Research Article
Abstract
Inflammatory bowel disease (IBD) is a chronic inflammatory disorder with rising incidence. Diseased tissues are heavily vascularized. Surprisingly, the pathogenic impact of the vasculature in IBD and the underlying regulatory mechanisms remain largely unknown. IFN-γ is a major cytokine in IBD pathogenesis, but in the context of the disease, it is almost exclusively its immune-modulatory and epithelial cell–directed functions that have been considered. Recent studies by our group demonstrated that IFN-γ also exerts potent effects on blood vessels. Based on these considerations, we analyzed the vessel-directed pathogenic functions of IFN-γ and found that it drives IBD pathogenesis through vascular barrier disruption. Specifically, we show that inhibition of the IFN-γ response in vessels by endothelial-specific knockout of IFN-γ receptor 2 ameliorates experimentally induced colitis in mice. IFN-γ acts pathogenic by causing a breakdown of the vascular barrier through disruption of the adherens junction protein VE-cadherin. Notably, intestinal vascular barrier dysfunction was also confirmed in human IBD patients, supporting the clinical relevance of our findings. Treatment with imatinib restored VE-cadherin/adherens junctions, inhibited vascular permeability, and significantly reduced colonic inflammation in experimental colitis. Our findings inaugurate the pathogenic impact of IFN-γ–mediated intestinal vessel activation in IBD and open new avenues for vascular-directed treatment of this disease.
Authors
Victoria Langer, Eugenia Vivi, Daniela Regensburger, Thomas H. Winkler, Maximilian J. Waldner, Timo Rath, Benjamin Schmid, Lisa Skottke, Somin Lee, Noo Li Jeon, Thomas Wohlfahrt, Viktoria Kramer, Philipp Tripal, Michael Schumann, Stephan Kersting, Claudia Handtrack, Carol I. Geppert, Karina Suchowski, Ralf H. Adams, Christoph Becker, Andreas Ramming, Elisabeth Naschberger, Nathalie Britzen-Laurent, Michael Stürzl
Abstract
We report on 2 patients with compound heterozygous mutations in forkhead box N1 (FOXN1), a transcription factor essential for thymic epithelial cell (TEC) differentiation. TECs are critical for T cell development. Both patients had a presentation consistent with T–/loB+NK+ SCID, with normal hair and nails, distinct from the classic nude/SCID phenotype in individuals with autosomal-recessive FOXN1 mutations. To understand the basis of this phenotype and the effects of the mutations on FOXN1, we generated mice using CRISPR-Cas9 technology to genocopy mutations in 1 of the patients. The mice with the Foxn1 compound heterozygous mutations had thymic hypoplasia, causing a T–B+NK+ SCID phenotype, whereas the hair and nails of these mice were normal. Characterization of the functional changes due to the Foxn1 mutations revealed a 5–amino acid segment at the end of the DNA-binding domain essential for the development of TECs but not keratinocytes. The transcriptional activity of this Foxn1 mutant was partly retained, indicating a region that specifies TEC functions. Analysis of an additional 9 FOXN1 mutations identified in multiple unrelated patients revealed distinct functional consequences contingent on the impact of the mutation on the DNA-binding and transactivation domains of FOXN1.
