Research Article
Abstract
Crohn’s disease (CD) is marked by recurring intestinal inflammation and tissue injury, often resulting in fibrostenosis and bowel obstruction, necessitating surgical intervention with high recurrence rates. To elucidate the mechanisms underlying fibrostenosis in CD, we analyzed the transcriptome of cells isolated from the transmural ileum of patients with CD, including a trio of lesions from each patient: non-affected, inflamed, and stenotic ileum samples, and compared them with samples from patients without CD. Our computational analysis revealed that profibrotic signals from a subset of monocyte-derived cells expressing CD150 induced a disease-specific fibroblast population, resulting in chronic inflammation and tissue fibrosis. The transcription factor TWIST1 was identified as a key modulator of fibroblast activation and extracellular matrix (ECM) deposition. Genetic and pharmacological inhibition of TWIST1 prevents fibroblast activation, reducing ECM production and collagen deposition. Our findings suggest that the myeloid-stromal axis may offer a promising therapeutic target to prevent fibrostenosis in CD.
Authors
Bo-Jun Ke, Saeed Abdurahiman, Francesca Biscu, Gaia Zanella, Gabriele Dragoni, Sneha Santhosh, Veronica De Simone, Anissa Zouzaf, Lies van Baarle, Michelle Stakenborg, Veronika Bosáková, Yentl Van Rymenant, Emile Verhulst, Sare Verstockt, Elliott Klein, Gabriele Bislenghi, Albert Wolthuis, Jan Frič, Christine Breynaert, Andre D’Hoore, Pieter Van der Veken, Ingrid De Meester, Sara Lovisa, Lukas J.A.C. Hawinkels, Bram Verstockt, Gert De Hertogh, Séverine Vermeire, Gianluca Matteoli
Abstract
The β-secretase β-site APP cleaving enzyme (BACE1) is a central drug target for Alzheimer’s disease. Clinically tested, BACE1-directed inhibitors also block the homologous protease BACE2. Yet little is known about physiological BACE2 substrates and functions in vivo. Here, we identify BACE2 as the protease shedding the lymphangiogenic vascular endothelial growth factor receptor 3 (VEGFR3). Inactivation of BACE2, but not BACE1, inhibited shedding of VEGFR3 from primary human lymphatic endothelial cells (LECs) and reduced release of the shed, soluble VEGFR3 (sVEGFR3) ectodomain into the blood of mice, nonhuman primates, and humans. Functionally, BACE2 inactivation increased full-length VEGFR3 and enhanced VEGFR3 signaling in LECs and also in vivo in zebrafish, where enhanced migration of LECs was observed. Thus, this study identifies BACE2 as a modulator of lymphangiogenic VEGFR3 signaling and demonstrates the utility of sVEGFR3 as a pharmacodynamic plasma marker for BACE2 activity in vivo, a prerequisite for developing BACE1-selective inhibitors for safer prevention of Alzheimer’s disease.
Authors
Andree Schmidt, Brian Hrupka, Frauke van Bebber, Sanjay Sunil Kumar, Xiao Feng, Sarah K. Tschirner, Marlene Aßfalg, Stephan A. Müller, Laura Sophie Hilger, Laura I. Hofmann, Martina Pigoni, Georg Jocher, Iryna Voytyuk, Emily L. Self, Mana Ito, Kana Hyakkoku, Akimasa Yoshimura, Naotaka Horiguchi, Regina Feederle, Bart De Strooper, Stefan Schulte-Merker, Eckhard Lammert, Dieder Moechars, Bettina Schmid, Stefan F. Lichtenthaler
Abstract
Preventing the onset of autoimmune type 1 diabetes (T1D) is feasible through pharmacological interventions that target molecular stress–responsive mechanisms. Cellular stresses, such as nutrient deficiency, viral infection, or unfolded proteins, trigger the integrated stress response (ISR), which curtails protein synthesis by phosphorylating eukaryotic translation initiation factor-2α (eIF2α). In T1D, maladaptive unfolded protein response (UPR) in insulin-producing β cells renders these cells susceptible to autoimmunity. We found that inhibition of the eIF2α kinase PKR-like ER kinase (PERK), a common component of the UPR and ISR, reversed the mRNA translation block in stressed human islets and delayed the onset of diabetes, reduced islet inflammation, and preserved β cell mass in T1D-susceptible mice. Single-cell RNA-Seq of islets from PERK-inhibited mice showed reductions in the UPR and PERK signaling pathways and alterations in antigen-processing and presentation pathways in β cells. Spatial proteomics of islets from these mice showed an increase in the immune checkpoint protein programmed death-ligand 1 (PD-L1) in β cells. Golgi membrane protein 1, whose levels increased following PERK inhibition in human islets and EndoC-βH1 human β cells, interacted with and stabilized PD-L1. Collectively, our studies show that PERK activity enhances β cell immunogenicity and that inhibition of PERK may offer a strategy for preventing or delaying the development of T1D.
