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Abstract
Targeted therapies and immunotherapy have shown promise in patients with non-small cell lung cancer (NSCLC). However, the majority of patients fail or become resistant to treatment, emphasizing the need for novel treatments. In this study, we confirm the prognostic value of AXL levels in NSCLC and demonstrate potent anti-tumor activity of the AXL-targeting antibody-drug conjugate enapotamab vedotin across different NSCLC subtypes in a mouse clinical trial of human NSCLC. Tumor regression or stasis was observed in 17/61 (28%) of the PDX models, and was associated with AXL mRNA expression levels. Significant single agent activity of enapotamab vedotin was validated in vivo in 9 of 10 AXL-expressing NSCLC xenograft models. In a panel of EGFR-mutant NSCLC cell lines rendered resistant to EGFR inhibitors (EGFRi) in vitro, we observed de novo or increased AXL protein expression concomitant with enapotamab vedotin-mediated cytotoxicity. Enapotamab vedotin also showed anti-tumor activity in vivo in 3 EGFR-mutant, EGFRi-resistant PDX models, including an osimertinib-resistant NSCLC PDX model. In summary, enapotamab vedotin has promising therapeutic potential in NSCLC. The safety and preliminary efficacy of enapotamab vedotin are currently being evaluated in the clinic across multiple solid tumor types, including NSCLC.
Authors
Louise A. Koopman, Mikkel G. Terp, Gijs G. Zom, Maarten L. Janmaat, Kirstine Jacobsen, Elke Gresnigt - Van den Heuvel, Marcel Brandhorst, Ulf Forssmann, Frederik M. de Bree, Nora Pencheva, Andreas Lingnau, Maria A. Zipeto, Paul W.H.I. Parren, Esther C.W. Breij, Henrik J. Ditzel
Abstract
Background. The presence of an early repolarization pattern (ERP) on the surface electrocardiogram (ECG) is associated with risk of ventricular fibrillation and sudden cardiac death. Family studies have shown that ERP is a highly heritable trait but molecular genetic determinants are unknown. Methods. To identify genetic susceptibility loci for ERP, we performed a GWAS and meta-analysis in 2,181 cases and 23,641 controls of European ancestry. Results. We identified a genome-wide significant (p<5E-8) locus in the KCND3 (potassium voltage gated channel subfamily D member 3) gene that was successfully replicated in additional 1,124 cases and 12,510 controls. A subsequent joint meta-analysis of the discovery and replication cohorts identified rs1545300 as the lead SNP at the KCND3 locus (OR 0.82 per minor T allele, p=7.7E-12), but did not reveal additional loci. Co-localization analyses indicate causal effects of KCND3 gene expression levels on ERP in both cardiac left ventricle and tibial artery. Conclusions. In this study we identified for the first time a genome-wide significant association of a genetic variant with ERP. Our findings of a locus in the KCND3 gene not only provide insights into the genetic determinants but also into the pathophysiological mechanism of ERP, discovering a promising candidate for functional studies. Funding. For detailed information per study, see Acknowledgments.
