Concise Communication

Abstract

The microbiome provides resistance to infection. However, mechanisms for this are poorly understood. Here we demonstrate in a murine model that colonization with the intestinal bacterium Clostridium scindens provided protection from Entamoeba histolytica colitis via innate immunity. Introduction of C. scindens into the gut microbiota epigenetically altered and expanded bone marrow granulocyte-monocyte-progenitors (GMPs) and resulted in increased intestinal neutrophils with subsequent challenge with E. histolytica. Introduction of C. scindens alone was sufficient to expand GMPs in gnotobiotic mice. Adoptive transfer of bone-marrow from C. scindens colonized-mice into naïve-mice protected against amebic colitis and increased intestinal neutrophils. Children without E. histolytica diarrhea also had a higher abundance of Lachnoclostridia. Because of the known ability of the Lachnoclostridia C. scindens to metabolize the bile salt cholate, we measured deoxycholate and discovered that it was increased in the sera of C. scindens colonized specific pathogen free and gnotobiotic mice, as well as in children protected from amebiasis. Administration of deoxycholate alone (in the absence of C. scindens) increased GMPs and provided protection from amebiasis. We have discovered a mechanism by which C. scindens and the microbially-metabolized bile salt deoxycholic acid alter hematopoietic precursors and provide innate protection from later infection with Entamoeba histolytica.

Authors

Stacey L. Burgess, Jhansi L. Leslie, Md. Jashim Uddin, David Noah Oakland, Carol A. Gilchrist, G. Brett Moreau, Koji Watanabe, Mahmoud M. Saleh, Morgan Simpson, Brandon A. Thompson, David T. Auble, Stephen D. Turner, Natasa Giallourou, Jonathan Swann, Zhen Pu, Jennie Z. Ma, Rashidul Haque, William A. Petri, Jr.

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Abstract

Molecular mechanisms governing the development of mammalian cochlea, the hearing organ, remain largely unknown. Through genome sequencing in three subjects from two families with non-syndromic cochlear aplasia, we identified homozygous 221 KB and 338 KB deletions in a non-coding region on chromosome 8 with an ~200 KB overlapping section. Genomic location of the overlapping deleted region was starting from ~350 KB downstream of GDF6. Otic lineage cells differentiated from induced pluripotent stem cells derived from an affected individual show reduced expression of GDF6 compared to control cells. A mouse knock-out of Gdf6 reveals cochlear aplasia closely resembling the human phenotype. We conclude that GDF6 plays a necessary role in early cochlear development controlled by cis-regulatory elements located within ~500 KB region of the genome in humans and that its disruption leads to deafness due to cochlear aplasia.

Authors

Guney Bademci, Clemer Abad, Filiz Basak Cengiz, Serhat Seyhan, Armagan Incesulu, Shengru Guo, Suat Fitoz, Emine Ikbal Atli, Nicholas C. Gosstola, Selma Demir, Brett M. Colbert, Gozde Cosar Seyhan, Claire J. Sineni, Duygu Duman, Hakan Gurkan, Cynthia Casson Morton, Derek M. Dykxhoorn, Katherina Walz, Mustafa Tekin

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Abstract

Infusion of the broadly neutralizing antibody VRC01 has been evaluated in HIV-1 chronically infected individuals. Here we studied how VRC01 infusions impacted viral rebound after cessation of antiretroviral therapy (ART) in 18 acutely-treated and durably-suppressed individuals. Viral rebound occurred in all individuals, yet VRC01 infusions modestly delayed rebound and participants who showed a faster decay of VRC01 in serum rebounded more rapidly (Rho=0.60, p=0.03). Participants with strains most sensitive to VRC01 or with VRC01 epitope motifs similar to known VRC01-susceptible strains rebounded later (Rho=-0.70, p<0.03). Upon rebound, HIV-1 sequences were indistinguishable from those sampled at diagnosis. Across the cohort, participant derived Env showed different sensitivity to VRC01 neutralization (including two resistant viruses), yet neutralization sensitivity was similar at diagnosis and post-rebound, indicating the lack of selection for VRC01-resistance during treatment interruption.Our results showed that viremia rebounded despite the absence of HIV-1 adaptation to VRC01 and an average VRC01 trough of 221µg/mL. While VRC01 levels were insufficient to prevent a resurgent infection, knowledge that they did not mediate Env mutations in acute-like viruses is relevant for antibody-based strategies in acute infection.