Authors
Qiumei Du, Larry K. Huynh, Fatma Coskun, Erika Molina, Matthew A. King, Prithvi Raj, Shaheen Khan, Igor Dozmorov, Christine M. Seroogy, Christian A. Wysocki, Grace T. Padron, Tyler R. Yates, M. Louise Markert, M. Teresa de la Morena, Nicolai S.C. van Oers
Abstract
Multiple sclerosis (MS) is a disabling disease of the CNS. Inflammatory features of MS include lymphocyte accumulations in the CNS and cerebrospinal fluid (CSF). The preclinical events leading to established MS are still enigmatic. Here we compared gene expression patterns of CSF cells from MS-discordant monozygotic twin pairs. Six “healthy” co-twins, who carry a maximal familial risk for developing MS, showed subclinical neuroinflammation (SCNI) with small MRI lesions. Four of these subjects had oligoclonal bands (OCBs). By single-cell RNA sequencing of 2752 CSF cells, we identified clonally expanded CD8+ T cells, plasmablasts, and, to a lesser extent, CD4+ T cells not only from MS patients but also from subjects with SCNI. In contrast to nonexpanded T cells, clonally expanded T cells showed characteristics of activated tissue-resident memory T (TRM) cells. The TRM-like phenotype was detectable already in cells from SCNI subjects but more pronounced in cells from patients with definite MS. Expanded plasmablast clones were detected only in MS and SCNI subjects with OCBs. Our data provide evidence for very early concomitant activation of 3 components of the adaptive immune system in MS, with a notable contribution of clonally expanded TRM-like CD8+ cells.
Authors
Eduardo Beltrán, Lisa Ann Gerdes, Julia Hansen, Andrea Flierl-Hecht, Stefan Krebs, Helmut Blum, Birgit Ertl-Wagner, Frederik Barkhof, Tania Kümpfel, Reinhard Hohlfeld, Klaus Dornmair
Abstract
To investigate the possibility that HIV-1 replication in lymph nodes sustains the reservoir during antiretroviral therapy (ART), we looked for evidence of viral replication in 5 donors after up to 13 years of viral suppression. We characterized proviral populations in lymph nodes and peripheral blood before and during ART, evaluated the levels of viral RNA expression in single lymph node and blood cells, and characterized the proviral integration sites in paired lymph node and blood samples. Proviruses with identical sequences, identical integration sites, and similar levels of RNA expression were found in lymph nodes and blood samples collected during ART, and no single sequence with significant divergence from the pretherapy population was detected in either blood or lymph nodes. These findings show that all detectable persistent HIV-1 infection is consistent with maintenance in lymph nodes by clonal proliferation of cells infected before ART and not by ongoing viral replication during ART.
Authors
William R. McManus, Michael J. Bale, Jonathan Spindler, Ann Wiegand, Andrew Musick, Sean C. Patro, Michele D. Sobolewski, Victoria K. Musick, Elizabeth M. Anderson, Joshua C. Cyktor, Elias K. Halvas, Wei Shao, Daria Wells, Xiaolin Wu, Brandon F. Keele, Jeffrey M. Milush, Rebecca Hoh, John W. Mellors, Stephen H. Hughes, Steven G. Deeks, John M. Coffin, Mary F. Kearney
Abstract
Cancer-associated fibroblasts (CAFs) are key actors in modulating the progression of many solid tumors, such as breast cancer (BC). Herein, we identify an integrin α11/PDGFRβ–positive CAF subset displaying tumor-promoting features in BC. In the preclinical MMTV-PyMT mouse model, integrin α11 deficiency led to a drastic reduction of tumor progression and metastasis. A clear association between integrin α11 and PDGFRβ was found at both transcriptional and histological levels in BC specimens. High stromal integrin α11/PDGFRβ expression was associated with high grades and poorer clinical outcome in human BC patients. Functional assays using 5 CAF subpopulations (1 murine, 4 human) revealed that integrin α11 promotes CAF invasion and CAF-induced tumor cell invasion upon PDGF-BB stimulation. Mechanistically, the proinvasive activity of integrin α11 relies on its ability to interact with PDGFRβ in a ligand-dependent manner and to promote its downstream JNK activation, leading to the production of tenascin C, a proinvasive matricellular protein. Pharmacological inhibition of PDGFRβ and JNK impaired tumor cell invasion induced by integrin α11+ CAFs. Collectively, our study uncovers an integrin α11+ subset of protumoral CAFs that exploits the PDGFRβ/JNK signaling axis to promote tumor invasiveness in BC.
