Gastroenterology
Abstract
As the interface between the gut microbiota and the mucosal immune system, there has been great interest in the maintenance of colonic epithelial integrity through mitochondrial oxidation of butyrate, a short-chain fatty acid produced by the gut microbiota. Herein, we showed that the intestinal epithelium can also oxidize long-chain fatty acids, and that luminally-delivered acylcarnitines in bile can be consumed via apical absorption by the intestinal epithelium resulting in mitochondrial oxidation. Finally, intestinal inflammation led to mitochondrial dysfunction in the apical domain of the surface epithelium that may reduce the consumption of fatty acids, contributing to higher concentrations of fecal acylcarnitines in murine Citrobacter rodentium-induced colitis and human inflammatory bowel disease. These results emphasized the importance of both the gut microbiota and the liver in the delivery of energy substrates for mitochondrial metabolism by the intestinal epithelium.
Authors
Sarah A. Smith, Sayaka A. Ogawa, Lillian Chau, Kelly A. Whelan, Kathryn E. Hamilton, Jie Chen, Lu Tan, Eric Z. Chen, Sue Keilbaugh, Franz Fogt, Meenakshi Bewtra, Jonathan Braun, Ramnik J. Xavier, Clary B. Clish, Barry Slaff, Aalim M. Weljie, Frederic D. Bushman, James D. Lewis, Hongzhe Li, Stephen R. Master, Michael J. Bennett, Hiroshi Nakagawa, Gary D. Wu
Abstract
A growing number of long non-coding RNAs (lncRNAs) have emerged as vital metabolic regulators. However, most human lncRNAs are non-conserved and highly tissue-specific, vastly limiting our ability to identify human lncRNA metabolic regulators (hLMRs). In this study, we establish a pipeline to identify putative hLMRs that are metabolically sensitive, disease-relevant, and population applicable. We first progressively processed multilevel human transcriptome data to select liver lncRNAs that exhibit highly dynamic expression in the general population, show differential expression in a nonalcoholic fatty liver disease (NAFLD) population, and response to dietary intervention in a small NAFLD cohort. We then experimentally demonstrated the responsiveness of selected hepatic lncRNAs to defined metabolic milieus in a liver-specific humanized mouse model. Furthermore, by extracting a concise list of protein-coding genes that are persistently correlated with lncRNAs in general and NAFLD populations, we predicted the specific function for each hLMR. Using gain- and loss-of-function approaches in humanized mice as well as ectopic expression in conventional mice, we validated the regulatory role of one non-conserved hLMR in cholesterol metabolism by coordinating with an RNA-binding protein, PTBP1, to modulate the transcription of cholesterol synthesis genes. Our work overcome the heterogeneity intrinsic to human data to enable the efficient identification and functional definition of disease-relevant human lncRNAs in metabolic homeostasis.
Authors
Xiangbo Ruan, Ping Li, Yonghe Ma, Chengfei Jiang, Yi Chen, Yu Shi, Nikhil Gupta, Fayaz Seifuddin, Mehdi Pirooznia, Yasuyuki Ohnishi, Nao Yoneda, Megumi Nishiwaki, Gabrijela Dumbovic, John L. Rinn, Yuichiro Higuchi, Kenji Kawai, Hiroshi Suemizu, Haiming Cao
Abstract
Protein tyrosine phosphatase non-receptor type 2 (PTPN2) recently emerged as a promising cancer immunotherapy target. We set to investigate the functional role of PTPN2 in the pathogenesis of human colorectal carcinoma (CRC) as its role in immune-silent solid tumors is poorly understood. We demonstrate that in human CRC, increased PTPN2 expression and activity correlated with disease progression and decreased immune responses in tumor tissues. Particularly, stage II and III tumors displayed enhanced PTPN2 protein expression in tumor-infiltrating T-cells and increased PTPN2 levels negatively correlated with PD1, CTLA4, STAT1 and granzyme A. In vivo, T-cell and dendritic cell-specific PTPN2 deletion reduced tumor burden in several CRC models by promoting CD44+ effector/memory T-cells, as well as CD8+ T-cell infiltration and cytotoxicity into the tumor. In direct relevance to CRC treatment, T-cell-specific PTPN2 deletion potentiated anti-PD-1 efficacy and induced anti-tumor memory formation upon tumor re-challenge in vivo. Our data suggest a role for PTPN2 in suppressing anti-tumor immunity and promoting tumor development in CRC patients. Our in vivo results uncover PTPN2 as a key player in controlling immunogenicity of CRC, with the strong potential to be exploited to promote cancer immunotherapy.
