Latest issue: April 18, 2019
In the issue
Abstract
Sarcomeric disarray is a hallmark of gene mutations in patients with hypertrophic cardiomyopathy (HCM). However, it is unknown when detrimental sarcomeric changes first occur and whether they originate in the developing embryonic heart. Furthermore, Rho kinase (ROCK) is a serine/threonine protein kinase that is critical for regulating the function of several sarcomeric proteins, and therefore, our aim was to determine whether disruption of ROCK signaling during the earliest stages of heart development would disrupt the integrity of sarcomeres, altering heart development and function. Using a mouse model in which the function of ROCK is specifically disrupted in embryonic cardiomyocytes, we demonstrate a progressive cardiomyopathy that first appeared as sarcomeric disarray during cardiogenesis. This led to abnormalities in the structure of the embryonic ventricular wall and compensatory cardiomyocyte hypertrophy during fetal development. This sarcomeric disruption and hypertrophy persisted throughout adult life, triggering left ventricular concentric hypertrophy with systolic dysfunction, and reactivation of fetal gene expression and cardiac fibrosis, all typical features of HCM. Taken together, our findings establish a mechanism for the developmental origin of the sarcomeric phenotype of HCM and suggest that variants in the ROCK genes or disruption of ROCK signaling could, in part, contribute to its pathogenesis.
Authors
Kate E. Bailey, Guy A. MacGowan, Simon Tual-Chalot, Lauren Phillips, Timothy J. Mohun, Deborah J. Henderson, Helen M. Arthur, Simon D. Bamforth, Helen M. Phillips
Abstract
AXL overexpression is a common resistance mechanism to anticancer therapies, including the resistance to BYL719 (Alpelisib) — the p110α isoform specific inhibitor of phosphoinositide 3-kinase (PI3K) — in esophagus squamous cell carcinoma (ESCC) and head and neck squamous cell carcinoma (HNSCC). However, the mechanisms underlying AXL overexpression in resistance to BYL719 remain elusive. Here, we demonstrate that the AP-1 transcription factors c-JUN and c-FOS regulate AXL overexpression in HNSCC and ESCC. The expression of AXL was correlated with that of c-JUN both in HNSCC patients and in HNSCC and ESCC cell lines. Silencing of c-JUN and c-FOS expression in tumor cells downregulated AXL expression and enhanced the sensitivity of human papilloma virus–positive (HPVPos) and –negative (HPVNeg) tumor cells to BYL719 in vitro. Blocking of JNK using SP600125 in combination with BYL719 showed a synergistic antiproliferative effect in vitro, which was accompanied by AXL downregulation and potent inhibition of the mTOR pathway. In vivo, the BYL719–SP600125 drug combination led to the arrest of tumor growth in cell line–derived and patient-derived xenograft models, as well as in syngeneic head and neck murine cancer models. Collectively, our data suggest that JNK inhibition, in combination with anti-PI3K therapy, is a new therapeutic strategy that should be tested in HPVPos and HPVNeg HNSCC and ESCC patients.
Authors
Mai Badarni, Manu Prasad, Noa Balaban, Jonathan Zorea, Ksenia M. Yegodayev, Joshua Ben-Zion, Anat Bahat Dinur, Reidar Grénman, Barak Rotblat, Limor Cohen, Moshe Elkabets
Abstract
Drug refractory epilepsy (RE) is a chronic neurological disease with varied etiology that represents a group of patients whose seizures do not respond to antiepileptic drugs. The immune system may have a role in seizure and epilepsy development, but the specific mechanisms of inflammation that lead to epileptogenesis and contribute to RE are unknown. Here, we used mass cytometry to comprehensively study the immune system of pediatric patients with RE and compared their immune profile and function with patients with age-matched autoimmune encephalitis (AIE) and healthy controls. Patients with RE and AIE displayed similar immune profiles overall, with changes in CD4+ and CD8+ T cell subsets and an unbalance toward proinflammatory IL-17 production. In addition, patients with RE uniquely showed an altered balance in NK cell subsets. A systems-level intercellular network analysis identified rewiring of the immune system, leading to loss of inhibitory/regulatory intercellular connections and emergence of proinflammatory pathogenic functions in neuroinflammatory immune cell networks in patients with AIE and RE. These data underscore the contribution of systemic inflammation to the pathogenesis of seizures and epileptogenesis and have direct translational implications in advancing diagnostics and therapeutics design.
Authors
Pavanish Kumar, Derrick Chan Wei Shih, Amanda Lim, Bhairav Paleja, Simon Ling, Lai Li Yun, Su Li Poh, Adeline Ngoh, Thaschawee Arkachaisri, Joo Guan Yeo, Salvatore Albani
Abstract
Changes in neuronal activity alter blood flow to match energy demand with the supply of oxygen and nutrients. This functional hyperemia is maintained by interactions among neurons, vascular cells, and glia. However, how changing neuronal activity prevalent at the onset of neurodegenerative disease affects neurovascular elements is unclear. Here, in mice with photoreceptor degeneration, a model of neuron-specific dysfunction, we combined the assessment of visual function, neurovascular unit structure, and blood-retina barrier permeability. We found that the rod loss paralleled remodeling of the neurovascular unit, comprising photoreceptors, retinal pigment epithelium, and Muller glia. When substantial visual function was still present, blood flow became disrupted and the blood-retina barrier began to fail, facilitating cone loss and vision decline. Thus, in contrast to the established view, the vascular deficit in neuronal degeneration is not a late consequence of neuronal dysfunction but is present early in the course of disease. These findings further establish the importance of vascular deficit and blood-retina barrier function in neuron-specific loss and highlight it as a target for early therapeutic intervention.