Authors

Evan M. Cale, Hongjun Bai, Meera Bose, Michael A. Messina, Donn Colby, Eric Sanders-Buell, Bethany L. Dearlove, Yifan Li, Emily Engeman, Daniel Silas, Anne Marie O’Sullivan, Brendan Mann, Suteeraporn Pinyakorn, Jintana Intasan, Khunthalee Benjapornpong, Carlo Sacdalan, Eugene Kroon, Nittaya Phanuphak, Robert Gramzinski, Sandhya Vasan, Merlin L. Robb, Nelson L. Michael, Rebecca M. Lynch, Robert Bailer, Amélie Pagliuzza, Nicolas Chomont, Amarendra Pegu, Nicole A. Doria-Rose, Lydie Trautmann, Trevor A. Crowell, John Mascola, Jintanat Ananworanich, Sodsai Tovanabutra, Morgane Rolland

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Abstract

Kallikrein-related peptidase 6 (KLK6) is a secreted serine protease hypothesized to promote inflammation via cleavage of protease-activated receptors (PAR)1 and PAR2. KLK6 levels are elevated in multiple inflammatory and autoimmune conditions, but no definitive role in pathogenesis has been established. Here, we show that skin-targeted overexpression of KLK6 causes generalized, severe psoriasiform dermatitis with spontaneous development of debilitating psoriatic arthritis-like joint disease. The psoriatic skin and joint phenotypes are reversed by normalization of skin KLK6 levels and attenuated following genetic elimination of PAR1 but not PAR2. Conservation of this regulatory pathway was confirmed in human psoriasis using vorapaxar, an FDA-approved PAR1 antagonist, on explanted lesional skin from psoriasis patients. Beyond defining a critical role for KLK6-PAR1 signaling in promoting psoriasis, our results demonstrate that KLK6-PAR1-mediated inflammation in the skin alone is sufficient to drive inflammatory joint disease. Further, we identify PAR1 as a promising cytokine-independent target in therapy of psoriasis and psoriatic arthritis.

Authors

Allison C. Billi, Jessica E. Ludwig, Yi Fritz, Richard Rozic, William R. Swindell, Lam C. Tsoi, Dennis Gruszka, Shahla Abdollahi-Roodsaz, Xianying Xing, Doina Diaconu, Ranjitha Uppala, Maya I. Camhi, Philip A. Klenotic, Mrinal K. Sarkar, M. Elaine Husni, Jose U. Scher, Christine McDonald, J. Michelle Kahlenberg, Ronald J. Midura, Johann E. Gudjonsson, Nicole L. Ward

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Abstract

Tyrosine kinase domain (TKD) mutations contribute to acquired resistance to FMS-like tyrosine kinase 3 (FLT3) inhibitors used to treat FLT3-mutant acute myeloid leukemia (AML). We report a cocrystal structure of FLT3 with a type I inhibitor, NCGC1481, that retained potent binding and activity against FLT3 TKD and gatekeeper mutations. Relative to the current generation of advanced FLT3 inhibitors, NCGC1481 exhibited superior antileukemic activity against the common, clinically relevant FLT3-mutant AML cells in vitro and in vivo.

Authors

LaQuita M. Jones, Katelyn Melgar, Lyndsey Bolanos, Kathleen Hueneman, Morgan M. Walker, Jian-Kang Jiang, Kelli M. Wilson, Xiaohu Zhang, Jian Shen, Fan Jiang, Patrick Sutter, Amy Wang, Xin Xu, Gregory J. Tawa, Scott B. Hoyt, Mark Wunderlich, Eric O’Brien, John P. Perentesis, Daniel T. Starczynowski, Craig J. Thomas