Authors
Egle Katkeviciute, Larissa Hering, Ana Montalban-Arques, Philipp Busenhart, Marlene Schwarzfischer, Roberto Manzini, Javier Conde, Kirstin Atrott, Silvia Lang, Gerhard Rogler, Elisabeth Naschberger, Vera S. Schellerer, Michael Stürzl, Andreas Rickenbacher, Matthias Turina, Achim Weber, Sebastian Leibl, Gabriel E. Leventhal, Mitchell Levesque, Onur Boyman, Michael Scharl, Marianne R. Spalinger
Abstract
Interstitial cells of Cajal (ICCs) are pacemaker cells in the intestine, and their function can be compromised due to loss of C-KIT expression. Macrophage activation has been identified in intestine affected by Hirschsprung disease associated enterocolitis (HAEC). In this study, we examined proinflammatory macrophage activation and explored the mechanisms by which this down-regulates C-KIT expression in ICCs in colon affected by HAEC. We found that macrophage activation and TNF-α production were dramatically increased in the proximal dilated colon of HAEC patients and 3-week old Ednrb-/- mice. Moreover, ICCs lost their C-KIT+ phenotype in the dilated colon, resulting in damaged pacemaker function and intestinal dysmotility. However, macrophage depletion or TNF-α neutralization led to recovery of ICC phenotype and restored their pacemaker function. In isolated ICCs, TNF-α-mediated phosphorylated-P65 induced over-expression of miR221, resulting in suppression of C-KIT expression and pacemaker currents. We also identified a TNF-α-NF-κB-miR221 pathway which downregulated C-KIT expression in ICCs in the colon affected by HAEC. These findings suggest the important roles of proinflammatory macrophage activation in a phenotypic switch of ICCs, representing a promising therapeutic target for HAEC.
Authors
Xuyong Chen, Xinyao Meng, Hongyi Zhang, Chenzhao Feng, Bin Wang, Ning Li, Khalid Mohamoud Abdullahi, Xiaojuan Wu, Jixin Yang, Zhi Li, Chunlei Jiao, Jia Wei, Xiaofeng Xiong, Kang Fu, Lei Yu, Gail Besner, Jiexiong Feng
Abstract
Arginase 1 (Arg1), which converts L-arginine into ornithine and urea, exerts pleiotropic immunoregulatory effects. However, the function of Arg1 in inflammatory bowel disease (IBD) remains poorly characterized. Here, we found that Arg1 expression correlated with the degree of inflammation in intestinal tissues from IBD patients. In mice, Arg1 was upregulated in an IL-4-/IL-13- and intestinal microbiota-dependent manner. Tie2-Cre+/-Arg1fl/fl mice lacking Arg1 in hematopoietic and endothelial cells recovered faster from experimental colitis than Arg1-expressing littermates. This correlated with decreased vessel density, compositional changes in intestinal microbiota, diminished infiltration by myeloid cells and an accumulation of intraluminal polyamines that promote epithelial healing. The pro-resolving effect of Arg1-deletion was reduced by an L-arginine-free diet, but rescued by simultaneous deletion of other L-arginine-metabolizing enzymes such as Arg2 or Nos2, demonstrating that protection from colitis requires L-arginine. Fecal microbiota transfers from Tie2-Cre+/-Arg1fl/fl mice into wild-type recipients ameliorated intestinal inflammation while transfers from wild-type littermates into Arg1-deficient mice prevented an advanced recovery from colitis. Thus, an increased availability of L-arginine as well as altered intestinal microbiota and metabolic products account for the accelerated resolution from colitis in the absence of Arg1. Consequently, the metabolism of L-arginine may serve as target for clinical intervention in IBD patients.
