing mice (mutant TP53, mutant PDGFRA, H3K27M). Six pediatric patients with glioma tolerated this combination without significant adverse events, and 4 patients with recurrent disease (n = 4) had a median overall survival of 8.5 months. Our results show that the efficacy of dasatinib treatment of PDGFRα-driven HGG was enhanced with everolimus and suggest a promising route for improving targeted therapy for this patient population.
Authors
Zachary Miklja, Viveka Nand Yadav, Rodrigo T. Cartaxo, Ruby Siada, Chase C. Thomas, Jessica R. Cummings, Brendan Mullan, Stefanie Stallard, Alyssa Paul, Amy K. Bruzek, Kyle Wierzbicki, Tao Yang, Taylor Garcia, Ian Wolfe, Marcia Leonard, Patricia L. Robertson, Hugh J.L. Garton, Daniel R. Wahl, Hemant Parmar, Jann N. Sarkaria, Cassie Kline, Sabine Mueller, Theodore Nicolaides, Chana Glasser, Sarah E.S. Leary, Sriram Venneti, Chandan Kumar-Sinha, Arul M. Chinnaiyan, Rajen Mody, Manjunath P. Pai, Timothy N. Phoenix, Bernard L. Marini, Carl Koschmann
×Abstract
BACKGROUND Idiopathic CD4 lymphopenia (ICL) is defined by persistently low CD4+ cell counts (<300 cells/μL) in the absence of a causal infection or immune deficiency and can manifest with opportunistic infections. Approximately 30% of ICL patients develop autoimmune disease. The prevalence and breadth of their autoantibodies, however, and their potential contribution to pathogenesis of ICL remain unclear.METHODS We hybridized 34 and 51 ICL patients’ sera to a 9,000-human-proteome array and to a 128-known-autoantigen array, respectively. Using a flow-based method, we characterized the presence of anti-lymphocyte Abs in the whole cohort of 72 patients, as well as the Ab functional capability of inducing Ab-dependent cell-mediated cytotoxicity (ADCC), complement deposition, and complement-dependent cytotoxicity (CDC). We tested ex vivo the activation of the classical complement pathway on ICL CD4+ T cells.RESULTS All ICL patients had a multitude of autoantibodies mostly directed against private (not shared) targets and unrelated quantitatively or qualitatively to the patients’ autoimmune disease status. The targets included lymphocyte intracellular and membrane antigens, confirmed by the detection by flow of IgM and IgG (mostly IgG1 and IgG4) anti–CD4+ cell Abs in 50% of the patients, with half of these cases triggering lysis of CD4+ T cells. We also detected in vivo classical complement activation on CD4+ T cells in 14% of the whole cohort.CONCLUSION Our data demonstrate that a high prevalence of autoantibodies in ICL, some of which are specific for CD4+ T cells, may contribute to pathogenesis, and may represent a potentially novel therapeutic target.TRIAL REGISTRATION ClinicalTrials.gov NCT00867269.FUNDING NIAID and National Institute of Arthritis and Musculoskeletal and Skin Diseases of the NIH.
Authors
Ainhoa Perez-Diez, Chun-Shu Wong, Xiangdong Liu, Harry Mystakelis, Jian Song, Yong Lu, Virginia Sheikh, Jeffrey S. Bourgeois, Andrea Lisco, Elizabeth Laidlaw, Cornelia Cudrici, Chengsong Zhu, Quan-Zhen Li, Alexandra F. Freeman, Peter R. Williamson, Megan Anderson, Gregg Roby, John S. Tsang, Richard Siegel, Irini Sereti
×Abstract
Therapy-induced neuroendocrine prostate cancer (t-NEPC) is a highly aggressive subtype of prostate cancer with poor patient survival. Emerging evidence indicates that t-NEPC can develop when prostate adenocarcinoma cells acquire cancer stem-like cell signaling in the presence of androgen receptor inhibition, followed by redifferentiation toward neuroendocrine lineage and subsequent t-NEPC progression. Whether the stem-like signaling is controlled by the core pluripotency stem cell genes (e.g., LIN28 and SOX2) remains unknown. Here, we report that the transcription of the LIN28B isoform and SOX2 were co-upregulated in t-NEPC patient tumors, patient-derived xenografts, transgenic mice, and cell models. Immunohistochemistry validated that LIN28B and SOX2 protein expression were elevated in t-NEPC patient biopsies. Using prostate adenocarcinoma and t-NEPC cell models, we demonstrated that LIN28B induced a stem-like gene network, neuroendocrine biomarkers, and neuroendocrine cell morphology. LIN28B depletion by CRISPR inhibited t-NEPC tumorigenesis and xenograft growth. These LIN28B functions were mediated mainly through the suppression of let-7 miRNA expression, resulting in de-repression of the transcription factor HMGA2 and HMGA2-mediated SOX2 expression. This study revealed a mechanism by which t-NEPC can develop through the LIN28B/let-7/SOX2 axis that regulates a cancer cell stem-like gene network, highlighting LIN28B as a potential therapeutic target in t-NEPC.
