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.
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
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.
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
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.
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
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.
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
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.
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
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.
Hideyuki Takeshima, Tohru Niwa, Satoshi Yamashita, Takeji Takamura-Enya, Naoko Iida, Mika Wakabayashi, Sohachi Nanjo, Masanobu Abe, Toshiro Sugiyama, Young-Joon Kim, Toshikazu Ushijima
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.
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
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.
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
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.
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
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.
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
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