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Abstract

Peptide MHC class II–based (pMHCII-based) nanomedicines trigger the formation of multicellular regulatory networks by reprogramming autoantigen-experienced CD4+ T cells into autoimmune disease–suppressing T regulatory type 1 (TR1) cells. We have shown that pMHCII-based nanomedicines displaying liver autoimmune disease–relevant yet ubiquitously expressed antigens can blunt various liver autoimmune disorders in a non–disease-specific manner without suppressing local or systemic immunity against infectious agents or cancer. Here, we show that such ubiquitous autoantigen-specific T cells are also awakened by extrahepatic tissue damage and that the corresponding TR1 progeny can suppress experimental autoimmune encephalomyelitis (EAE) and pancreatic β cell autoreactivity. In mice having EAE, nanomedicines displaying either ubiquitous or CNS-specific epitopes triggered the formation and expansion of cognate TR1 cells and their recruitment to the CNS-draining lymph nodes, sparing their liver-draining counterparts. Surprisingly, in mice having both liver autoimmunity and EAE, liver inflammation sequestered these ubiquitous or even CNS-specific TR1 cells away from the CNS, abrogating their antiencephalitogenic activity. In these mice, only the ubiquitous antigen-specific TR1 cells suppressed liver autoimmunity. Thus, the scope of antigen spreading in autoimmune disorders is larger than previously anticipated, involving specificities expected to be silenced by mechanisms of tolerance; the regulatory activity, but not the retention of autoreactive TR1 cells, requires local autoantigen expression.

Authors

Channakeshava Sokke Umeshappa, Jacques Mbongue, Santiswarup Singha, Saswat Mohapatra, Jun Yamanouchi, Justin A. Lee, Roopa Hebbandi Nanjundappa, Kun Shao, Urs Christen, Yang Yang, Kristofor K. Ellestad, Pere Santamaria

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Abstract

Notch signaling is a highly conserved intercellular pathway with tightly regulated and pleiotropic roles in normal tissue development and homeostasis. Dysregulated Notch signaling has also been implicated in human disease, including multiple forms of cancer, and represents an emerging therapeutic target. Successful development of such therapeutics requires a detailed understanding of potential on-target toxicities. Here, we identify autosomal dominant mutations of the canonical Notch ligand Jagged1 (or JAG1) as a cause of peripheral nerve disease in 2 unrelated families with the hereditary axonal neuropathy Charcot-Marie-Tooth disease type 2 (CMT2). Affected individuals in both families exhibited severe vocal fold paresis, a rare feature of peripheral nerve disease that can be life-threatening. Our studies of mutant protein posttranslational modification and localization indicated that the mutations (p.Ser577Arg, p.Ser650Pro) impair protein glycosylation and reduce JAG1 cell surface expression. Mice harboring heterozygous CMT2-associated mutations exhibited mild peripheral neuropathy, and homozygous expression resulted in embryonic lethality by midgestation. Together, our findings highlight a critical role for JAG1 in maintaining peripheral nerve integrity, particularly in the recurrent laryngeal nerve, and provide a basis for the evaluation of peripheral neuropathy as part of the clinical development of Notch pathway–modulating therapeutics.

Authors

Jeremy M. Sullivan, William W. Motley, Janel O. Johnson, William H. Aisenberg, Katherine L. Marshall, Katy E.S. Barwick, Lingling Kong, Jennifer S. Huh, Pamela C. Saavedra-Rivera, Meriel M. McEntagart, Marie-Helene Marion, Lucy A. Hicklin, Hamid Modarres, Emma L. Baple, Mohamed H. Farah, Aamir R. Zuberi, Cathleen M. Lutz, Rachelle Gaudet, Bryan J. Traynor, Andrew H. Crosby, Charlotte J. Sumner

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Abstract

Chronic pancreatitis (CP) is considered an irreversible fibroinflammatory pancreatic disease. Despite numerous animal model studies, questions remain about local immune characteristics in human CP. We profiled pancreatic immune cell characteristics in control organ donors and CP patients that included hereditary and idiopathic CP undergoing total pancreatectomy with islet auto-transplantation. Flow cytometric analysis revealed a significant increase in the frequency of CD68+ macrophages in idiopathic CP. In contrast, hereditary CP showed a significant increase in CD3+ T cell frequency, which prompted us to investigate the T cell receptor β (TCRβ) repertoire in CP and controls. TCRβ-sequencing revealed a significant increase in TCRβ repertoire diversity and reduced clonality in both CP groups versus controls. Interestingly, we observed differences in Vβ-Jβ gene family usage between hereditary and idiopathic CP and a positive correlation of TCRβ rearrangements with disease severity scores. Immunophenotyping analyses in hereditary and idiopathic CP pancreata indicate differences in innate and adaptive immune responses, which highlights differences in immunopathogenic mechanism of disease among subtypes of CP. TCR repertoire analysis further suggests a role for specific T cell responses in hereditary versus idiopathic CP pathogenesis providing new insights into immune responses associated with human CP.