Authors
Jessica Lovnicki, Yu Gan, Tingting Feng, Yinan Li, Ning Xie, Chia-Hao Ho, Ahn R. Lee, Xufeng Chen, Lucia Nappi, Bo Han, Ladan Fazli, Jiaoti Huang, Martin E. Gleave, Xuesen Dong
×Abstract
Beclin 2 plays a critical role in metabolic regulation and obesity, but its functions in innate immune signaling and cancer development remain largely unknown. Here, we identified Beclin 2 as a critical negative regulator of inflammation and lymphoma development. Mice with homozygous ablation of BCL2-interacting protein 2 (Becn2) developed splenomegaly and lymphadenopathy and markedly increased ERK1/2 and NF-κB signaling for proinflammatory cytokine production. Beclin 2 targeted the key signaling kinases MEKK3 and TAK1 for degradation through an ATG9A-dependent, but ATG16L/Beclin 1/LC3–independent, autophagic pathway. Mechanistically, Beclin 2 recruited MEKK3 or TAK1 through ATG9A to form a complex (Beclin 2-ATG9A-MEKK3) on ATG9A+ vesicles upon ULK1 activation. Beclin 2 further interacted with STX5 and STX6 to promote the fusion of MEKK3- or TAK1-associated ATG9A+ vesicles to phagophores for subsequent degradation. Importantly, Becn2-deficient mice had a markedly increased incidence of lymphoma development, with persistent STAT3 activation. Myeloid-specific ablation of MEKK3 (Map3k3) completely rescued the phenotypes (splenomegaly, higher amounts of proinflammatory cytokines, and cancer incidence) of Becn2-deficient mice. Hence, our findings have identified an important role of Beclin 2 in the negative regulation of innate immune signaling and tumor development through an ATG9A-dependent, but ATG16L/Beclin 1/LC3–independent, autophagic pathway, thus providing a potential target for the treatment of inflammatory diseases and cancer.
Authors
Motao Zhu, Guangtong Deng, Peng Tan, Changsheng Xing, Cuiping Guan, Chongming Jiang, Yinlong Zhang, Bo Ning, Chaoran Li, Bingnan Yin, Kaifu Chen, Yuliang Zhao, Helen Y. Wang, Beth Levine, Guangjun Nie, Rong-Fu Wang
×Abstract
Chronic inflammation is deeply involved in various human disorders, such as cancer, neurodegenerative disorders, and metabolic disorders. Induction of epigenetic alterations, especially aberrant DNA methylation, is one of the major mechanisms, but how it is induced is still unclear. Here, we found that expression of TET genes, methylation erasers, was downregulated in inflamed mouse and human tissues, and that this was caused by upregulation of TET-targeting miRNAs such as MIR20A, MIR26B, and MIR29C, likely due to activation of NF-κB signaling downstream of IL-1β and TNF-α. However, TET knockdown induced only mild aberrant methylation. Nitric oxide (NO), produced by NOS2, enhanced enzymatic activity of DNA methyltransferases (DNMTs), methylation writers, and NO exposure induced minimal aberrant methylation. In contrast, a combination of TET knockdown and NO exposure synergistically induced aberrant methylation, involving genomic regions not methylated by either alone. The results showed that a vicious combination of TET repression, due to NF-κB activation, and DNMT activation, due to NO production, is responsible for aberrant methylation induction in human tissues.