Authors
Elena Ivanova, Nazia M. Alam, Glen T. Prusky, Botir T. Sagdullaev
Abstract
MHC I–restricted epitopes of chicken ovalbumin (OVA) were originally identified using CD8+ T cells as probes. Here, using bioinformatics tools, we identify 4 additional epitopes in OVA in addition to a cryptic epitope. Each additional epitope is presented in vivo, as deduced from the lack of CD8+ T cell response to it in OVA-transgenic mice. In addition, CD8 responses to the previously known epitopes and those identified in this study are examined in C57BL/6J mice exposed to the OVA-expressing tumor E.G7 in several ways. No responses to any epitope, including SIINFEKL, are detected in mice with growing E.G7 or mice immunized with the tumor. Only in E.G7-bearing mice treated with an anti–CTLA-4 antibody, which depletes tumor-infiltrating regulatory T cells, are CD8 responses to SIINFEKL and the epitope EKYNLTSVL identified in this study detected. Finally, all epitopes fail to treat mice with preexisting tumors. These observations force an important reconsideration of the common assumptions about the therapeutic value of neoepitopes detected by CD8 responses in tumor-bearing hosts.
Authors
Sukrut Hemant Karandikar, John Sidney, Alessandro Sette, Mark Joseph Selby, Alan Jerry Korman, Pramod Kumar Srivastava
Abstract
Tregs are key modulators of inflammation and are important for the maintenance of peripheral tolerance. Adoptive immunotherapy with polyclonal Tregs holds promise in organ transplantation, graft-versus-host disease, and autoimmune diseases but may be enhanced by antigen-specific, long-lived Tregs. We modified primary human Tregs with chimeric antigen receptors (CARs) bearing different costimulatory domains and performed in vitro analyses of their phenotype and function. While neither the presence of a CAR nor the type of costimulation domain influenced Foxp3 expression in Tregs, the costimulation domain of the CARs affected CAR-Treg surface phenotype and functions, such as cytokine production. Furthermore, signaling from the CD28 costimulation domain maintained CAR-Treg suppressor function, whereas 4-1B costimulation did not. In vivo, CAR-Tregs accumulated at sites expressing target antigen and suppressed antigen-specific effector T cell responses; however, only CAR-Tregs with CD28 signaling domains were potent inhibitors of effector T cell–mediated graft rejection in vivo. Our findings support the use of CD28-based CAR-Tregs for tissue-specific immune suppression in the clinic.
Authors
Angela C. Boroughs, Rebecca C. Larson, Bryan D. Choi, Amanda A. Bouffard, Lauren S. Riley, Erik Schiferle, Anupriya S. Kulkarni, Curtis L. Cetrulo, David Ting, Bruce R. Blazar, Shadmehr Demehri, Marcela V. Maus
Abstract
The endoplasmic reticulum (ER) of cancer cells needs to adapt to the enhanced proteotoxic stress associated with the accumulation of unfolded, misfolded, and transformation-associated proteins. One way by which tumors thrive in the context of ER stress is by promoting ER-associated degradation (ERAD), although the mechanisms are poorly understood. Here, we show that the small p97/VCP-interacting protein (SVIP), an endogenous inhibitor of ERAD, undergoes DNA hypermethylation–associated silencing in tumorigenesis to achieve this goal. SVIP exhibits tumor suppressor features and its recovery is associated with increased ER stress and growth inhibition. Proteomic and metabolomic analyses show that cancer cells with epigenetic loss of SVIP are depleted in mitochondrial enzymes and oxidative respiration activity. This phenotype is reverted upon SVIP restoration. The dependence of SVIP-hypermethylated cancer cells on aerobic glycolysis and glucose was also associated with sensitivity to an inhibitor of the glucose transporter GLUT1. This could be relevant to the management of tumors carrying SVIP epigenetic loss, because these occur in high-risk patients who manifest poor clinical outcomes. Overall, our study provides insights into how epigenetics helps deal with ER stress and how SVIP epigenetic loss in cancer may be amenable to therapies that target glucose transporters.