Authors
Alexander Teumer, Teresa Trenkwalder, Thorsten Kessler, Yalda Jamshidi, Marten E. van den Berg, Bernhard Kaess, Christopher P. Nelson, Rachel Bastiaenen, Marzia De Bortoli, Alessandra Rossini, Isabel Deisenhofer, Klaus Stark, Solmaz Assa, Peter S. Braund, Claudia Cabrera, Anna F. Dominiczak, Martin Gögele, Leanne M. Hall, M. Arfan Ikram, Maryam Kavousi, Karl J. Lackner, Christian Müller, Thomas Münzel, Matthias Nauck, Sandosh Padmanabhan, Norbert Pfeiffer, Tim D. Spector, Andre G. Uitterlinden, Niek Verweij, Uwe Völker, Helen R. Warren, Mobeen Zafar, Stephan B. Felix, Jan A. Kors, Harold Snieder, Patricia B. Munroe, Cristian Pattaro, Christian Fuchsberger, Georg Schmidt, Ilja M. Nolte, Heribert Schunkert, Peter Pramstaller, Philipp S. Wild, Pim van der Harst, Bruno H. Stricker, Renate B. Schnabel, Nilesh J. Samani, Christian Hengstenberg, Marcus Dörr, Elijah R. Behr, Wibke Reinhard
Abstract
High levels of circulating miR-16 in the serum of multiple myeloma (MM) patients are independently associated with longer survival. Although the tumor suppressor function of intracellular miR-16 in cancer cells, including MM plasma cells (PCs), has been highly elucidated, its extracellular role in maintaining a non-supportive cancer microenvironment has not been fully explored. Here, we show that miR-16 can be actively secreted by MM cells through extracellular vesicles (EVs), and its extracellular and intracellular levels are directly correlated. We also show that EVs isolated from MM patients and from the conditioned media of MM-PCs can differentiate circulating monocytes to M2-tumor supportive macrophages (TAMs) and that the presence of higher levels of extracellular miR-16 counteracts this effect. In agreement with these observations, our data show that miR-16 directly targets the IKKα/β complex of the NF-kB canonical pathway, which is known to play a critical role in polarizing macrophages toward an M2 phenotype. By using a miR-15a-16-1 knockout mouse model, we also show that loss of the miR-16 cluster supports polarization to M2-macrophages. Finally, we demonstrate the therapeutic benefit of miR-16 overexpression in potentiating the anti-MM activity by a proteasome inhibitor in the presence of MM resident bone marrow TAM.
Authors
Jihane Khalife, Jayeeta Ghose, Marianna Martella, Domenico Viola, Alberto Rocci, Estelle Troadec, Cesar Terrazas, Abhay R. Satoskar, Emine Gulsen Gunes, Ada Dona, James F. Sanchez, P. Leif Bergsagel, Marta Chesi, Alex Pozhitkov, Steven Rosen, Guido Marcucci, Jonathan J. Keats, Craig C. Hofmeister, Amrita Krishnan, Enrico Caserta, Flavia Pichiorri
Abstract
Anemia of β-thalassemias is caused by ineffective erythropoiesis and reduced red cell survival. Several lines of evidence indicate that iron/heme restriction is a potential therapeutic strategy for the disease. Glycine is a key initial substrate for heme and globin synthesis. We provide evidence that bitopertin, a glycine transport inhibitor administered orally, improves anemia, reduces hemolysis, diminishes ineffective erythropoiesis, and increases red cell survival in a mouse model of β-thalassemia (Hbbth3/+ mice). Bitopertin ameliorates erythroid oxidant damage as indicated by a reduction in membrane-associated free α–globin chain aggregates, in reactive oxygen species cellular content, in membrane-bound hemichromes and in HRI activation and elF2α phosphorylation. The improvement of β-thalassemic ineffective erythropoiesis is associated with diminished mTOR activation, Rab5, Lamp1, and p62 accumulation, indicating an improved autophagy. Bitopertin also upregulates liver hepcidin and diminishes liver iron overload. The hematologic improvements achieved by bitopertin are blunted by the concomitant administration of the iron chelator deferiprone, suggesting that an excessive restriction of iron availability might negate the beneficial effects of bitopertin. These data provide important and clinically relevant insights into glycine restriction and reduced heme synthesis strategies for the treatment of β-thalassemia.