Authors
Charanya Muralidharan, Fei Huang, Jacob R. Enriquez, Jiayi E. Wang, Jennifer B. Nelson, Titli Nargis, Sarah C. May, Advaita Chakraborty, Kayla T. Figatner, Svetlana Navitskaya, Cara M. Anderson, Veronica Calvo, David Surguladze, Mark J. Mulvihill, Xiaoyan Yi, Soumyadeep Sarkar, Scott A. Oakes, Bobbie-Jo M. Webb-Robertson, Emily K. Sims, Kirk A. Staschke, Decio L. Eizirik, Ernesto S. Nakayasu, Michael E. Stokes, Sarah A. Tersey, Raghavendra G. Mirmira
Abstract
We report that diazepam binding inhibitor (DBI) is a glial messenger mediating crosstalk between satellite glial cells (SGCs) and sensory neurons in the dorsal root ganglion (DRG). DBI is highly expressed in SGCs of mice, rats, and humans, but not in sensory neurons or most other DRG-resident cells. Knockdown of DBI results in a robust mechanical hypersensitivity without major effects on other sensory modalities. In vivo overexpression of DBI in SGCs reduces sensitivity to mechanical stimulation and alleviates mechanical allodynia in neuropathic and inflammatory pain models. We further show that DBI acts as an unconventional agonist and positive allosteric modulator at the neuronal GABAA receptors, particularly strongly affecting those with a high-affinity benzodiazepine binding site. Such receptors are selectively expressed by a subpopulation of mechanosensitive DRG neurons, and these are also more enwrapped with DBI-expressing glia, as compared with other DRG neurons, suggesting a mechanism for a specific effect of DBI on mechanosensation. These findings identified a communication mechanism between peripheral neurons and SGCs. This communication modulates pain signaling and can be targeted therapeutically.
Authors
Xinmeng Li, Arthur Silveira Prudente, Vincenzo Prato, Xianchuan Guo, Han Hao, Frederick Jones, Sofia Figoli, Pierce Mullen, Yujin Wang, Raquel Tonello, Sang Hoon Lee, Shihab Shah, Benito Maffei, Temugin Berta, Xiaona Du, Nikita Gamper
Abstract
Cell cycle regulation is largely abnormal in cancers. Molecular understanding and therapeutic targeting of the aberrant cell cycle are essential. Here, we identified that an underappreciated serine/threonine kinase, cyclin-dependent kinase–like 3 (CDKL3), crucially drives rapid cell cycle progression and cell growth in cancers. With regard to mechanism, CDKL3 localizes in the nucleus and associates with specific cyclin to directly phosphorylate retinoblastoma (Rb) for quiescence exit. In parallel, CDKL3 prevents the ubiquitin-proteasomal degradation of cyclin-dependent kinase 4 (CDK4) by direct phosphorylation on T172 to sustain G1 phase advancement. The crucial function of CDKL3 in cancers was demonstrated both in vitro and in vivo. We also designed, synthesized, and characterized a first-in-class CDKL3-specific inhibitor, HZ1. HZ1 exhibits greater potency than CDK4/6 inhibitor in pan-cancer treatment by causing cell cycle arrest and overcomes acquired resistance to CDK4/6 inhibitor. In particular, CDKL3 has significant clinical relevance in colon cancer, and the effectiveness of HZ1 was demonstrated by murine and patient-derived cancer models. Collectively, this work presents an integrated paradigm of cancer cell cycle regulation and suggests CDKL3 targeting as a feasible approach in cancer treatment.