Authors
Hideyuki Takeshima, Tohru Niwa, Satoshi Yamashita, Takeji Takamura-Enya, Naoko Iida, Mika Wakabayashi, Sohachi Nanjo, Masanobu Abe, Toshiro Sugiyama, Young-Joon Kim, Toshikazu Ushijima
×Abstract
Immune checkpoint blockade (ICB) has revolutionized cancer therapeutics. Desmoplastic malignancies, such as cholangiocarcinoma (CCA), have an abundant tumor immune microenvironment (TIME). However, to date, ICB monotherapy in such malignancies has been ineffective. Herein, we identify tumor-associated macrophages (TAMs) as the primary source of programmed death–ligand 1 (PD-L1) in human and murine CCA. In a murine model of CCA, recruited PD-L1+ TAMs facilitated CCA progression. However, TAM blockade failed to decrease tumor progression due to a compensatory emergence of granulocytic myeloid-derived suppressor cells (G-MDSCs) that mediated immune escape by impairing T cell response. Single-cell RNA sequencing (scRNA-Seq) of murine tumor G-MDSCs highlighted a unique ApoE G-MDSC subset enriched with TAM blockade; further analysis of a human scRNA-Seq data set demonstrated the presence of a similar G-MDSC subset in human CCA. Finally, dual inhibition of TAMs and G-MDSCs potentiated ICB. In summary, our findings highlight the therapeutic potential of coupling ICB with immunotherapies targeting immunosuppressive myeloid cells in CCA.
Authors
Emilien Loeuillard, Jingchun Yang, EeeLN Buckarma, Juan Wang, Yuanhang Liu, Caitlin Conboy, Kevin D. Pavelko, Ying Li, Daniel O’Brien, Chen Wang, Rondell P. Graham, Rory L. Smoot, Haidong Dong, Sumera Ilyas
×Abstract
Alarmins, sequestered self-molecules containing damage-associated molecular patterns, are released during tissue injury to drive innate immune cell proinflammatory responses. Whether endogenous negative regulators controlling early immune responses are also released at the site of injury is poorly understood. Herein, we establish that the stromal cell–derived alarmin interleukin 33 (IL-33) is a local factor that directly restricts the proinflammatory capacity of graft-infiltrating macrophages early after transplantation. By assessing heart transplant recipient samples and using a mouse heart transplant model, we establish that IL-33 is upregulated in allografts to limit chronic rejection. Mouse cardiac transplants lacking IL-33 displayed dramatically accelerated vascular occlusion and subsequent fibrosis, which was not due to altered systemic immune responses. Instead, a lack of graft IL-33 caused local augmentation of proinflammatory iNOS+ macrophages that accelerated graft loss. IL-33 facilitated a metabolic program in macrophages associated with reparative and regulatory functions, and local delivery of IL-33 prevented the chronic rejection of IL-33–deficient cardiac transplants. Therefore, IL-33 represents what we believe is a novel regulatory alarmin in transplantation that limits chronic rejection by restraining the local activation of proinflammatory macrophages. The local delivery of IL-33 in extracellular matrix–based materials may be a promising biologic for chronic rejection prophylaxis.
Authors
Tengfang Li, Zhongqiang Zhang, Joe G. Bartolacci, Gaelen K. Dwyer, Quan Liu, Lisa R. Mathews, Murugesan Velayutham, Anna S. Roessing, Yoojin C. Lee, Helong Dai, Sruti Shiva, Martin H. Oberbarnscheidt, Jenna L. Dziki, Steven J. Mullet, Stacy G. Wendell, James D. Wilkinson, Steven A. Webber, Michelle Wood-Trageser, Simon C. Watkins, Anthony J. Demetris, George S. Hussey, Stephen F. Badylak, Hēth R. Turnquist
×Abstract
The development of broadly neutralizing antibodies (BNAbs) in HIV infection is a result of long-term coevolutionary interaction between viruses and antibodies. Understanding how this interaction promotes the increase of neutralization breadth during infection will improve the way in which AIDS vaccine strategies are designed. In this paper, we used SIV-infected rhesus macaques as a model to study the development of neutralization breadth by infecting rhesus macaques with longitudinal NAb escape variants and evaluating the kinetics of NAb response and viral evolution. We found that the infected macaques developed a stepwise NAb response against escape variants and increased neutralization breadth during the course of infection. Furthermore, the increase of neutralization breadth correlated with the duration of infection but was independent of properties of the inoculum, viral loads, or viral diversity during infection. These results imply that the duration of infection was the main factor driving the development of BNAbs. These data suggest the importance of novel immunization strategies to induce effective NAb response against HIV infection by mimicking long-term infection.