Authors
Alessandro Matte, Enrica Federti, Michael Winter, Annette Koerner, Anja Harmeier, Norman Mazer, Tomas Tomka, Maria Luisa Di Paolo, Luigia De Falco, Immacolata Andolfo, Elisabetta Beneduce, Achille Iolascon, Alejandra Macias-Garcia, Jane-Jane Chen, Anne Janin, Christophe Leboeuf, Francesco Turrini, Carlo Brugnara, Lucia De Franceschi
Abstract
Platelet inositol hexakisphosphate kinase 1 (IP6K1) has been shown to control systemic inflammation. Herein, we examined if platelets and IP6K1 regulate pancreatic tissue injury via formation of NETs in experimental models of acute pancreatitis (AP) in mice. By use of electron microscopy abundant NET formation was observed in the inflamed pancreas. These NETs contained numerous microparticles (MP) expressing CD41 or Mac-1. Platelet depletion reduced deposition of NET-MP complexes in the inflamed pancreas. Circulating platelet-neutrophil aggregates (PNA) were increased and inhibition of P-selectin not only disrupted PNA formation but also reduced NETs formation in the inflamed pancreas. NETs depleted of MPs had lower capacity to provoke amylase secretion and STAT-3 phosphorylation in acinar cells. Taurocholate-induced NETs formation, inflammation and tissue damage in the pancreas were decreased in IP6K1-deficient mice. Thrombin stimulation of mixtures of wild-type platelets and neutrophils resulted in NETs formation but not when IP6K1-deficient platelets were incubated with wild-type neutrophils. Polyphosphate rescue restored thrombin-induced NET formation in mixtures of IP6K1-deficient platelets and wild-type neutrophils. Platelet IP6K1 regulates NET-MP complex formation in the pancreas of mice during induction of AP. Targeting platelet IP6K1 might useful to decrease NET-dependent pancreatic tissue inflammation and tissue injury in patients with AP.
Authors
Raed Madhi, Milladur Rahman, Dler Taha, Johan Linders, Mohammed Merza, Yongzhi Wang, Matthias Mörgelin, Henrik Thorlacius
Abstract
The red blood cell (RBC) storage lesion is a multi-parametric response that occurs during storage at 4°C, but its impact on transfused patients remains unclear. In studies of the RBC storage lesion, the temperature transition from cold storage to normal body temperature that occurs during transfusion has received limited attention. We hypothesized that multiple deleterious events might occur in this period of increasing temperature. We show dramatic alterations in several properties of therapeutic blood units stored at 4°C after warming them to normal body temperature (37°C), as well as febrile temperature (40°C). In particular, the intracellular content and redox state of nicotinamide adenine dinucleotide phosphate [NADP(H)] were directly affected by post-storage incubation at 37°C, as well as by pro-oxidant storage conditions. Modulation of the NADPH-producing pentose phosphate pathway, but not the prevention of hemoglobin autoxidation by conversion of oxyhemoglobin to carboxyhemoglobin, provided protection against storage-induced alterations in RBCs, demonstrating the central role of NADPH in mitigating increased susceptibility of stored RBCs to oxidative stress. We propose that assessing RBCs oxidative status after restoration of body temperature provides a sensitive tool to detect storage-related alterations, and has the potential to improve the quality of stored RBCs for transfusion.
Authors
Aline Roch, Nicholas J. Magon, Jessica Maire, Cacang Suarna, Anita Ayer, Sophie Waldvogel, Beat. A. Imhof, Mark J. Koury, Roland Stocker, Marc Schapira
Abstract
Islet transplantation can restore lost glycemic control in type 1 diabetes subjects, but is restricted in its clinical application by limiting supplies of islets and the need for heavy immune suppression to prevent rejection. TNFAIP3, encoding the ubiquitin editing enzyme A20, regulates the activation of immune cells by raising NF-κB signalling thresholds. Here we show that increasing A20 expression in allogeneic islet grafts resulted in permanent survival for approximately 45% of recipients, and > 80% survival when combined with subtherapeutic rapamycin. Allograft survival was dependent upon regulatory T cells, was antigen-specific and grafts showed reduced expression of inflammatory factors. Transplantation of islets with A20 containing a loss-of-function variant (I325N) resulted in increased RIPK1 ubiquitination and NF-κB signalling, graft hyper-inflammation and acute allograft rejection. Overexpression of A20 in human islets potently reduced expression of inflammatory mediators with no impact on glucose stimulated insulin secretion. Therapeutic administration of A20 raises inflammatory signalling thresholds to favour immune tolerance and promotes islet allogeneic survival. Clinically this would allow for reduced immunosuppression and support the use of alternate islet sources.