Authors
Irina Primac, Erik Maquoi, Silvia Blacher, Ritva Heljasvaara, Jan Van Deun, Hilde Y.H. Smeland, Annalisa Canale, Thomas Louis, Linda Stuhr, Nor Eddine Sounni, Didier Cataldo, Taina Pihlajaniemi, Christel Pequeux, Olivier De Wever, Donald Gullberg, Agnès Noel
Abstract
The majority of patients with diabetic macular edema (DME), the most common cause of vision loss in working-age Americans, do not respond adequately to current therapies targeting VEGFA. Here, we show that expression of angiopoietin-like 4 (ANGPTL4), a HIF-1–regulated gene product, is increased in the eyes of diabetic mice and patients with DME. We observed that ANGPTL4 and VEGF act synergistically to destabilize the retinal vascular barrier. Interestingly, while ANGPTL4 modestly enhanced tyrosine phosphorylation of VEGF receptor 2, promotion of vascular permeability by ANGPTL4 was independent of this receptor. Instead, we found that ANGPTL4 binds directly to neuropilin 1 (NRP1) and NRP2 on endothelial cells (ECs), leading to rapid activation of the RhoA/ROCK signaling pathway and breakdown of EC-EC junctions. Treatment with a soluble fragment of NRP1 (sNRP1) prevented ANGPTL4 from binding to NRP1 and blocked ANGPTL4-induced activation of RhoA as well as EC permeability in vitro and retinal vascular leakage in diabetic animals in vivo. In addition, sNRP1 reduced the stimulation of EC permeability by aqueous fluid from patients with DME. Collectively, these data identify the ANGPTL4/NRP/RhoA pathway as a therapeutic target for the treatment of DME.
Authors
Akrit Sodhi, Tao Ma, Deepak Menon, Monika Deshpande, Kathleen Jee, Aumreetam Dinabandhu, Jordan Vancel, Daoyuan Lu, Silvia Montaner
Abstract
Membrane repair is essential to cell survival. In skeletal muscle, injury often associates with plasma membrane disruption. Additionally, muscular dystrophy is linked to mutations in genes that produce fragile membranes or reduce membrane repair. Methods to enhance repair and reduce susceptibility to injury could benefit muscle in both acute and chronic injury settings. Annexins are a family of membrane-associated Ca2+-binding proteins implicated in repair, and annexin A6 was previously identified as a genetic modifier of muscle injury and disease. Annexin A6 forms the repair cap over the site of membrane disruption. To elucidate how annexins facilitate repair, we visualized annexin cap formation during injury. We found that annexin cap size positively correlated with increasing Ca2+ concentrations. We also found that annexin overexpression promoted external blebs enriched in Ca2+ and correlated with a reduction of intracellular Ca2+ at the injury site. Annexin A6 overexpression reduced membrane injury, consistent with enhanced repair. Treatment with recombinant annexin A6 protected against acute muscle injury in vitro and in vivo. Moreover, administration of recombinant annexin A6 in a model of muscular dystrophy reduced serum creatinine kinase, a biomarker of disease. These data identify annexins as mediators of membrane-associated Ca2+ release during membrane repair and annexin A6 as a therapeutic target to enhance membrane repair capacity.
Authors
Alexis R. Demonbreun, Katherine S. Fallon, Claire C. Oosterbaan, Elena Bogdanovic, James L. Warner, Jordan J. Sell, Patrick G. Page, Mattia Quattrocelli, David Y. Barefield, Elizabeth M. McNally
Abstract
Essentially all Staphylococcus aureus (S. aureus) bacteria that gain access to the circulation are plucked out of the bloodstream by the intravascular macrophages of the liver — the Kupffer cells. It is also thought that these bacteria are disseminated via the bloodstream to other organs. Our data show that S. aureus inside Kupffer cells grew and escaped across the mesothelium into the peritoneal cavity and immediately infected GATA-binding factor 6–positive (GATA6+) peritoneal cavity macrophages. These macrophages provided a haven for S. aureus, thereby delaying the neutrophilic response in the peritoneum by 48 hours and allowing dissemination to various peritoneal and retroperitoneal organs including the kidneys. In mice deficient in GATA6+ peritoneal macrophages, neutrophils infiltrated more robustly and reduced S. aureus dissemination. Antibiotics administered i.v. did not prevent dissemination into the peritoneum or to the kidneys, whereas peritoneal administration of vancomycin (particularly liposomal vancomycin with optimized intracellular penetrance capacity) reduced kidney infection and mortality, even when administered 24 hours after infection. These data indicate that GATA6+ macrophages within the peritoneal cavity are a conduit of dissemination for i.v. S. aureus, and changing the route of antibiotic delivery could provide a more effective treatment for patients with peritonitis-associated bacterial sepsis.