Authors
Fan Wu, Ilnour Ourmanov, Andrea Kirmaier, Sivan Leviyang, Celia LaBranche, Jinghe Huang, Sonya Whitted, Kenta Matsuda, David Montefiori, Vanessa M. Hirsch
×Abstract
Late-onset inflammatory toxicities resembling hemophagocytic lymphohistiocytosis (HLH) or macrophage activation syndrome (MAS) occur after chimeric antigen receptor T cell (CAR T cell) infusion and represent a therapeutic challenge. Given the established link between perforin deficiency and primary HLH, we investigated the role of perforin in anti-CD19 CAR T cell efficacy and HLH-like toxicities in a syngeneic murine model. Perforin contributed to both CD8+ and CD4+ CAR T cell cytotoxicity but was not required for in vitro or in vivo leukemia clearance. Upon CAR-mediated in vitro activation, perforin-deficient CAR T cells produced higher amounts of proinflammatory cytokines compared with WT CAR T cells. Following in vivo clearance of leukemia, perforin-deficient CAR T cells reexpanded, resulting in splenomegaly with disruption of normal splenic architecture and the presence of hemophagocytes, which are findings reminiscent of HLH. Notably, a substantial fraction of patients who received anti-CD22 CAR T cells also experienced biphasic inflammation, with the second phase occurring after the resolution of cytokine release syndrome, resembling clinical manifestations of HLH. Elevated inflammatory cytokines such as IL-1β and IL-18 and concurrent late CAR T cell expansion characterized the HLH-like syndromes occurring in the murine model and in humans. Thus, a murine model of perforin-deficient CAR T cells recapitulated late-onset inflammatory toxicities occurring in human CAR T cell recipients, providing therapeutically relevant mechanistic insights.
Authors
Kazusa Ishii, Marie Pouzolles, Christopher D. Chien, Rebecca A. Erwin-Cohen, M. Eric Kohler, Haiying Qin, Haiyan Lei, Skyler Kuhn, Amanda K. Ombrello, Alina Dulau-Florea, Michael A. Eckhaus, Haneen Shalabi, Bonnie Yates, Daniel A. Lichtenstein, Valérie S. Zimmermann, Taisuke Kondo, Jack F. Shern, Howard A. Young, Naomi Taylor, Nirali N. Shah, Terry J. Fry
×Abstract
Cells sense the extracellular environment and mechanical stimuli and translate these signals into intracellular responses through mechanotransduction, which alters cell maintenance, proliferation, and differentiation. Here we use a mouse model of trauma-induced heterotopic ossification (HO) to examine how cell-extrinsic forces impact mesenchymal progenitor cell (MPC) fate. After injury, single-cell (sc) RNA sequencing of the injury site reveals an early increase in MPC genes associated with pathways of cell adhesion and ECM-receptor interactions, and MPC trajectories to cartilage and bone. Immunostaining uncovers active mechanotransduction after injury with increased focal adhesion kinase signaling and nuclear translocation of transcriptional coactivator TAZ, inhibition of which mitigates HO. Similarly, joint immobilization decreases mechanotransductive signaling, and completely inhibits HO. Joint immobilization decreases collagen alignment and increases adipogenesis. Further, scRNA sequencing of the HO site after injury with or without immobilization identifies gene signatures in mobile MPCs correlating with osteogenesis, and signatures from immobile MPCs with adipogenesis. scATAC-seq in these same MPCs confirm that in mobile MPCs, chromatin regions around osteogenic genes are open, whereas in immobile MPCs, regions around adipogenic genes are open. Together these data suggest that joint immobilization after injury results in decreased ECM alignment, altered MPC mechanotransduction, and changes in genomic architecture favoring adipogenesis over osteogenesis, resulting in decreased formation of HO.
Authors
Amanda K. Huber, Nicole Patel, Chase A. Pagani, Simone Marini, Karthik R. Padmanabhan, Daniel L. Matera, Mohamed Said, Charles Hwang, Ginny Ching-Yun Hsu, Andrea A. Poli, Amy L. Strong, Noelle D. Visser, Joseph A. Greenstein, Reagan Nelson, Shuli Li, Michael T. Longaker, Yi Tang, Stephen J. Weiss, Brendon M. Baker, Aaron W. James, Benjamin Levi
×Abstract
Tumor immunosuppression is a limiting factor for successful cancer therapy. The lipid sphingosine-1-phosphate (S1P), which signals through 5 distinct G protein–coupled receptors (S1PR1–5), has emerged as an important regulator of carcinogenesis. However, the utility of targeting S1P in tumors is hindered by S1P’s impact on immune cell trafficking. Here, we report that ablation of the immune cell–specific receptor S1PR4, which plays a minor role in immune cell trafficking, delayed tumor development and improved therapy success in murine models of mammary and colitis-associated colorectal cancer through increased CD8+ T cell abundance. Transcriptome analysis revealed that S1PR4 affected proliferation and survival of CD8+ T cells in a cell-intrinsic manner via the expression of Pik3ap1 and Lta4h. Accordingly, PIK3AP1 expression was connected to increased CD8+ T cell proliferation and clinical parameters in human breast and colon cancer. Our data indicate a so-far-unappreciated tumor-promoting role of S1P by restricting CD8+ T cell expansion via S1PR4.