Authors
Haijiao Zhang, Jiahui Lin, Shaoqin Zheng, Lanjing Ma, Zhongqiu Pang, Hongyi Yin, Chengcheng Meng, Yinuo Wang, Qing Han, Xi Zhang, Zexu Li, Liu Cao, Lijun Liu, Teng Fei, Daming Gao, Liang Yang, Xueqiang Peng, Chen Ding, Shixue Wang, Ren Sheng
Abstract
Myostatin (MSTN) has long been recognized as a critical regulator of muscle mass. Recently, there has been increasing interest in its role in metabolism. In our study, we specifically knocked out MSTN in brown adipose tissue (BAT) from mice (MSTNΔUCP1) and found that the mice gained more weight than did controls when fed a high-fat diet, with progressive hepatosteatosis and impaired skeletal muscle activity. RNA-Seq analysis indicated signatures of mitochondrial dysfunction and inflammation in the MSTN-ablated BAT. Further studies demonstrated that Kruppel-like factor 4 (KLF4) was responsible for the metabolic phenotypes observed, whereas fibroblast growth factor 21 (FGF21) contributed to the microenvironment communication between adipocytes and macrophages induced by the loss of MSTN. Moreover, the MSTN/SMAD2/3-p38 signaling pathway mediated the expression of KLF4 and FGF21 in adipocytes. In summary, our findings suggest that brown adipocyte–derived MSTN regulated BAT thermogenesis via autocrine and paracrine effects on adipocytes or macrophages, ultimately regulating systemic energy homeostasis.
Authors
Hui Wang, Shanshan Guo, Huanqing Gao, Jiyang Ding, Hongyun Li, Xingyu Kong, Shuang Zhang, Muyang He, Yonghao Feng, Wei Wu, Kexin Xu, Yuxuan Chen, Hanyin Zhang, Tiemin Liu, Xingxing Kong
Abstract
Leukemia relapse is a major cause of death after allogeneic hematopoietic cell transplantation (allo-HCT). We tested the potential of targeting T cell (Tc) immunoglobulin and mucin-containing molecule 3 (TIM-3) for improving graft-versus-leukemia (GVL) effects. We observed differential expression of TIM-3 ligands when hematopoietic stem cells overexpressed certain oncogenic-driver mutations. Anti–TIM-3 Ab treatment improved survival of mice bearing leukemia with oncogene-induced TIM-3 ligand expression. Conversely, leukemia cells with low ligand expression were anti–TIM-3 treatment resistant. In vitro, TIM-3 blockade or genetic deletion in CD8+ Tc enhanced Tc activation, proliferation, and IFN-γ production while enhancing GVL effects, preventing Tc exhaustion, and improving Tc cytotoxicity and glycolysis in vivo. Conversely, TIM-3 deletion in myeloid cells did not affect allogeneic Tc proliferation and activation in vitro, suggesting that anti–TIM-3 treatment–mediated GVL effects are Tc induced. In contrast to anti–programmed cell death protein 1 (anti–PD-1) and anti–cytotoxic T lymphocyte–associated protein 4 (anti–CTLA-4) treatment, anti–TIM-3-treatment did not enhance acute graft-versus-host disease (aGVHD). TIM-3 and its ligands were frequently expressed in acute myeloid leukemia (AML) cells of patients with post–allo-HCT relapse. We decipher the connections between oncogenic mutations found in AML and TIM-3 ligand expression and identify anti–TIM-3 treatment as a strategy for enhancing GVL effects via metabolic and transcriptional Tc reprogramming without exacerbation of aGVHD. Our findings support clinical testing of anti–TIM-3 Ab in patients with AML relapse after allo-HCT.
Authors
Nana Talvard-Balland, Lukas M. Braun, Karen O. Dixon, Melissa Zwick, Helena Engel, Alina Hartmann, Sandra Duquesne, Livius Penter, Geoffroy Andrieux, Lukas Rindlisbacher, Andrea Acerbis, Jule Ehmann, Christoph Köllerer, Michela Ansuinelli, Andres Rettig, Kevin Moschallski, Petya Apostolova, Tilman Brummer, Anna L. Illert, Markus A. Schramm, Yurong Cheng, Anna Köttgen, Justus Duyster, Hans D. Menssen, Jerome Ritz, Bruce R. Blazar, Melanie Boerries, Annette Schmitt-Gräff, Nurefsan Sariipek, Peter Van Galen, Joerg M. Buescher, Nina Cabezas-Wallscheid, Heike L. Pahl, Erika L. Pearce, Robert J. Soiffer, Catherine J. Wu, Luca Vago, Burkhard Becher, Natalie Köhler, Tobias Wertheimer, Vijay K. Kuchroo, Robert Zeiser
Abstract
IgG4-related disease (IgG4-RD) is a systemic immune-mediated fibroinflammatory disease whose pathomechanisms remain poorly understood. Here, we identified gene variants in familial IgG4-RD and determined their functional consequences. All 3 affected members of the family shared variants of the transcription factor IKAROS, encoded by IKZF1, and the E3 ubiquitin ligase UBR4. The IKAROS variant increased binding to the FYN promoter, resulting in higher transcription of FYN in T cells. The UBR4 variant prevented the lysosomal degradation of the phosphatase CD45. In the presence of elevated FYN, CD45 functioned as a positive regulatory loop, lowering the threshold for T cell activation. Consequently, T cells from the affected family members were hyperresponsive to stimulation. When transduced with a low-avidity, autoreactive T cell receptor, their T cells responded to the autoantigenic peptide. In parallel, high expression of FYN in T cells biased their differentiation toward Th2 polarization by stabilizing the transcription factor JunB. This bias was consistent with the frequent atopic manifestations in patients with IgG4-RD, including the affected family members in the present study. Building on the functional consequences of these 2 variants, we propose a disease model that is not only instructive for IgG4-RD but also for atopic diseases and autoimmune diseases associated with an IKZF1 risk haplotype.