Authors
Pere Llinàs-Arias, Margalida Rosselló-Tortella, Paula López-Serra, Montserrat Pérez-Salvia, Fernando Setién, Silvia Marin, Juan P. Muñoz, Alexandra Junza, Jordi Capellades, María E. Calleja-Cervantes, Humberto J. Ferreira, Manuel Castro de Moura, Marina Srbic, Anna Martínez-Cardús, Carolina de la Torre, Alberto Villanueva, Marta Cascante, Oscar Yanes, Antonio Zorzano, Catia Moutinho, Manel Esteller
Abstract
Many lung diseases result from a failure of efficient regeneration of damaged alveolar epithelial cells (AECs) after lung injury. During regeneration, AEC2s proliferate to replace lost cells, after which proliferation halts and some AEC2s transdifferentiate into AEC1s to restore normal alveolar structure and function. Although the mechanisms underlying AEC2 proliferation have been studied, the mechanisms responsible for halting proliferation and inducing transdifferentiation are poorly understood. To identify candidate signaling pathways responsible for halting proliferation and inducing transdifferentiation, we performed single-cell RNA sequencing on AEC2s during regeneration in a murine model of lung injury induced by intratracheal LPS. Unsupervised clustering revealed distinct subpopulations of regenerating AEC2s: proliferating, cell cycle arrest, and transdifferentiating. Gene expression analysis of these transitional subpopulations revealed that TGF-β signaling was highly upregulated in the cell cycle arrest subpopulation and relatively downregulated in transdifferentiating cells. In cultured AEC2s, TGF-β was necessary for cell cycle arrest but impeded transdifferentiation. We conclude that during regeneration after LPS-induced lung injury, TGF-β is a critical signal halting AEC2 proliferation but must be inactivated to allow transdifferentiation. This study provides insight into the molecular mechanisms regulating alveolar regeneration and the pathogenesis of diseases resulting from a failure of regeneration.
Authors
Kent A. Riemondy, Nicole L. Jansing, Peng Jiang, Elizabeth F. Redente, Austin E. Gillen, Rui Fu, Alyssa J. Miller, Jason R. Spence, Anthony N. Gerber, Jay R. Hesselberth, Rachel L. Zemans
Abstract
Because injured mitochondria can accelerate cell death through the elaboration of oxidative free radicals and other mediators, it is striking that proliferator γ coactivator 1-α (PGC1α), a stimulator of increased mitochondrial abundance, protects stressed renal cells instead of potentiating injury. Here, we report that PGC1α’s induction of lysosomes via transcription factor EB (TFEB) may be pivotal for kidney protection. CRISPR and stable gene transfer showed that PGC1α-KO tubular cells were sensitized to the genotoxic stressor cisplatin, whereas Tg cells were protected. The biosensor mitochondrial-targeted Keima (mtKeima) unexpectedly revealed that cisplatin blunts mitophagy both in cells and mice. PGC1α and its downstream mediator NAD+ counteracted this effect. PGC1α did not consistently affect known autophagy pathways modulated by cisplatin. Instead RNA sequencing identified coordinated regulation of lysosomal biogenesis via TFEB. This effector pathway was sufficiently important that inhibition of TFEB or lysosomes unveiled a striking harmful effect of excess PGC1α in cells and conditional mice. These results uncover an unexpected effect of cisplatin on mitophagy and PGC1α’s reliance on lysosomes for kidney protection. Finally, the data illuminate TFEB as a potentially novel target for renal tubular stress resistance.
Authors
Matthew R. Lynch, Mei T. Tran, Kenneth M. Ralto, Zsuzsanna K. Zsengeller, Vinod Raman, Swati S. Bhasin, Nuo Sun, Xiuying Chen, Daniel Brown, Ilsa I. Rovira, Kensei Taguchi, Craig R. Brooks, Isaac E. Stillman, Manoj K. Bhasin, Toren Finkel, Samir M. Parikh
Abstract
In clinical breast cancer intervention, selection of the optimal treatment protocol based on predictive biomarkers remains an elusive goal. Here, we present a modeling tool to predict the likelihood of breast cancer response to neoadjuvant chemotherapy using patient-specific tumor vasculature biomarkers. A semiautomated analysis was implemented and performed on 3990 histological images from 48 patients, with 10–208 images analyzed for each patient. We applied a histology-based mathematical model to 30 resected primary breast cancer tumors and then evaluated a cohort of 18 patients undergoing neoadjuvant chemotherapy, collecting pre- and posttreatment pathology specimens and MRI data. We found that core biopsy samples can be used with acceptable accuracy to determine histological parameters representative of the whole tissue region. Analysis of model histology parameters obtained from tumor vasculature measurements, specifically diffusion distance divided by the radius of the drug-delivering blood vessel (L/rb) and blood volume fraction (BVF), provides a statistically significant separation of patients obtaining a pathologic complete response (pCR) from those who do not. With this model, it is feasible to evaluate primary breast tumor vasculature biomarkers in a patient-specific manner, thereby allowing a precision approach to breast cancer treatment.