Authors
Julia Baier, Maximilian Gänsbauer, Claudia Giessler, Harald Arnold, Mercedes Muske, Ulrike Schleicher, Soeren Lukassen, Arif B. Ekici, Manfred Rauh, Christoph Daniel, Arndt Hartmann, Benjamin Schmid, Philipp Tripal, Katja Dettmer, Peter J. Oefner, Raja Atreya, Stefan Wirtz, Christian Bogdan, Jochen Mattner
Abstract
Immune checkpoint blockade (ICB) has revolutionized cancer therapeutics. Desmoplastic malignancies such as cholangiocarcinoma (CCA) have an abundant tumor immune microenvironment (TIME). However, to date ICB monotherapy in such malignancies has been ineffective. Herein, we identify that tumor-associated macrophages (TAMs) are the primary source of PD-L1 in human and murine CCA. In a murine model of CCA, recruited PD-L1+ TAMs facilitate CCA progression. However, TAM blockade failed to decrease tumor progression due to a compensatory emergence of granulocytic-myeloid-derived suppressor cells (G-MDSCs) that mediated immune escape by impairing T-cell response. Single-cell RNA sequencing (scRNA-seq) of murine tumor G-MDSCs highlighted a novel ApoE G-MDSC subset enriched with TAM blockade; further analysis of a human scRNA-seq dataset demonstrated the presence of a similar G-MDSC subset in human CCA. Finally, dual inhibition of TAMs and G-MDSCs potentiated ICB. In summary, our findings highlight the therapeutic potential of coupling ICB with immunotherapies targeting immunosuppressive myeloid cells in CCA.
Authors
Emilien Loeuillard, Jingchun Yang, EeeLN Buckarma, Juan Wang, Yuanhang Liu, Caitlin B. Conboy, Kevin D. Pavelko, Ying Li, Daniel O'Brien, Chen Wang, Rondell P. Graham, Rory L. Smoot, Haidong Dong, Sumera Rizvi
Abstract
Gasdermin D (GSDMD) induces pyroptosis via the pore-forming activity of its N-terminal domain, cleaved by activated caspases associated with the release of IL-1β. Here, we report a nonpyroptotic role of full-length GSDMD in guiding the release of IL-1β–containing small extracellular vesicles (sEVs) from intestinal epithelial cells (IECs). In response to caspase-8 inflammasome activation, GSDMD, chaperoned by Cdc37/Hsp90, recruits the E3 ligase, NEDD4, to catalyze polyubiquitination of pro–IL-1β, serving as a signal for cargo loading into secretory vesicles. GSDMD and IL-1β colocalize with the exosome markers CD63 and ALIX intracellularly, and GSDMD and NEDD4 are required for release of CD63+ sEVs containing IL-1β, GSDMD, NEDD4, and caspase-8. Importantly, increased expression of epithelial-derived GSDMD is observed both in patients with inflammatory bowel disease (IBD) and those with experimental colitis. While GSDMD-dependent release of IL-1β–containing sEVs is detected in cultured colonic explants from colitic mice, GSDMD deficiency substantially attenuates disease severity, implicating GSDMD-mediated release of IL-1β sEVs in the pathogenesis of intestinal inflammation, such as that observed in IBD.
Authors
Katarzyna Bulek, Junjie Zhao, Yun Liao, Nitish Rana, Daniele Corridoni, Agne Antanaviciute, Xing Chen, Han Wang, Wen Qian, William A. Miller-Little, Shadi Swaidani, Fangqiang Tang, Belinda B. Willard, Keith McCrae, Zizhen Kang, George R. Dubyak, Fabio Cominelli, Alison Simmons, Theresa T. Pizarro, Xiaoxia Li
Abstract
Chronic infections can lead to carcinogenesis through inflammation-related mechanisms. Chronic infection of the human gastric mucosa with Helicobacter pylori is a well-known risk factor for gastric cancer. However, the mechanisms underlying H. pylori–induced gastric carcinogenesis are incompletely defined. We aimed to screen and clarify the functions of long noncoding RNAs (lncRNAs) that are differentially expressed in H. pylori–related gastric cancer. We found that lncRNA SNHG17 was upregulated by H. pylori infection and markedly increased the levels of double-strand breaks (DSBs). SNHG17 overexpression correlated with poor overall survival in patients with gastric cancer. The recruitment of NONO by overabundant nuclear SNHG17, along with the role of cytoplasmic SNHG17 as a decoy for miR-3909, which regulates Rad51 expression, shifted the DSB repair balance from homologous recombination toward nonhomologous end joining. Notably, during chronic H. pylori infection, SNHG17 knockdown inhibited chromosomal aberrations. Our findings suggest that spatially independent deregulation of the SNHG17/NONO and SNHG17/miR-3909/RING1/Rad51 pathways upon H. pylori infection promotes tumorigenesis in gastric cancer by altering the DNA repair system, which is critical for the maintenance of genomic stability. Upregulation of SNHG17 by H. pylori infection might be an undefined link between cancer and inflammation.