Authors
Catherine Olesch, Evelyn Sirait-Fischer, Matthias Berkefeld, Annika F. Fink, Rosa M. Susen, Birgit Ritter, Birgitta E. Michels, Dieter Steinhilber, Florian R. Greten, Rajkumar Savai, Kazuhiko Takeda, Bernhard Brüne, Andreas Weigert
×Abstract
Transcription infidelity (TI) is a mechanism that increases RNA and protein diversity. We found that single-base omissions (i.e., gaps) occurred at significantly higher rates in the RNA of highly allergenic legumes. Transcripts from peanut, soybean, sesame, and mite allergens contained a higher density of gaps than those of nonallergens. Allergen transcripts translate into proteins with a cationic carboxy terminus depleted in hydrophobic residues. In mice, recombinant TI variants of the peanut allergen Ara h 2, but not the canonical allergen itself, induced, without adjuvant, the production of anaphylactogenic specific IgE (sIgE), binding to linear epitopes on both canonical and TI segments of the TI variants. The removal of cationic proteins from bovine lactoserum markedly reduced its capacity to induce sIgE. In peanut-allergic children, the sIgE reactivity was directed toward both canonical and TI segments of Ara h 2 variants. We discovered 2 peanut allergens, which we believe to be previously unreported, because of their RNA-DNA divergence gap patterns and TI peptide amino acid composition. Finally, we showed that the sIgE of children with IgE-negative milk allergy targeted cationic proteins in lactoserum. We propose that it is not the canonical allergens, but their TI variants, that initiate sIgE isotype switching, while both canonical and TI variants elicit clinical allergic reactions.
Authors
Benoit Thouvenot, Olivier Roitel, Julie Tomasina, Benoit Hilselberger, Christelle Richard, Sandrine Jacquenet, Françoise Codreanu-Morel, Martine Morisset, Gisèle Kanny, Etienne Beaudouin, Christine Delebarre-Sauvage, Thierry Olivry, Claude Favrot, Bernard E. Bihain
×Abstract
Senescent cells (SnCs) are implicated in the pathogenesis of age-related diseases including osteoarthritis (OA), in part via expression of a senescence-associated secretory phenotype (SASP) that includes immunologically relevant factors and cytokines. In a model of posttraumatic OA (PTOA), anterior cruciate ligament transection (ACLT) induced a type 17 immune response in the articular compartment and draining inguinal lymph nodes (LNs) that paralleled expression of the senescence marker p16INK4a (Cdkn2a) and p21 (Cdkn1a). Innate lymphoid cells, γδ+ T cells, and CD4+ T cells contributed to IL-17 expression. Intra-articular injection of IL-17–neutralizing antibody reduced joint degeneration and decreased expression of the senescence marker Cdkn1a. Local and systemic senolysis was required to attenuate tissue damage in aged animals and was associated with decreased IL-17 and increased IL-4 expression in the articular joint and draining LNs. In vitro, we found that Th17 cells induced senescence in fibroblasts and that SnCs skewed naive T cells toward Th17 or Th1, depending on the presence of TGF-β. The SASP profile of the inflammation-induced SnCs included altered Wnt signaling, tissue remodeling, and cell-cycle pathways not previously implicated in senescence. These findings provide molecular targets and mechanisms for senescence induction and therapeutic strategies to support tissue healing in an aged environment.