Authors
Terisse A. Brocato, Ursa Brown-Glaberman, Zhihui Wang, Reed G. Selwyn, Colin M. Wilson, Edward F. Wyckoff, Lesley C. Lomo, Jennifer L. Saline, Anupama Hooda-Nehra, Renata Pasqualini, Wadih Arap, C. Jeffrey Brinker, Vittorio Cristini
Abstract
Zebrafish are increasingly utilized to model cardiomyopathies and regeneration. Current methods evaluating cardiac function have known limitations, fail to reliably detect focal mechanics, and are not readily feasible in zebrafish. We developed a semiautomated, open-source method — displacement analysis of myocardial mechanical deformation (DIAMOND) — for quantitative assessment of 4D segmental cardiac function. We imaged transgenic embryonic zebrafish in vivo using a light-sheet fluorescence microscopy system with 4D cardiac motion synchronization. Our method permits the derivation of a transformation matrix to quantify the time-dependent 3D displacement of segmental myocardial mass centroids. Through treatment with doxorubicin, and by chemically and genetically manipulating the myocardial injury–activated Notch signaling pathway, we used DIAMOND to demonstrate that basal ventricular segments adjacent to the atrioventricular canal display the highest 3D displacement and are also the most susceptible to doxorubicin-induced injury. Thus, DIAMOND provides biomechanical insights into in vivo segmental cardiac function scalable to high-throughput research applications.
Authors
Junjie Chen, Yichen Ding, Michael Chen, Jonathan Gau, Nelson Jen, Chadi Nahal, Sally Tu, Cynthia Chen, Steve Zhou, Chih-Chiang Chang, Jintian Lyu, Xiaolei Xu, Tzung K. Hsiai, René R. Sevag Packard
Abstract
The identification of new sources of β cells is an important endeavor with therapeutic implications for diabetes. Insulin resistance, in physiological states such as pregnancy or in pathological states such as type 2 diabetes (T2D), is characterized by a compensatory increase in β cell mass. To explore the existence of a dynamic β cell reserve, we superimposed pregnancy on the liver-specific insulin receptor–KO (LIRKO) model of insulin resistance that already exhibits β cell hyperplasia and used lineage tracing to track the source of new β cells. Although both control and LIRKO mice displayed increased β cell mass in response to the relative insulin resistance of pregnancy, the further increase in mass in the latter supported a dynamic source that could be traced to pancreatic ducts. Two observations support the translational significance of these findings. First, NOD/SCID-γ LIRKO mice that became pregnant following cotransplantation of human islets and human ducts under the kidney capsule showed enhanced β cell proliferation and an increase in ductal cells positive for transcription factors expressed during β cell development. Second, we identified duct cells positive for immature β cell markers in pancreas sections from pregnant humans and in individuals with T2D. Taken together, during increased insulin demand, ductal cells contribute to the compensatory β cell pool by differentiation/neogenesis.
Authors
Ercument Dirice, Dario F. De Jesus, Sevim Kahraman, Giorgio Basile, Raymond W.S. Ng, Abdelfattah El Ouaamari, Adrian Kee Keong Teo, Shweta Bhatt, Jiang Hu, Rohit N. Kulkarni
Abstract
Fibrosis is a hallmark in the pathogenesis of various diseases, with very limited therapeutic solutions. A key event in the fibrotic process is the expression of contractile proteins, including α-smooth muscle actin (αSMA) by fibroblasts, which become myofibroblasts. Here, we report the results of a high-throughput screening of a library of approved drugs that led to the discovery of haloperidol, a common antipsychotic drug, as a potent inhibitor of myofibroblast activation. We show that haloperidol exerts its antifibrotic effect on primary murine and human fibroblasts by binding to sigma receptor 1, independent from the canonical transforming growth factor-β signaling pathway. Its mechanism of action involves the modulation of intracellular calcium, with moderate induction of endoplasmic reticulum stress response, which in turn abrogates Notch1 signaling and the consequent expression of its targets, including αSMA. Importantly, haloperidol also reduced the fibrotic burden in 3 different animal models of lung, cardiac, and tumor-associated fibrosis, thus supporting the repurposing of this drug for the treatment of fibrotic conditions.
Authors
Michael Rehman, Simone Vodret, Luca Braga, Corrado Guarnaccia, Fulvio Celsi, Giulia Rossetti, Valentina Martinelli, Tiziana Battini, Carlin Long, Kristina Vukusic, Tea Kocijan, Chiara Collesi, Nadja Ring, Natasa Skoko, Mauro Giacca, Giannino Del Sal, Marco Confalonieri, Marcello Raspa, Alessandro Marcello, Michael P. Myers, Sergio Crovella, Paolo Carloni, Serena Zacchigna
Abstract
The recent Zika virus (ZIKV) epidemic in the Americas has revealed rare but serious manifestations of infection. ZIKV has emerged in regions endemic for dengue virus (DENV), a closely related mosquito-borne flavivirus. Cross-reactive antibodies confound studies of ZIKV epidemiology and pathogenesis. The immune responses to ZIKV may be different in people, depending on their DENV immune status. Here, we focus on the human B cell and antibody response to ZIKV as a primary flavivirus infection to define the properties of neutralizing and protective antibodies generated in the absence of preexisting immunity to DENV. The plasma antibody and memory B cell response is highly ZIKV type–specific, and ZIKV-neutralizing antibodies mainly target quaternary structure epitopes on the viral envelope. To map viral epitopes targeted by protective antibodies, we isolated 2 type-specific monoclonal antibodies (mAbs) from a ZIKV case. Both mAbs were strongly neutralizing in vitro and protective in vivo. The mAbs recognize distinct epitopes centered on domains I and II of the envelope protein. We also demonstrate that the epitopes of these mAbs define antigenic regions commonly targeted by plasma antibodies in individuals from endemic and nonendemic regions who have recovered from ZIKV infections.