Authors
Nathan W. Zammit, Stacey N. Walters, Karen L. Seeberger, Philip J. O’Connell, Gregory S. Korbutt, Shane T. Grey
Abstract
Mesenchymal stromal/stem cell (MSC) therapy has shown promise in experimental models of idiopathic pulmonary fibrosis (IPF). The aim of this study was to test the therapeutic effects of MSC-extracellular vesicles/exosomes (MEx) in a bleomycin-induced pulmonary fibrosis model and investigate putative mechanisms of action. Exosomes were isolated from media conditioned by human bone marrow MSCs. Adult mice (C57BL/6 strain) were challenged with endotracheal instillation of bleomycin and treated with MEx concurrently or for reversal models, at day 7 or 21. Experimental groups were assessed at day 7 and/or at day 14 or 28. Bleomycin-challenged mice presented with severe septal thickening and prominent fibrosis, and this was effectively prevented or reversed by a single dose of MEx. Furthermore, MEx therapy modulated whole lung macrophage phenotype and shifted the proportion of lung ‘proinflammatory’ classical monocytes, non-classical monocytes and alveolar macrophages to favor the monocyte/macrophage profiles of untreated-control mice. A parallel immunomodulatory effect was demonstrated in the bone marrow. Notably, transplantation of MEx-preconditioned bone marrow-derived monocytes alleviated core features of pulmonary fibrosis and lung inflammation. Proteomic analysis further revealed a signature enriched in non-inflammatory monocyte genes following MEx therapy supporting the immuno-regulatory, anti-inflammatory effect of MEx.We conclude that a bolus dose of MEx prevents and reverts core features of bleomycin-induced pulmonary fibrosis, and that the beneficial actions of MEx may be mediated via systemic modulation of monocyte phenotypes.
Authors
Nahal Mansouri, Gareth R. Willis, Angeles Fernandez-Gonzalez, Monica Reis, Sina Nassiri, Alex Mitsialis, Stella Kourembanas
Abstract
The acute respiratory distress syndrome (ARDS) is an inflammatory lung disorder that frequently complicates critical illness, and most commonly occurs in the setting of sepsis. Although a number of clinical and environmental risk factors for ARDS have been described, not all patients with risk factors develop the syndrome, raising the possibility of genetic underpinnings for ARDS susceptibility. We have previously reported that circulating cell-free hemoglobin (CFH) is elevated during sepsis, and higher levels are associated with worse outcomes. CFH is rapidly scavenged by the plasma protein haptoglobin (Hp). A common HP genetic variant HP2 is unique to humans and represents 60% of the HP allele frequency in populations of European ancestry. The HP2 gene product has reduced ability to inhibit CFH-mediated inflammation and oxidative stress compared to the alternative HP1. We hypothesized that the HP2 variant increases ARDS susceptibility during sepsis when plasma CFH levels are elevated. In a murine model of sepsis with elevated CFH levels, transgenic mice homozygous for Hp2 had increased lung inflammation, pulmonary vascular permeability, lung apoptosis, and mortality compared to mice homozygous for the alternative allele Hp1. We then tested the clinical relevance of our findings in a prospective observational cohort study of 496 septic critically ill adults, and found that the HP2 variant was significantly associated with increased ARDS susceptibility (odds ratio 1.41 per HP2 allele, 95% confidence interval 1.06 – 1.88, P = 0.018) after controlling for clinical risk factors and plasma CFH. This relationship between the HP2 genetic variant and ARDS risk was only seen in patients with elevated plasma CFH levels. These observations identify the HP2 variant as a novel genetic ARDS risk factor during sepsis, and may have important implications in the study and treatment of ARDS.