Authors
Heather J. Faust, Hong Zhang, Jin Han, Matthew T. Wolf, Ok Hee Jeon, Kaitlyn Sadtler, Alexis N. Peña, Liam Chung, David R. Maestas Jr., Ada J. Tam, Drew M. Pardoll, Judith Campisi, Franck Housseau, Daohong Zhou, Clifton O. Bingham III, Jennifer H. Elisseeff
×Abstract
To improve the clinical outcome of adoptive NK cell therapy in patients with solid tumors, NK cells need to persist within the tumor microenvironment (TME) in which the abundance of ROS could dampen antitumor immune responses. In the present study, we demonstrated that IL-15–primed NK cells acquired resistance against oxidative stress through the thioredoxin system activated by mTOR. Mechanistically, the activation of thioredoxin showed dependence on localization of thioredoxin-interacting protein. We show that NK cells residing in the tumor core expressed higher thiol densities that could aid in protecting other lymphocytes against ROS within the TME. Furthermore, the prognostic value of IL15 and the NK cell gene signature in tumors may be influenced by tobacco smoking history in patients with non–small-cell lung cancer (NSCLC). Collectively, the levels of reducing antioxidants in NK cells may not only predict better tumor penetrance but potentially even the immune therapy response.
Authors
Ying Yang, Shi Yong Neo, Ziqing Chen, Weiyingqi Cui, Yi Chen, Min Guo, Yongfang Wang, Haiyan Xu, Annina Kurzay, Evren Alici, Lars Holmgren, Felix Haglund, Kai Wang, Andreas Lundqvist
×Abstract
Using the Nephrotic Syndrome Study Network Consortium data set and other publicly available transcriptomic data sets, we identified retinoic acid receptor responder protein 1 (RARRES1) as a gene whose expression positively correlated with renal function decline in human glomerular disease. The glomerular expression of RARRES1, which is largely restricted to podocytes, increased in focal segmental glomerulosclerosis (FSGS) and diabetic kidney disease (DKD). TNF-α was a potent inducer of RARRES1 expression in cultured podocytes, and transcriptomic analysis showed the enrichment of cell death pathway genes with RARRES1 overexpression. The overexpression of RARRES1 indeed induced podocyte apoptosis in vitro. Notably, this effect was dependent on its cleavage in the extracellular domain, as the mutation of its cleavage site abolished the apoptotic effect. Mechanistically, the soluble RARRES1 was endocytosed and interacted with and inhibited RIO kinase 1 (RIOK1), resulting in p53 activation and podocyte apoptosis. In mice, podocyte-specific overexpression of RARRES1 resulted in marked glomerular injury and albuminuria, while the overexpression of RARRES1 cleavage mutant had no effect. Conversely, podocyte-specific knockdown of Rarres1 in mice ameliorated glomerular injury in the setting of adriamycin-induced nephropathy. Our study demonstrates an important role and the mechanism of RARRES1 in podocyte injury in glomerular disease.
Authors
Anqun Chen, Ye Feng, Han Lai, Wenjun Ju, Zhengzhe Li, Yu Li, Andrew Wang, Quan Hong, Fang Zhong, Chengguo Wei, Jia Fu, Tianjun Guan, Bichen Liu, Matthias Kretzler, Kyung Lee, John Cijiang He
×Abstract
Postnatal failure of oligodendrocyte maturation has been proposed as a cellular mechanism of diffuse white matter injury (WMI) in premature infants. However, the molecular mechanisms for oligodendrocyte maturational failure remain unclear. In neonatal mice and cultured differentiating oligodendrocytes, sublethal intermittent hypoxic (IH) stress activated cyclophilin D–dependent mitochondrial proton leak and uncoupled mitochondrial respiration, leading to transient bioenergetic stress. This was associated with development of diffuse WMI: poor oligodendrocyte maturation, diffuse axonal hypomyelination, and permanent sensorimotor deficit. In normoxic mice and oligodendrocytes, exposure to a mitochondrial uncoupler recapitulated the phenotype of WMI, supporting the detrimental role of mitochondrial uncoupling in the pathogenesis of WMI. Compared with WT mice, cyclophilin D–knockout littermates did not develop bioenergetic stress in response to IH challenge and fully preserved oligodendrocyte maturation, axonal myelination, and neurofunction. Our study identified the cyclophilin D–dependent mitochondrial proton leak and uncoupling as a potentially novel subcellular mechanism for the maturational failure of oligodendrocytes and offers a potential therapeutic target for prevention of diffuse WMI in premature infants experiencing chronic IH stress.