Authors
Matthew H. Collins, Huy A. Tu, Ciara Gimblet-Ochieng, Guei-Jiun Alice Liou, Ramesh S. Jadi, Stefan W. Metz, Ashlie Thomas, Benjamin D. McElvany, Edgar Davidson, Benjamin J. Doranz, Yaoska Reyes, Natalie M. Bowman, Sylvia Becker-Dreps, Filemón Bucardo, Helen M. Lazear, Sean A. Diehl, Aravinda M. de Silva
Abstract
Human placenta development and a successful pregnancy is incumbent upon precise oxygen-dependent control of trophoblast migration/invasion. Persistent low oxygen leading to failed trophoblast invasion promotes inadequate spiral artery remodeling, a characteristic of preeclampsia. Angiomotin (AMOT) is a multifaceted scaffolding protein involved in cell polarity and migration, yet its upstream regulation and significance in the human placenta remain unknown. Herein, we show that AMOT is primarily expressed in migratory extravillous trophoblast cells (EVTs) of the intermediate and distal anchoring column. Its expression increases after 10 weeks of gestation when oxygen tension rises and EVT migration/invasion peaks. Time-lapse imaging confirmed that the AMOT 80-kDa isoform promotes migration of trophoblastic JEG3 and HTR-8/SVneo cells. In preeclampsia, however, AMOT expression is decreased and its localization to migratory fetomaternal interface EVTs is disrupted. We demonstrate that Jumonji C domain–containing protein 6 (JMJD6), an oxygen sensor, positively regulates AMOT via oxygen-dependent lysyl hydroxylation. Furthermore, in vitro and ex vivo studies show that transforming growth factor-β (TGF-β) regulates AMOT expression, its interaction with polarity protein PAR6, and its subcellular redistribution from tight junctions to cytoskeleton. Our data reveal an oxygen- and TGF-β–driven migratory function for AMOT in the human placenta, and implicate its deficiency in impaired trophoblast migration that plagues preeclampsia.
Authors
Abby Farrell, Sruthi Alahari, Leonardo Ermini, Andrea Tagliaferro, Michael Litvack, Martin Post, Isabella Caniggia
Abstract
Bacterial permeability family member A1 (BPIFA1), also known as short palate, lung, and nasal epithelium clone 1 (SPLUNC1), is a protein involved in the antiinflammatory response. The goal of this study was to determine whether BPIFA1 expression in asthmatic airways is regulated by genetic variations, altering epithelial responses to type 2 cytokines (e.g., IL-13). Nasal epithelial cells from patients with mild to severe asthma were collected from the National Heart, Lung, and Blood Institute Severe Asthma Research Program centers, genotyped for rs750064, and measured for BPIFA1. To determine the function of rs750064, cells were cultured at air-liquid interface and treated with IL-13 with or without recombinant human BPIFA1 (rhBPIFA1). Noncultured nasal cells with the rs750064 CC genotype had significantly less BPIFA1 mRNA expression than the CT and TT genotypes. Cultured CC versus CT and TT cells without stimulation maintained less BPIFA1 expression. With IL-13 treatment, CC genotype cells secreted more eotaxin-3 than CT and TT genotype cells. Also, rhBPIFA1 reduced IL-13–mediated eotaxin-3. BPIFA1 mRNA levels negatively correlated with serum IgE and fractional exhaled nitric oxide. Baseline FEV1% levels were lower in the asthma patients with the CC genotype (n = 1,016). Our data suggest that less BPIFA1 in asthma patients with the CC allele may predispose them to greater eosinophilic inflammation, which could be attenuated by rhBPIFA1 protein therapy.
Authors
Niccolette Schaefer, Xingnan Li, Max A. Seibold, Nizar N. Jarjour, Loren C. Denlinger, Mario Castro, Andrea M. Coverstone, W. Gerald Teague, Jonathan Boomer, Eugene R. Bleecker, Deborah A. Meyers, Wendy C. Moore, Gregory A. Hawkins, John Fahy, Brenda R. Phillips, David T. Mauger, Azzeddine Dakhama, Shaan Gellatly, Nicole Pavelka, Reena Berman, Y. Peter Di, Sally E. Wenzel, Hong Wei Chu
Abstract
Autoimmune disease is 4 times more common in women than men. This bias is largely unexplained. Female skin is “autoimmunity prone,” showing upregulation of many proinflammatory genes, even in healthy women. We previously identified VGLL3 as a putative transcription cofactor enriched in female skin. Here, we demonstrate that skin-directed overexpression of murine VGLL3 causes a severe lupus-like rash and systemic autoimmune disease that involves B cell expansion, autoantibody production, immune complex deposition, and end-organ damage. Excess epidermal VGLL3 drives a proinflammatory gene expression program that overlaps with both female skin and cutaneous lupus. This includes increased B cell–activating factor (BAFF), the only current biologic target in systemic lupus erythematosus (SLE); IFN-κ, a key inflammatory mediator in cutaneous lupus; and CXCL13, a biomarker of early-onset SLE and renal involvement. Our results demonstrate that skin-targeted overexpression of the female-biased factor VGLL3 is sufficient to drive cutaneous and systemic autoimmune disease that is strikingly similar to SLE. This work strongly implicates VGLL3 as a pivotal orchestrator of sex-biased autoimmunity.