Authors
Qingxiang Liu, Yanyan Zheng, Ines Sturmlechner, Abhinav Jain, Maryam Own, Qiankun Yang, Huimin Zhang, Filippo Pinto e Vairo, Karen Cerosaletti, Jane H. Buckner, Kenneth J. Warrington, Matthew J. Koster, Cornelia M. Weyand, Jörg J. Goronzy
Abstract
Tolerance of mouse kidney allografts arises in grafts that develop regulatory tertiary lymphoid organs (rTLOs). Single-cell RNA-seq (scRNA-seq) data and adoptive transfer of alloreactive T cells after transplantation showed that cytotoxic CD8+ T cells are reprogrammed within the accepted graft to an exhausted/regulatory-like phenotype mediated by IFN-γ. Establishment of rTLOs was required because adoptive transfer of alloreactive T cells prior to transplantation results in kidney allograft rejection. Despite the presence of intragraft CD8+ cells with a regulatory phenotype, they were not essential for the induction and maintenance of kidney allograft tolerance since renal allotransplantation into CD8-KO recipients resulted in acceptance and not rejection. Analysis of scRNA-seq data from allograft kidneys and malignant tumors identified similar regulatory-like cell types within the T cell clusters and trajectory analysis showed that cytotoxic CD8+ T cells are reprogrammed into an exhausted/regulatory-like phenotype intratumorally. Induction of cytotoxic CD8+ T cell dysfunction of infiltrating cells appears to be a beneficial mechanistic pathway that protects the kidney allotransplant from rejection through a process we call “defensive tolerance.” This pathway has implications for our understanding of allotransplant tolerance and tumor resistance to host immunity.
Authors
Takahiro Yokose, Edward S. Szuter, Ivy Rosales, Michael T. Guinn, Andrew S. Liss, Taisuke Baba, David A. Ruddy, Michelle Piquet, Jamil Azzi, A. Benedict Cosimi, Paul S. Russell, Joren C. Madsen, Robert B. Colvin, Alessandro Alessandrini
Abstract
Reelin (RELN) is a secreted glycoprotein essential for cerebral cortex development. In humans, recessive RELN variants cause cortical and cerebellar malformations, while heterozygous variants were associated with epilepsy, autism, and mild cortical abnormalities. However, the functional effects of RELN variants remain unknown. We identified inherited and de novo RELN missense variants in heterozygous patients with neuronal migration disorders (NMDs) as diverse as pachygyria and polymicrogyria. We investigated in culture and in the developing mouse cerebral cortex how different variants impacted RELN function. Polymicrogyria-associated variants behaved as gain-of-function, showing an enhanced ability to induce neuronal aggregation, while those linked to pachygyria behaved as loss-of-function, leading to defective neuronal aggregation/migration. The pachygyria-associated de novo heterozygous RELN variants acted as dominant-negative by preventing WT RELN secretion in culture, animal models, and patients, thereby causing dominant NMDs. We demonstrated how mutant RELN proteins in vitro and in vivo predict cortical malformation phenotypes, providing valuable insights into the pathogenesis of such disorders.
Authors
Martina Riva, Sofia Ferreira, Kotaro Hayashi, Yoann Saillour, Vera P. Medvedeva, Takao Honda, Kanehiro Hayashi, Claire Altersitz, Shahad Albadri, Marion Rosello, Julie Dang, Malo Serafini, Frédéric Causeret, Olivia J. Henry, Charles-Joris Roux, Céline Bellesme, Elena Freri, Dragana Josifova, Elena Parrini, Renzo Guerrini, Filippo Del Bene, Kazunori Nakajima, Nadia Bahi-Buisson, Alessandra Pierani
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ISSN: 0021-9738 (print), 1558-8238 (online)