Authors
V. Eric Kerchberger, Julie A. Bastarache, Ciara M. Shaver, Hiromasa Nagata, J. Brennan McNeil, Stuart R. Landstreet, Nathan D. Putz, Wen-Kuang Yu, Jordan Jesse, Nancy E. Wickersham, Tatiana N. Sidorova, David R. Janz, Chirag R. Parikh, Edward D. Siew, Lorraine B. Ware
Abstract
Background: Hydroxymethyl-glutaryl-coenzyme A reductase inhibitors (‘statins’) are prescribed to millions of people. Statins are anti-inflammatory independent of their cholesterol-reducing effects. To date, most reports on the immune effects of statins have assayed a narrow array of variables and have focused on cell lines, rodent models, or patient cohorts. We sought to define the effect of rosuvastatin on the ‘immunome’ of healthy, normocholesterolemic subjects. Methods: Prospective study of rosuvastatin (20 mg/day x 28 days) in 18 statin-naïve adults with low density lipoprotein-cholesterol <130 mg/dL. A panel of >180 immune/biochemical/endocrinologic variables was measured at baseline, and days 14, 28, and 42 (14 days after drug withdrawal). Drug effect was evaluated using linear mixed effects models. Potential interactions between drug and baseline high-sensitivity C-reactive protein (hsCRP) were evaluated. Results: A wide array of immune measures changed (nominal p<0.05) during rosuvastatin treatment, although the changes were modest in magnitude and few met a false discovery rate of 0.05. Among changes noted were a concordant increase in pro-inflammatory cytokines (IFNγ, IL-1β, IL-5, IL-6, TNFα) and peripheral blood neutrophil frequency, and a decline in activated T regulatory cell frequency. Several drug effects were significantly modified by baseline hsCRP, and some did not resolve after drug withdrawal. Among other unexpected rosuvastatin effects were changes in erythrocyte indices, glucose-regulatory hormones, CD8+ T cells, and haptoglobin. Conclusion: Rosuvastatin induces modest changes in immunologic and metabolic measures in normocholesterolemic subjects, with several effects dependent upon baseline CRP. Future, larger studies are warranted to validate these changes and their physiological significance.
Authors
Peer W. F. Karmaus, Min Shi, Shira Perl, Angélique Biancotto, Julián Candia, Foo Cheung, Yuri Kotliarov, Neal Young, Michael B. Fessler
Abstract
Accumulation of lysosomal storage material and late-stage neurodegeneration are hallmarks of lysosomal storage disorders (LSDs) affecting the brain. Yet, for most LSDs, including CLN3 disease, the most common form of childhood dementia, it is unclear what mechanisms drive neurologic symptoms. Do deficits arise from loss of function of the mutated protein or toxicity from storage accumulation? Here, using in vitro voltage sensitive dye imaging and in vivo electrophysiology, we find progressive hippocampal dysfunction occurs prior to notable lysosomal storage and neuronal loss in two CLN3 disease mouse models. Pharmacologic reversal of lysosomal storage deposition in young mice does not rescue this circuit dysfunction. Additionally, we find that CLN3 disease mice lose an electrophysiologic marker of new memory encoding – hippocampal sharp wave ripples. This discovery, which is also seen in Alzheimer’s disease, suggests the possibility of a shared electrophysiologic signature of dementia. Overall, our data describes new insights into previously unknown network-level changes occurring in LSDs affecting the central nervous system, and highlight the need for new therapeutic interventions targeting early circuit defects.