Authors
Allison C. Billi, Mehrnaz Gharaee-Kermani, Joseph Fullmer, Lam C. Tsoi, Brett D. Hill, Dennis Gruszka, Jessica Ludwig, Xianying Xing, Shannon Estadt, Sonya J. Wolf, Syed Monem Rizvi, Celine C. Berthier, Jeffrey B. Hodgin, Maria A. Beamer, Mrinal K. Sarkar, Yun Liang, Ranjitha Uppala, Shuai Shao, Chang Zeng, Paul W. Harms, Monique E. Verhaegen, John J. Voorhees, Fei Wen, Nicole L. Ward, Andrzej A. Dlugosz, J. Michelle Kahlenberg, Johann E. Gudjonsson
Abstract
BACKGROUND. The goal of antiretroviral therapy (ART) is to suppress HIV-1 replication and reconstitute CD4+ T cells. Here, we report on HIV-infected individuals who had a paradoxical decline in CD4+ T cells despite ART-mediated suppression of plasma HIV-1 load (pVL). We defined such an immunological outcome as extreme immune decline (EXID). METHODS. EXID’s clinical and immunological characteristics were compared to immunological responders (IRs), immunological nonresponders (INRs), healthy controls (HCs), and idiopathic CD4+ lymphopenia (ICL) patients. T cell immunophenotyping and assembly/activation of inflammasomes were evaluated by flow cytometry. PBMC transcriptome analysis and genetic screening for pathogenic variants were performed. Levels of cytokines/chemokines were measured by electrochemiluminescence. Luciferase immunoprecipitation system and NK-mediated antibody-dependent cellular cytotoxicity (ADCC) assays were used to identify anti-lymphocyte autoantibodies. RESULTS. EXIDs were infected with non-B HIV-1 subtypes and after 192 weeks of consistent ART-mediated pVL suppression had a median CD4+ decrease of 157 cells/μl, compared with CD4+ increases of 193 cells/μl and 427 cells/μl in INR and IR, respectively. EXID had reduced naive CD4+ T cells, but similar proportions of cycling CD4+ T cells and HLA-DR+CD38+CD8+ T cells compared with IR and INR. Levels of inflammatory cytokines were also similar in EXID and INR, but the IL-7 axis was profoundly perturbed compared with HC, IR, INR, and ICL. Genes involved in T cell and monocyte/macrophage function, autophagy, and cell migration were differentially expressed in EXID. Two of the 5 EXIDs had autoantibodies causing ADCC, while 2 different EXIDs had an increased inflammasome/caspase-1 activation despite consistently ART-suppressed pVL. CONCLUSIONS. EXID is a distinct immunological outcome compared with previously described INR. Anti–CD4+ T cell autoantibodies and aberrant inflammasome/caspase-1 activation despite suppressed HIV-1 viremia are among the mechanisms responsible for EXID.
Authors
Andrea Lisco, Chun-Shu Wong, Silvia Lucena Lage, Itzchak Levy, Jason Brophy, Jeffrey Lennox, Maura Manion, Megan V. Anderson, Yolanda Mejia, Christopher Grivas, Harry Mystakelis, Peter D. Burbelo, Ainhoa Perez-Diez, Adam Rupert, Craig A. Martens, Sarah L. Anzick, Caryn Morse, Shanna Chan, Claire Deleage, Irini Sereti
Abstract
Biased agonism is a paradigm that may explain the selective activation of a signaling pathway via a GPCR that activates multiple signals. The autoantibody-induced inactivation of the calcium-sensing receptor (CaSR) causes acquired hypocalciuric hypercalcemia (AHH). Here, we describe an instructive case of AHH in which severe hypercalcemia was accompanied by an increased CaSR antibody titer. These autoantibodies operated as biased allosteric modulators of CaSR by targeting its Venus flytrap domain near the Ca2+-binding site. A positive allosteric modulator of CaSR, cinacalcet, which targets its transmembrane domain, overcame this autoantibody effect and successfully corrected the hypercalcemia in this patient. Hence, this is the first study to our knowledge that identifies the interaction site of a disease-causing GPCR autoantibody working as its biased allosteric modulator and demonstrates that cinacalcet can correct the AHH autoantibody effects both in vitro and in our AHH patient. Our observations provide potentially new insights into how biased agonism works and how to design a biased allosteric modulator of a GPCR. Our observations also indicate that the diagnosis of AHH is important because the severity of hypercalcemia may become fatal if the autoantibody titer increases. Calcimimetics may serve as good treatment options for some patients with severe AHH.