Authors
Selina K. Jorch, Bas G.J. Surewaard, Mokarram Hossain, Moritz Peiseler, Carsten Deppermann, Jennifer Deng, Ania Bogoslowski, Fardau van der Wal, Abdelwahab Omri, Michael J. Hickey, Paul Kubes
Abstract
Serine-rich splicing factor 3 (SRSF3) plays a critical role in liver function and its loss promotes chronic liver damage and regeneration. As a consequence, genetic deletion of SRSF3 in hepatocytes caused progressive liver disease and ultimately led to hepatocellular carcinoma. Here we show that SRSF3 is decreased in human liver samples with nonalcoholic fatty liver disease (NAFLD), nonalcoholic steatohepatitis (NASH), or cirrhosis that was associated with alterations in RNA splicing of known SRSF3 target genes. Hepatic SRSF3 expression was similarly decreased and RNA splicing dysregulated in mouse models of NAFLD and NASH. We showed that palmitic acid–induced oxidative stress caused conjugation of the ubiquitin-like NEDD8 protein to SRSF3 and proteasome-mediated degradation. SRSF3 was selectively neddylated at lysine 11 and mutation of this residue (SRSF3-K11R) was sufficient to prevent both SRSF3 degradation and alterations in RNA splicing. Lastly, prevention of SRSF3 degradation in vivo partially protected mice from hepatic steatosis, fibrosis, and inflammation. These results highlight a neddylation-dependent mechanism regulating gene expression in the liver that is disrupted in early metabolic liver disease and may contribute to the progression to NASH, cirrhosis, and ultimately hepatocellular carcinoma.
Authors
Deepak Kumar, Manasi Das, Consuelo Sauceda, Lesley G. Ellies, Karina Kuo, Purva Parwal, Mehak Kaur, Lily Jih, Gautam K. Bandyopadhyay, Douglas Burton, Rohit Loomba, Olivia Osborn, Nicholas J.G. Webster
Abstract
The rate of disease progression in autosomal-dominant polycystic kidney disease (ADPKD) has high intrafamilial variability, suggesting that environmental factors may play a role. We hypothesized that a prevalent form of renal insult may accelerate cystic progression and investigated tubular crystal deposition. We report that calcium oxalate (CaOx) crystal deposition led to rapid tubule dilation, activation of PKD-associated signaling pathways, and hypertrophy in tubule segments along the affected nephrons. Blocking mTOR signaling blunted this response and inhibited efficient excretion of lodged crystals. This mechanism of “flushing out” crystals by purposefully dilating renal tubules has not to our knowledge been previously recognized. Challenging PKD rat models with CaOx crystal deposition or inducing calcium phosphate deposition by increasing dietary phosphorus intake led to increased cystogenesis and disease progression. In a cohort of patients with ADPKD, lower levels of urinary excretion of citrate, an endogenous inhibitor of calcium crystal formation, were correlated with increased disease severity. These results suggest that PKD progression may be accelerated by commonly occurring renal crystal deposition that could be therapeutically controlled by relatively simple measures.
Authors
Jacob A. Torres, Mina Rezaei, Caroline Broderick, Louis Lin, Xiaofang Wang, Bernd Hoppe, Benjamin D. Cowley Jr., Vincenzo Savica, Vicente E. Torres, Saeed Khan, Ross P. Holmes, Michal Mrug, Thomas Weimbs
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ISSN: 0021-9738 (print), 1558-8238 (online)