Sepsis survivors exhibit impaired responsiveness to antigen (Ag) challenge associated with increased mortality from infection. The contribution of follicular dendritic cells (FDCs) in the impaired humoral response in sepsis-surviving mice is investigated in this study. We demonstrated that mice subjected to sepsis from cecal ligation and puncture (CLP) have reduced NP-specific high-affinity class-switched Ig antibodies compared to sham control mice following immunization with the T-dependent Ag, NP-CGG. NP-specific germinal center (GC) B cells in CLP mice exhibited reduced TNFα and AID mRNA expression compared to sham mice. CLP mice showed a reduction in FDC clusters, a reduced binding of immune complexes on FDCs, and reduced mRNA expression of CR2, ICAM-1, VCAM-1, FcγRIIB, TNFR1, IKK2 and LTbR compared to sham mice. Adoptive transfer studies showed there was no B cell-intrinsic defect. In summary, our data suggest that the reduced Ag-specific antibody response in CLP mice is secondary to a disruption in FDC and GC B cell function.
Minakshi Rana, Andrea La Bella, Rivka Lederman, Bruce T. Volpe, Barbara Sherry, Betty Diamond
Interstitial kidney inflammation is present in various nephritides in which serum IL-23 is elevated. Here we show that IL-23 receptor (IL-23R) expressing murine and human renal tubular epithelial cells (TEC) respond to IL-23 by inducing intracellular calcium flux, enhanced glycolysis, and the upregulation of calcium/calmodulin kinase IV (CaMK4) which results in suppression of the expression of the arginine degrading enzyme arginase 1 (ARG1) thus increasing in situ levels of free L-Arginine (Arg). Limited availability of Arg suppresses the ability of infiltrating T cells to proliferate and produce inflammatory cytokines. TEC from humans and mice with nephritis express increased levels of IL-23R and CaMK4 but reduced levels of ARG1. TEC-specific deletion of Il23r or Camk4 suppressed inflammation whereas deletion of Arg1 exacerbated inflammation in different murine disease models. Finally, TEC-specific delivery of a CaMK4 inhibitor specifically curbed the renal inflammation in lupus-prone mice without affecting systemic inflammation. Our data offer first evidence on the immunosuppressive capacity of TEC through a mechanism that involves competitive uptake of Arg and signify the importance of modulation of an inflammatory cytokine on the function of non-lymphoid cells, which leads to the establishment of an inflammatory microenvironment. New approaches to treat kidney inflammation should consider restoring the immunosuppressive capacity of TEC.
Hao Li, Maria G. Tsokos, Rhea Bhargava, Iannis E. Adamopoulos, Hanni Menn-Josephy, Isaac E. Stillman, Philip Rosenstiel, Jarrat Jordan, George C. Tsokos
Severe asthma remains challenging to manage with limited treatment options. We have previously shown that targeting smooth muscle integrin α5β1 interaction with fibronectin can mitigate the effects of airway hyperresponsiveness by impairing force transmission. In this paper we show that another member of the integrin superfamily, integrin α2β1, is present in airway smooth muscle and capable of regulating force transmission via cellular tethering to the matrix protein collagen I, and to a lesser degree, laminin-111. The addition of an inhibitor of integrin α2β1 impaired IL-13-enhanced contraction in mouse tracheal rings and human bronchial rings, and abrogated the exaggerated bronchoconstriction induced by allergen sensitization and challenge. We confirmed that this effect was not due to alterations in classic intracellular myosin light chain phosphorylation regulating muscle shortening. Although IL-13 did not affect surface expression of α2β1, it did increase α2β1-mediated adhesion and the level of expression of an activation-specific epitope on the β1 subunit. We developed a method to simultaneously quantify airway narrowing and muscle shortening using two-photon microscopy and demonstrated that inhibition of α2β1 mitigated IL-13-enhanced airway narrowing without altering muscle shortening by impairing the tethering of muscle to the surrounding matrix. Our data identify cell-matrix tethering as an attractive therapeutic target to mitigate the severity of airway contraction in asthma.
Sean Liu, Uyen Ngo, Xin-Zi Tang, Xin Ren, Wenli Qiu, Xiaozhu Huang, William DeGrado, Christopher D.C. Allen, Hyunil Jo, Dean Sheppard, Aparna B. Sundaram
Chronic cellular stress associated with neurodegenerative disease can result in the persistence of stress granule (SG) structures, membraneless organelles that form in response to cellular stress. In Huntington’s disease (HD), chronic expression of mutant huntingtin generates various forms of cellular stress, including activation of the unfolded protein response and oxidative stress. However, it has yet to be determined whether SGs are a feature of HD neuropathology. We examined the miRNA composition of extracellular vesicles (EVs) present in the cerebrospinal fluid (CSF) of HD patients and show that a subset of their target mRNAs were differentially expressed in the prefrontal cortex of HD patients. Of these targets, SG components were enriched, including the SG nucleating Ras GTPase-activating protein-binding protein 1 (G3BP1). We investigated localization and levels of G3BP1 and found a significant increase in the density of G3BP1-positive granules in the cortex and hippocampus of R6/2 transgenic mice and in the superior frontal cortex of HD patient brains. Intriguingly, we also observed that the SG-associated TAR DNA-Binding Protein-43 (TDP43), a nuclear RNA/DNA binding protein, was mislocalized to the cytoplasm of G3BP1-granule positive HD cortical neurons. These findings suggest that G3BP1 SG dynamics may play a role in the pathophysiology of HD.