Authors
Rebecca C. Ahrens-Nicklas, Luis Tecedor, Arron Hall, Elena Lysenko, Akiva S. Cohen, Beverly L. Davidson, Eric D. Marsh
Abstract
Background: In this study, we aimed to identify the lipidomic predictors of early type-2 diabetic kidney disease (DKD) progression which are currently undefined DKD progression. Methods: This longitudinal study included 92 American Indians with type-2 diabetes. Serum lipids (406 from 18 classes) were quantified using mass spectrometry from baseline samples when iothalamate glomerular filtration rate (GFR) was ≥90 mL/min. Affymetrix GeneChip Array was used to measure renal transcript expression. DKD Progression was defined as ≥40% decline in GFR during follow up. Results: Participants had a mean age of 45±9 years and median urine albumin-creatinine ratio of 43 (interquartile range 11 to 144). The 32 progressors had significantly higher relative abundance of polyunsaturated triacylglycerols (TAG)s and a lower abundance of C16-20 acylcarnitines (AC)s (p<0.001). In a Cox regression model the main effect terms of unsaturated free fatty acids and phosphatidylethanolamines and the interaction terms of C16-20 ACs and short, low-double-bond TAGs by categories of albuminuria independently predicted progression of DKD. Renal expression of acetyl-CoA carboxylase encoding gene (ACACA) correlated with serum diacylglycerols in the glomerular compartment (r=0.36, p=0.006), and with low-double-bond TAGs in the tubulointerstitial compartment (r=0.52, p<0.001). Conclusion: Collectively, the findings reveal a previously unrecognized link between lipid markers of impaired mitochondrial β-oxidation and enhanced lipogenesis, with DKD progression, in individuals with preserved GFR. Renal acetyl-CoA carboxylase activation accompanies these lipidomic changes and suggests that it may be the underlying mechanism linking lipid abnormalities to DKD progression. Funding: R24DK082841, K08DK106523, R03DK121941, P30DK089503, P30DK081943, P30DK020572
Authors
Farsad Afshinnia, Viji Nair, Jiahe Lin, Thekkelnaycke M. Rajendiran, Tanu Soni, Jaeman Byun, Kumar Sharma, Patrice E. Fort, Thomas W. Gardner, Helen C. Looker, Robert G. Nelson, Frank C. Brosius, Eva L. Feldman, George Michailidis, Matthias Kretzler, Subramaniam Pennathur
Abstract
Kabuki syndrome 1 (KS1) is a Mendelian disorder of the epigenetic machinery caused by mutations in the gene encoding KMT2D, which methylates lysine 4 on histone H3 (H3K4). KS1 is characterized by intellectual disability, postnatal growth retardation, and distinct craniofacial dysmorphisms. A mouse model (Kmt2d+/bGeo) exhibits features of the human disorder and has provided insight into other phenotypes; however, the mechanistic basis of skeletal abnormalities and growth retardation remains elusive. Using high-resolution micro-computed tomography we show that Kmt2d+/bGeo mice have shortened long bones and ventral bowing of skulls. In vivo expansion of growth plates within skulls and long bones suggests disrupted endochondral ossification as a common disease mechanism. Stable chondrocyte cell lines harboring inactivating mutations in Kmt2d exhibit precocious differentiation, further supporting this mechanism. A known inducer of chondrogenesis, SOX9, and its targets show markedly increased expression in Kmt2d-/- chondrocytes. By transcriptome profiling, we identify Shox2 as a putative KMT2D target. We propose that decreased KMT2D-mediated H3K4me3 at Shox2 releases Sox9 inhibition and thereby leads to enhanced chondrogenesis, providing a novel and plausible explanation for precocious chondrocyte differentiation. Our findings provide insight into the pathogenesis of growth retardation in KS1 and suggest novel therapeutic approaches for this and related disorders.