Authors
Noriko Makita, Takao Ando, Junichiro Sato, Katsunori Manaka, Koji Mitani, Yasuko Kikuchi, Takayoshi Niwa, Masanori Ootaki, Yuko Takeba, Naoki Matsumoto, Atsushi Kawakami, Toshihisa Ogawa, Masaomi Nangaku, Taroh Iiri
Abstract
Myelomonocytic cells are critically involved in iron turnover as aged RBC recyclers. Human monocytes are divided in 3 subpopulations of classical, intermediate, and nonclassical cells, differing in inflammatory and migratory phenotype. Their functions in iron homeostasis are, however, unclear. Here, we asked whether the functional diversity of monocyte subsets translates into differences in handling physiological and pathological iron species. By microarray data analysis and flow cytometry we identified a set of iron-related genes and proteins upregulated in classical and, in part, intermediate monocytes. These included the iron exporter ferroportin (FPN1), ferritin, transferrin receptor, putative transporters of non-transferrin-bound iron (NTBI), and receptors for damaged erythrocytes. Consequently, classical monocytes displayed superior scavenging capabilities of potentially toxic NTBI, which were augmented by blocking iron export via hepcidin. The same subset and, to a lesser extent, the intermediate population, efficiently cleared damaged erythrocytes in vitro and mediated erythrophagocytosis in vivo in healthy volunteers and patients having received blood transfusions. To summarize, our data underline the physiologically important function of the classical and intermediate subset in clearing NTBI and damaged RBCs. As such, these cells may play a nonnegligible role in iron homeostasis and limit iron toxicity in iron overload conditions.
Authors
David Haschka, Verena Petzer, Florian Kocher, Christoph Tschurtschenthaler, Benedikt Schaefer, Markus Seifert, Sieghart Sopper, Thomas Sonnweber, Clemens Feistritzer, Tara L. Arvedson, Heinz Zoller, Reinhard Stauder, Igor Theurl, Guenter Weiss, Piotr Tymoszuk
In-Press Preview - More
Abstract
Epidemiological findings indicate that coinfection with influenza viruses is associated with an increased risk of death in patients suffering from tuberculosis but the underlying pathomechanisms are not well understood. In this study, we demonstrate that influenza A virus (IAV) coinfection rapidly impairs control of Mycobacterium tuberculosis (Mtb) in C57BL/6 mice. IAV coinfection was associated with significantly increased bacterial loads, reduced survival and a substantial modulation of innate and adaptive immune defenses including an impaired onset and development of Mtb-specific CD4+ T cell responses and the accumulation of macrophages with increased arginase-1 production in the lungs. Our findings strongly indicate that IAV coinfection compromises the host’s ability to control Mtb infection via the production of IL-10 which was rapidly induced upon viral infection. The blockade of IL-10 receptor signaling reduced the bacterial load in coinfected mice to a level comparable with that in Mtb-only-infected animals. Taken together, our data suggest that IL-10 signaling constitutes a major pathway that enhances susceptibility to Mtb during concurrent IAV infection.
Authors
Sarah Ring, Lars Eggers, Jochen Behrends, Adam Wutkowski, Dominik Schwudke, Andrea Kröger, Alexandra Maximiliane Hierweger, Christoph Hölscher, Gülsah Gabriel, Bianca Schneider
Abstract
Gain of the long arm of chromosome 17 (17q) is a cytogenetic hallmark of high-risk neuroblastoma, yet its contribution to neuroblastoma pathogenesis remains incompletely understood. Combining whole-genome and RNA sequencing of neuroblastomas, we identified the prohibitin (PHB) gene as highly expressed in tumors with 17q gain. High PHB expression correlated with poor prognosis and was associated with loss of gene expression programs promoting neuronal development and differentiation. PHB depletion induced differentiation and apoptosis and slowed cell cycle progression of neuroblastoma cells, at least in part through impaired ERK1/2 activation. Conversely, ectopic expression of PHB was sufficient to increase proliferation of neuroblastoma cells and was associated with suppression of markers associated with neuronal differentiation and favorable neuroblastoma outcome. Thus, PHB is a 17q oncogene in neuroblastoma that promotes tumor cell proliferation, and de-differentiation.