Isabella I. Sanchez, Thai B. Nguyen, Whitney E. England, Ryan G. Lim, Anthony Q. Vu, Ricardo Miramontes, Lauren M. Byrne, Sebastian Markmiller, Alice L. Lau, Iliana Orellana, Maurice A. Curtis, Richard Lewis Maxwell Faull, Gene W. Yeo, Christie D. Fowler, Jack C. Reidling, Edward J. Wild, Robert C. Spitale, Leslie M. Thompson
T cells are involved in control of COVID-19, but limited knowledge is available on the relationship between antigen-specific T cell response and disease severity. Here, we assessed the magnitude, function and phenotype of SARS-CoV-2-specific CD4 T cells in 95 hospitalized COVID-19 patients (38 of them being HIV-1 and/or tuberculosis (TB) co-infected) and 38 non-COVID-19 patients, using flow cytometry. We showed that SARS-CoV-2-specific CD4 T cell attributes, rather than magnitude, associates with disease severity, with severe disease being characterized by poor polyfunctional potential, reduced proliferation capacity and enhanced HLA-DR expression. Moreover, HIV-1 and TB co-infection skewed the SARS-CoV-2 T cell response. HIV-1 mediated CD4 T cell depletion associated with suboptimal T cell and humoral immune responses to SARS-CoV-2; and a decrease in the polyfunctional capacity of SARS-CoV-2-specific CD4 T cells was observed in COVID-19 patients with active TB. Our results also revealed that COVID-19 patients displayed reduced frequency of Mtb-specific CD4 T cells, with possible implications for TB disease progression. There results corroborate the important role of SARS-CoV-2-specific T cells in COVID-19 pathogenesis and support the concept of altered T cell functions in patients with severe disease.
Catherine Riou, Elsa Du Bruyn, Cari Stek, Remy Daroowala, Rene T. Goliath, Fatima Abrahams, Qonita Said-Hartley, Brian W. Allwood, Nei-Yuan Hsiao, Katalin A. Wilkinson, Cecilia S. Lindestam Arlehamn, Alessandro Sette, Sean Wasserman, Robert J. Wilkinson
Cutaneous melanoma remains the most lethal skin cancer, and ranks third among all malignancies in terms of years of life lost. Despite the advent of immune checkpoint and targeted therapies, only roughly half of patients with advanced melanoma achieves a durable remission. SIRT5 is a member of the sirtuin family of protein deacylases that regulate metabolism and other biological processes. Germline Sirt5 deficiency is associated with mild phenotypes in mice. Here we show that SIRT5 is required for proliferation and survival across all cutaneous melanoma genotypes tested, as well as uveal melanoma, a genetically distinct melanoma subtype that arises in the eye and is incurable once metastatic. Likewise, SIRT5 is required for efficient tumor formation by melanoma xenografts and in an autochthonous mouse Braf;Pten-driven melanoma model. Via metabolite and transcriptomic analyses, we find that SIRT5 is required to maintain histone acetylation and methylation levels in melanoma cells, thereby promoting proper gene expression. SIRT5-dependent genes notably include MITF, a key lineage-specific survival oncogene in melanoma, and the c-MYC proto-oncogene. SIRT5 may represent a novel, druggable genotype-independent addiction in melanoma.
William Giblin, Lauren Bringman-Rodenbarger, Angela H. Guo, Surinder Kumar, Alexander C. Monovich, Ahmed M. Mostafa, Mary E. Skinner, Michelle Azar, Ahmed S.A. Mady, Carolina H. Chung, Namrata Kadambi, Keith-Allen Melong, Ho-Joon Lee, Li Zhang, Peter Sajjakulnukit, Sophie Trefely, Erika L. Varner, Sowmya Iyer, Min Wang, James S. Wilmott, H. Peter Soyer, Richard A. Sturm, Antonia L. Pritchard, Aleodor A. Andea, Richard A. Scolyer, Mitchell S. Stark, David A. Scott, Douglas R. Fullen, Marcus W. Bosenberg, Sriram Chandrasekaran, Zaneta Nikolovska-Coleska, Monique E. Verhaegen, Nathaniel W. Snyder, Miguel N. Rivera, Andrei Osterman, Costas A. Lyssiotis, David B. Lombard
Down syndrome (DS), caused by trisomy of chromosome 21, occurs in 1 of every 800 live births. Early defects in cortical development likely account for the cognitive impairments in DS, although the underlying molecular mechanism remains elusive. Here, we performed histological assays and unbiased single-cell RNA sequencing (scRNA-seq) analysis on cerebral organoids derived from four euploid cell lines and from induced pluripotent stem cells (iPSCs) from three individuals with trisomy 21 to explore cell type-specific abnormalities associated with DS during early brain development. We found that neurogenesis was significantly affected based on diminished proliferation and decreased expression of layer II and IV markers in cortical neurons in the subcortical regions; this may be responsible for the reduced size of the organoids. Furthermore, suppression of the DSCAM-PAK1 pathway which showed enhanced activities in DS) via CRISPR/Cas9, CRISPRi or small-molecule inhibitor treatment reverses abnormal neurogenesis, thereby increasing the size of organoids derived from DS iPSCs. Our study demonstrated that 3D cortical organoids developed in vitro are a valuable model of DS and provided a direct link between dysregulation of the DSCAM-PAK1 pathway and developmental brain defects in DS.