Authors
Zoya Niatsetskaya, Sergey Sosunov, Anna Stepanova, James Goldman, Alexander Galkin, Maria Neginskaya, Evgeny Pavlov, Vadim Ten
×Haploinsufficiency of immune checkpoint receptor CTLA4 induces a distinct neuroinflammatory disorder
Abstract
BACKGROUND Cytotoxic T lymphocyte antigen 4 (CTLA4) is essential for immune homeostasis. Genetic mutations causing haploinsufficiency (CTLA4h) lead to a phenotypically heterogenous, immune-mediated disease that can include neuroinflammation. The neurological manifestations of CTLA4h are poorly characterized.METHODS We performed an observational natural history study of 50 patients with CTLA4h who were followed at the NIH. We analyzed clinical, radiological, immunological, and histopathological data.RESULTS Evidence for neuroinflammation was observed in 32% (n = 16 of 50) of patients in this cohort by magnetic resonance imaging (MRI) and/or by cerebrospinal fluid analysis. Clinical symptoms were commonly absent or mild in severity, with headaches as the leading complaint (n = 13 of 16). The most striking findings were relapsing, large, contrast-enhancing focal lesions in the brain and spinal cord observed on MRI. We detected inflammation in the cerebrospinal fluid and leptomeninges before the parenchyma. Brain biopsies of inflammatory lesions from 10 patients showed perivascular and intraparenchymal mixed cellular infiltrates with little accompanying demyelination or neuronal injury.CONCLUSIONS Neuroinflammation due to CTLA4h is mediated primarily by an infiltrative process with a distinct and striking dissociation between clinical symptoms and radiological findings in the majority of patients.FUNDING NIAID, NIH, Division of Intramural Research, NINDS, NIH, Division of Intramural Research, and the National Multiple Sclerosis Society–American Brain Foundation.TRIAL REGISTRATION ClinicalTrials.gov NCT00001355.
Authors
Matthew K. Schindler, Stefania Pittaluga, Yoshimi Enose-Akahata, Helen C. Su, V. Koneti Rao, Amy Rump, Steven Jacobson, Irene Cortese, Daniel S. Reich, Gulbu Uzel
×Abstract
Pathologic lymphatic remodeling in lymphedema evolves during periods of tissue inflammation and hypoxia through poorly defined processes. In human and mouse lymphedema, there is a significant increase of hypoxia inducible factor 1 α (HIF-1α), but a reduction of HIF-2α protein expression in lymphatic endothelial cells (LECs). We questioned whether dysregulated expression of these transcription factors contributes to disease pathogenesis and found that LEC-specific deletion of Hif2α exacerbated lymphedema pathology. Even without lymphatic vascular injury, the loss of LEC-specific Hif2α caused anatomic pathology and a functional decline in fetal and adult mice. These findings suggest that HIF-2α is an important mediator of lymphatic health. HIF-2α promoted protective phosphorylated TIE2 (p-TIE2) signaling in LECs, a process also replicated by upregulating TIE2 signaling through adenovirus-mediated angiopoietin-1 (Angpt1) gene therapy. Our study suggests that HIF-2α normally promotes healthy lymphatic homeostasis and raises the exciting possibility that restoring HIF-2α pathways in lymphedema could mitigate long-term pathology and disability.
Authors
Xinguo Jiang, Wen Tian, Eric J. Granucci, Allen B. Tu, Dongeon Kim, Petra Dahms, Shravani Pasupneti, Gongyong Peng, Yesl Kim, Amber H. Lim, F. Hernan Espinoza, Matthew Cribb, J. Brandon Dixon, Stanley G. Rockson, Gregg L. Semenza, Mark R. Nicolls
×Abstract
During hemolysis, macrophages in the liver phagocytose damaged erythrocytes to prevent the toxic effects of cell-free hemoglobin and heme. It remains unclear how this homeostatic process modulates phagocyte functions in inflammatory diseases. Using a genetic mouse model of spherocytosis and single-cell RNA sequencing, we found that erythrophagocytosis skewed liver macrophages into an antiinflammatory phenotype that we defined as MarcohiHmoxhiMHC class IIlo erythrophagocytes. This phenotype transformation profoundly mitigated disease expression in a model of an anti-CD40–induced hyperinflammatory syndrome with necrotic hepatitis and in a nonalcoholic steatohepatitis model, representing 2 macrophage-driven sterile inflammatory diseases. We reproduced the antiinflammatory erythrophagocyte transformation in vitro by heme exposure of mouse and human macrophages, yielding a distinctive transcriptional signature that segregated heme-polarized from M1- and M2-polarized cells. Mapping transposase-accessible chromatin in single cells by sequencing defined the transcription factor NFE2L2/NRF2 as a critical driver of erythrophagocytes, and Nfe2l2/Nrf2 deficiency restored heme-suppressed inflammation. Our findings point to a pathway that regulates macrophage functions to link erythrocyte homeostasis with innate immunity.