Authors
Ian C. MacArthur, Yi Bei, Heathcliff Dorado García, Michael V. Ortiz, Joern Toedling, Filippos Klironomos, Jana Rolff, Angelika Eggert, Johannes H. Schulte, Alex Kentsis, Anton G. Henssen
Abstract
The bone marrow microenvironment (BMME) contributes to the regulation of hematopoietic stem cell (HSC) function, though its role in age-associated lineage skewing is poorly understood. Here we show that dysfunction of aged marrow macrophages (Mφs) directs HSC platelet-bias. Mφs from the marrow of aged mice and humans exhibited an activated phenotype, with increased expression of inflammatory signals. Aged marrow Mφs also displayed decreased phagocytic function. Senescent neutrophils, typically cleared by marrow Mφs, were markedly increased in aged mice, consistent with functional defects in Mφ phagocytosis and efferocytosis. In aged mice, Interleukin 1B (IL1B) was elevated in the bone marrow and caspase 1 activity, which can process pro-IL1B, was increased in marrow Mφs and neutrophils. Mechanistically, IL1B signaling was necessary and sufficient to induce a platelet bias in HSCs. In young mice, depletion of phagocytic cell populations or loss of the efferocytic receptor Axl expanded platelet-biased HSCs. Our data support a model wherein increased inflammatory signals and decreased phagocytic function of aged marrow Mφs induce the acquisition of platelet bias in aged HSCs. This work highlights the instructive role of Mφs and IL1B in the age-associated lineage-skewing of HSCs, and reveals the therapeutic potential of their manipulation as antigeronic targets.
Authors
Benjamin J. Frisch, Corey M. Hoffman, Sarah E. Latchney, Mark W. LaMere, Jason Myers, John Ashton, Allison J. Li, Jerry Saunders, James Palis, Archibald S. Perkins, Amanda McCabe, Julianne N. Smith, Kathleen E. McGrath, Fatima Rivera-Escalera, Andrew McDavid, Jane L. Liesveld, Vyacheslav A. Korshunov, Michael R. Elliott, Katherine C. MacNamara, Michael W. Becker, Laura M. Calvi
Abstract
Children with trisomy 21 (Down syndrome [DS]) have a 130-fold increased incidence of Hirschsprung Disease (HSCR), a developmental defect where the enteric nervous system (ENS) is missing from distal bowel (i.e., distal bowel is aganglionic). Treatment for HSCR is surgical resection of aganglionic bowel, but many children have bowel problems after surgery. Post-surgical problems like enterocolitis and soiling are especially common in children with DS. To determine how trisomy 21 affects ENS development, we evaluated the ENS in two DS mouse models, Ts65Dn and Tc1. These mice are trisomic for many chromosome 21 homologous genes, including Dscam and Dyrk1a, which are hypothesized to contribute to HSCR risk. Ts65Dn and Tc1 mice have normal ENS precursor migration at E12.5 and almost normal myenteric plexus structure as adults. However, Ts65Dn and Tc1 mice have markedly reduced submucosal plexus neuron density throughout the bowel. Surprisingly, the submucosal neuron defect in Ts65Dn mice is not due to excess Dscam or Dyrk1a, since normalizing copy number for these genes does not rescue the defect. These findings suggest the possibility that the high frequency of bowel problems in children with DS and HSCR may occur because of additional unrecognized problems with ENS structure.
Authors
Ellen M. Schill, Christina M. Wright, Alisha Jamil, Jonathan M. LaCombe, Randall J. Roper, Robert O. Heuckeroth
Abstract
Background: Current dosing of intrapleural fibrinolytic therapy (IPFT) in adults with complicated parapneumonic effusion (CPE) / empyema is empiric, as dose-escalation trials have not previously been conducted. We hypothesized that LTI-01 (scuPA), which is relatively resistant to PA inhibitor-1 (PAI-1), would be well-tolerated. Methods: This was an open-label, dose-escalation trial of LTI-01 IPFT at 50,000-800,000 IU daily for up to 3 days in adults with loculated CPE/empyema and failed pleural drainage. The primary objective was to evaluate safety and tolerability, and secondary objectives included assessments of processing and bioactivity of scuPA in blood and pleural fluid (PF), and early efficacy. Results: LTI-01 was well tolerated with no bleeding, treatment-emergent adverse events or surgical referrals (n=14 subjects). uPA antigen increased in PFs at 3 hours after LTI-01 (p<0.01) but not in plasma. PF saturated active PAI-1, generated PAI-1-resistant bioactive complexes, increased PA and fibrinolytic activities and D-dimers. There was no systemic fibrinogenolysis, nor increments in plasma D-dimer. Decreased pleural opacities occurred in all but one subject. Both subjects receiving 800,000 IU required two doses to relieve pleural sepsis, with two other subjects similarly responding at lower doses. Conclusion: LTI-01 IPFT was well-tolerated at these doses with no safety concerns. Bioactivity of LTI-01 IPFT was confirmed, limited to PFs where its processing simulated that previously reported in preclinical studies. Preliminary efficacy signals including reduction of pleural opacity were observed.
Authors
Lutz Beckert, Ben Brockway, Graham Simpson, Anne Marie Southcott, Y.C. Gary Lee, Najib Rahman, Richard W. Light, Steven Shoemaker, John Gillies, Andrey A. Komissarov, Galina Florova, Timothy Ochran, William Bradley, Harrison Ndetan, Karan P. Singh, Krishna Sarva, Steven Idell
View more articles by topic:
JCI Insight is a peer-reviewed journal published by the American Society for Clinical Investigation dedicated to well-executed preclinical and clinical research studies. The journal was founded in 2016 and is headed by Editor in Chief Howard Rockman.
Submit your work