Authors

Bomi Lee, Julia Z. Adamska, Hong Namkoong, Melena D. Bellin, Joshua J. Wilhelm, Gregory L. Szot, David M. Louis, Mark M. Davis, Stephen Pandol, Aida Habtezion

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Abstract

Treating neuropathic pain is challenging and novel non-opioid based medicines are needed. Using unbiased receptomics, transcriptomic analyses, immunofluorescence and in situ hybridization, we found the expression of the orphan GPCR (oGPCR) Gpr160 and GPR160 increased in the rodent dorsal horn of the spinal cord (DH-SC) following traumatic nerve injury. Genetic and immunopharmacological approaches demonstrated that GPR160 inhibition in the spinal cord prevented and reversed neuropathic pain in male and female rodents without altering normal pain response. GPR160 inhibition in the spinal cord attenuated sensory processing in the thalamus, a key relay in the sensory discriminative pathways of pain. We also identified cocaine- and amphetamine-regulated transcript peptide (CARTp) as a GPR160 ligand. Inhibiting endogenous CARTp signaling in spinal cord attenuated neuropathic pain, whereas exogenous intrathecal (i.th.) CARTp evoked painful hypersensitivity through GPR160-dependent ERK and cAMP response element-binding protein (CREB). Our findings de-orphanize GPR160, identify it as a determinant of neuropathic pain and potential therapeutic target, and provide insights to its signaling pathways. CARTp is involved in many diseases including depression, reward and addiction, de-orphanization of GPR160 is a major step forward understanding the role of CARTp signaling in health and disease.

Authors

Gina LC Yosten, Caron M. Harada, Christopher J. Haddock, Luigino Antonio Giancotti, Grant R. Kolar, Ryan Patel, Chun Guo, Zhoumou Chen, Jinsong Zhang, Timothy M. Doyle, Anthony H. Dickenson, Willis K. Samson, Daniela Salvemini

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Abstract

Risk for childhood asthma is conferred by alleles within the 17q21 locus affecting ORMDL sphingolipid biosynthesis regulator 3 (ORMDL3) expression. ORMDL3 inhibits sphingolipid de novo synthesis. Although the effects of 17q21 genotypes on sphingolipid synthesis in human asthma remain unclear, both decreased sphingolipid synthesis and ORMDL3 overexpression are linked to airway hyperreactivity. To characterize the relationship of genetic asthma susceptibility with sphingolipid synthesis, we analyzed asthma-associated 17q21 genotypes (rs7216389, rs8076131, rs4065275, rs12603332, and rs8067378) in both children with asthma and those without asthma, quantified plasma and whole-blood sphingolipids, and assessed sphingolipid de novo synthesis in peripheral blood cells by measuring the incorporation of stable isotope–labeled serine (substrate) into sphinganine and sphinganine-1-phosphate. Whole-blood dihydroceramides and ceramides were decreased in subjects with the 17q21 asthma–risk alleles rs7216389 and rs8076131. Children with nonallergic asthma had lower dihydroceramides, ceramides, and sphingomyelins than did controls. Children with allergic asthma had higher dihydroceramides, ceramides, and sphingomyelins compared with children with nonallergic asthma. Additionally, de novo sphingolipid synthesis was lower in children with asthma compared with controls. These findings connect genetic 17q21 variations that are associated with asthma risk and higher ORMDL3 expression to lower sphingolipid synthesis in humans. Altered sphingolipid synthesis may therefore be a critical factor in asthma pathogenesis and may guide the development of future therapeutics.

Authors

Jennie G. Ono, Benjamin I. Kim, Yize Zhao, Paul J. Christos, Yohannes Tesfaigzi, Tilla S. Worgall, Stefan Worgall

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