Authors
Marc Pfefferlé, Giada Ingoglia, Christian A. Schaer, Ayla Yalamanoglu, Raphael Buzzi, Irina L. Dubach, Ge Tan, Emilio Y. López-Cano, Nadja Schulthess, Kerstin Hansen, Rok Humar, Dominik J. Schaer, Florence Vallelian
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BACKGROUND Induction of innate immune memory, also termed trained immunity, by the antituberculosis vaccine bacillus Calmette-Guérin (BCG) contributes to protection against heterologous infections. However, the overall impact of BCG vaccination on the inflammatory status of an individual is not known; while induction of trained immunity may suggest increased inflammation, BCG vaccination has been epidemiologically associated with a reduced incidence of inflammatory and allergic diseases.METHODS We investigated the impact of BCG (BCG-Bulgaria, InterVax) vaccination on systemic inflammation in a cohort of 303 healthy volunteers, as well as the effect of the inflammatory status on the response to vaccination. A targeted proteome platform was used to measure circulating inflammatory proteins before and after BCG vaccination, while ex vivo Mycobacterium tuberculosis– and Staphylococcus aureus–induced cytokine responses in peripheral blood mononuclear cells were used to assess trained immunity.RESULTS While BCG vaccination enhanced cytokine responses to restimulation, it reduced systemic inflammation. This effect was validated in 3 smaller cohorts, and was much stronger in men than in women. In addition, baseline circulating inflammatory markers were associated with ex vivo cytokine responses (trained immunity) after BCG vaccination.CONCLUSION The capacity of BCG to enhance microbial responsiveness while dampening systemic inflammation should be further explored for potential therapeutic applications.FUNDING Netherlands Organization for Scientific Research, European Research Council, and the Danish National Research Foundation.
Authors
Valerie A.C.M. Koeken, L. Charlotte J. de Bree, Vera P. Mourits, Simone J.C.F.M. Moorlag, Jona Walk, Branko Cirovic, Rob J.W. Arts, Martin Jaeger, Helga Dijkstra, Heidi Lemmers, Leo A.B. Joosten, Christine S. Benn, Reinout van Crevel, Mihai G. Netea
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BACKGROUND The antituberculosis vaccine bacillus Calmette-Guérin (BCG) reduces overall infant mortality. Induction of innate immune memory, also termed trained immunity, contributes toward protection against heterologous infections. Since immune cells display oscillations in numbers and function throughout the day, we investigated the effect of BCG administration time on the induction of trained immunity.METHODS Eighteen volunteers were vaccinated with BCG at 6 pm and compared with 36 age- and sex-matched volunteers vaccinated between 8 am and 9 am. Peripheral blood mononuclear cells were stimulated with Staphylococcus aureus and Mycobacterium tuberculosis before, as well as 2 weeks and 3 months after, BCG vaccination. Cytokine production was measured to assess the induction of trained immunity and adaptive responses, respectively. Additionally, the influence of vaccination time on induction of trained immunity was studied in an independent cohort of 302 individuals vaccinated between 8 am and 12 pm with BCG.RESULTS Compared with evening vaccination, morning vaccination elicited both a stronger trained immunity and adaptive immune phenotype. In a large cohort of 302 volunteers, early morning vaccination resulted in a superior cytokine production capacity compared with later morning. A cellular, rather than soluble, substrate of the circadian effect of BCG vaccination was demonstrated by the enhanced capacity to induce trained immunity in vitro in morning- compared with evening-isolated monocytes.CONCLUSIONS BCG vaccination in the morning induces stronger trained immunity and adaptive responses compared with evening vaccination. Future studies should take vaccine administration time into account when studying specific and nonspecific effects of vaccines; early morning should be the preferred moment of BCG administration.FUNDING The Netherlands Organization for Scientific Research, the European Research Council, and the Danish National Research Foundation.
Authors
L. Charlotte J. de Bree, Vera P. Mourits, Valerie A.C.M. Koeken, Simone J.C.F.M. Moorlag, Robine Janssen, Lukas Folkman, Daniele Barreca, Thomas Krausgruber, Victoria Fife-Gernedl, Boris Novakovic, Rob J.W. Arts, Helga Dijkstra, Heidi Lemmers, Christoph Bock, Leo A.B. Joosten, Reinout van Crevel, Christine S. Benn, Mihai G. Netea
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ISSN: 0021-9738 (print), 1558-8238 (online)