Authors
Taotao Han, Xiaohui Jing, Jiayu Bao, Lianmei Zhao, Aidong Zhang, Renling Miao, Hui Guo, Baoguo Zhou, Shang Zhang, Jiazeng Sun, Juan Shi
Abstract
The tight junction protein claudin-2 is upregulated in disease. Although many studies have linked intestinal barrier loss to local and systemic disease, these have relied on macromolecular probes. In vitro analyses show however that these probes cannot be accommodated by size- and charge-selective claudin-2 channels. We sought to define the impact of claudin-2 channels on disease. Transgenic claudin-2 overexpression or IL-13-induced claudin-2 upregulation increased intestinal small cation permeability in vivo. IL-13 did not however affect permeability in claudin-2-knockout mice. Claudin-2 is therefore necessary and sufficient to effect size- and charge-selective permeability increases in vivo. In chronic disease, T-cell transfer colitis severity was augmented or diminished in claudin-2 transgenic or knockout mice, respectively. We translated in vitro data suggesting that casein kinase-2 (CK2) inhibition blocks claudin-2 channel function and found that CK2 inhibition prevented IL-13-induced, claudin-2-mediated permeability increases in vivo. In chronic immune-mediated colitis, CK2 inhibition attenuated progression in claudin-2-sufficient, but not claudin-2-knockout, mice, i.e., the effect was claudin-2-dependent. Paracellular flux mediated by claudin-2 channels can therefore promote immune-mediated colitis progression. Although the mechanisms by which claudin-2 channels intensify disease remain to be defined, these data suggest that claudin-2 may be an accessible target in immune-mediated disorders, including inflammatory bowel disease.
Authors
Preeti Raju, Nitesh Shashikanth, Pei-Yun Tsai, Pawin Pongkorpsakol, Sandra Chanez-Parades, Peter R. Steinhagen, Wei-Ting Kuo, Gurminder Singh, Sachiko Tsukita, Jerrold R. Turner
Abstract
Enteric neuronal degeneration, as seen in inflammatory bowel disease, obesity, and diabetes, can lead to gastrointestinal dysmotility. Pyroptosis is a novel form of programmed cell death but little is known about its role in enteric neuronal degeneration. We observed higher levels of cleaved caspase-1, a marker of pyroptosis, in myenteric ganglia of overweight and obese human subjects compared with normal-weight subjects. Western diet–fed (WD-fed) mice exhibited increased myenteric neuronal pyroptosis, delayed colonic transit, and impaired electric field stimulation–induced colonic relaxation responses. WD increased TLR4 expression and cleaved caspase-1 in myenteric nitrergic neurons. Overactivation of nitrergic neuronal NF-κB signaling resulted in increased pyroptosis and delayed colonic motility. In caspase-11–deficient mice, WD did not induce nitrergic myenteric neuronal pyroptosis and colonic dysmotility. To understand the contributions of saturated fatty acids and bacterial products to the steps leading to enteric neurodegeneration, we performed in vitro experiments using mouse enteric neurons. Palmitate and lipopolysaccharide (LPS) increased nitrergic, but not cholinergic, enteric neuronal pyroptosis. LPS gained entry to the cytosol in the presence of palmitate, activating caspase-11 and gasdermin D, leading to pyroptosis. These results support a role of the caspase-11–mediated pyroptotic pathway in WD-induced myenteric nitrergic neuronal degeneration and colonic dysmotility, providing important therapeutic targets for enteric neuropathy.
Authors
Lan Ye, Ge Li, Anna Goebel, Abhinav V. Raju, Feng Kong, Yanfei Lv, Kailin Li, Yuanjun Zhu, Shreya Raja, Peijian He, Fang Li, Simon Musyoka Mwangi, Wenhui Hu, Shanthi Srinivasan
Copyright © 2020 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)