Xiao-Yan Tang, Lei Xu, Jingshen Wang, Yuan Hong, Yuanyuan Wang, Qian Zhu, Da Wang, Xin-Yue Zhang, Chun-Yue Liu, Kai-Heng Fang, Xiao Han, Shihua Wang, Xin Wang, Min Xu, Anita Bhattacharyya, Xing Guo, Mingyan Lin, Yan Liu
Myelofibrosis (MF) are a non-BCR-ABL myeloproliferative neoplasms (NMPs) associated with poor outcomes. Current treatment has little effect on the natural history of the disease. MF results from complex interactions between 1) the malignant clone, 2) an inflammatory context, and 3) remodeling of the bone marrow (BM) microenvironment. Each of these points is a potential target of PPAR-y activation. Here, we demonstrated the therapeutic potential of PPAR-y agonists in resolving MF in three mouse models. We showed that PPAR-y agonists reduce myeloproliferation, modulate inflammation, and protect the BM stroma in vitro and ex vivo. Activation of PPAR-y constitutes a relevant therapeutic target in MF and our data support the possibility of using PPAR-y agonists in clinical practice.
Juliette Lambert, Joseph Saliba, Carolina Calderon, Karine Sii-Felice, Mohammad Salma, Valérie Edmond, Jean-Claude Alvarez, Marc Delord, Caroline Marty, Isabelle Plo, Jean-Jacques Kiladjian, Eric Soler, William Vainchenker, Jean-Luc Villeval, Philippe Rousselot, Stephane Prost
Melanoma dedifferentiation has been reported as a state of cellular resistance to targeted therapies and immunotherapies as cancer cells revert to a more primitive cellular phenotype. Here we show that, counterintuitively, the biopsies of patient tumors that respond to anti-programmed cell death receptor 1 (PD-1) therapy decreased expression of melanocytic markers and increased neural crest markers, suggesting treatment-induced dedifferentiation. When modeling the effects in vitro, we documented that melanoma cell lines that were originally melanocytic differentiated underwent a process of neural crest dedifferentiation when continuously exposed to interferon gamma, through global chromatin landscape changes leading to enrichment in specific hyperaccessible chromatin regions. The interferon gamma-induced dedifferentiation signature corresponded with improved outcomes in patients with melanoma, challenging the notion that neural crest dedifferentiation is entirely an adverse phenotype.
Yeon Joo Kim, Katherine M. Sheu, Jennifer Tsoi, Gabriel Abril-Rodriguez, Egmidio Medina, Catherine S. Grasso, Davis Y. Torrejon, Ameya S. Champhekar, Kevin Litchfield, Charles Swanton, Daniel E. Speiser, Philip O. Scumpia, Alexander Hoffmann, Thomas G. Graeber, Cristina Puig-Saus, Antoni Ribas
Disrupting transmission of Borrelia burgdorferi (B. burgdorferi ) from infected ticks to humans is one strategy to prevent the significant morbidity from Lyme disease. We have previously shown that an anti-OspA human monoclonal antibody, 2217, prevents transmission of B. burgdorferi from infected ticks in animal models. Maintenance of a protective plasma concentration of a human monoclonal antibody for tick season presents a significant challenge for a pre-exposure prophylaxis strategy. Here, we describe the optimization of 2217 by amino acid substitutions (LS, M428L and N434S) into the Fc domain. The LS mutation led to a twofold-increase in half-life in cynomolgus monkeys. In a rhesus macaque model, 2217LS protected animals from tick transmission of spirochetes at a dose of 3 mg/kg. Crystallographic analysis of Fab in complex with OspA reveals that 2217 binds a novel epitope that is highly conserved among the B. burgdorferi, B. garinii, and B. afzelii species. Unlike most vaccines that may require boosters to achieve protection, our work supports the development of 2217LS as an effective pre-exposure prophylaxis in Lyme-endemic regions with a single dose at the beginning of tick season offering immediate protection that remains for the duration of exposure risk.
Zachary A. Schiller, Michael J. Rudolph, Jacqueline R. Toomey, Monir Ejemel, Alan LaRochelle, Simon A. Davis, Havard S. Lambert, Aurélie Kern, Amanda C. Tardo, Colby A. Souders, Eric Peterson, Rebecca D. Cannon, Chandrashekar Ganesa, Frank Fazio, Nicholas J. Mantis, Lisa A. Cavacini, John Sullivan-Bolyai, Linden T. Hu, Monica E. Embers, Mark S. Klempner, Yang Wang
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