Authors
Jill A. Fahrner, Wan-Ying Lin, Ryan C. Riddle, Leandros Boukas, Valerie B. DeLeon, Sheetal Chopra, Susan E. Lad, Teresa Romeo Luperchio, Kasper D. Hansen, Hans T. Bjornsson
Abstract
Transcriptomic profiling classifies pancreatic ductal adenocarcinoma (PDAC) into several molecular subtypes with distinctive histological and clinical characteristics. However, little is known about the molecular mechanisms that define each subtype and their correlation with clinical outcome. Mutant KRAS is the most prominent driver in PDAC, present in over 90% of tumors, but the dependence of tumors on oncogenic KRAS signaling varies between subtypes. In particular, squamous subtype are relatively independent of oncogenic KRAS signaling and typically display much more aggressive clinical behavior versus progenitor subtype. Here, we identified that YAP1 activation is enriched in the squamous subtype and associated with poor prognosis. Activation of YAP1 in progenitor subtype cancer cells profoundly enhanced malignant phenotypes and transformed progenitor subtype cells into squamous subtype. Conversely, depletion of YAP1 specifically suppressed tumorigenicity of squamous subtype PDAC cells. Mechanistically, we uncovered a significant positive correlation between WNT5A expression and the YAP1 activity in human PDAC, and demonstrated that WNT5A overexpression led to YAP1 activation and recapitulated YAP1-dependent but Kras-independent phenotype of tumor progression and maintenance. Thus, our study identifies YAP1 oncogene as a major driver of squamous subtype PDAC and uncovers the role of WNT5A in driving PDAC malignancy through activation of the YAP pathway.
Authors
Bo Tu, Jun Yao, Sammy Ferri-Borgogno, Jun Zhao, Shujuan Chen, Qiuyun Wang, Liang Yan, Xin Zhou, Cihui Zhu, Seungmin Bang, Qing Chang, Christopher A. Bristow, Ya'an Kang, Hongwu Zheng, Huamin Wang, Jason B. Fleming, Michael Kim, Timothy P. Heffernan, Giulio F. Draetta, Duojia Pan, Anirban Maitra, Wantong Yao, Sonal Gupta, Haoqiang Ying
Abstract
Middle East respiratory syndrome coronavirus (MERS-CoV) emerged in 2012 in the Kingdom of Saudi Arabia and has caused over 2400 cases and more than 800 deaths. Epidemiological studies identified diabetes as the primary comorbidity associated with severe and/or lethal MERS-CoV infection. Understanding how diabetes affects MERS is important due to the global burden of diabetes and pandemic potential of MERS-CoV. We used a model in which mice were made susceptible to MERS-CoV by expressing human DPP4 and type 2 diabetes was induced by administering a high fat diet. Upon infection with MERS-CoV, diabetic mice had a prolonged phase of severe disease and delayed recovery which was independent of virus titers. Histological analysis revealed that diabetic mice had delayed inflammation which was then prolonged through 21 dpi. Diabetic mice had fewer inflammatory monocyte/macrophages and CD4+ T cells which correlated with lower levels of Ccl2 and Cxcl10 expression. Diabetic mice also had lower levels of Tnfa, Il6, Il12b, and Arg1 expression and higher levels of Il17a expression. These data suggest that the increased disease severity observed in individuals with MERS and comorbid type 2 diabetes is likely due to a dysregulated immune response which results in more severe and prolonged lung pathology.
Authors
Kirsten A. Kulcsar, Christopher M. Coleman, Sarah E. Beck, Matthew B. Frieman
Abstract
Increased fibrosis is a characteristic remodeling response to biomechanical and neurohumoral stress and a determinant of cardiac mechanical and electrical dysfunction in disease. Stress-induced activation of cardiac fibroblasts (CF) is a critical step in the fibrotic response, although the precise sequence of events underlying activation of these critical cells in vivo remain unclear. Here, we test the hypothesis that a βIV-spectrin/STAT3 complex is essential for maintenance of a quiescent phenotype (basal non-activated state) in CFs. We report increased fibrosis, decreased cardiac function, and electrical impulse conduction defects in genetic and acquired mouse models of βIV-spectrin deficiency. Loss of betaIV-spectrin function promotes STAT3 nuclear accumulation and transcriptional activity, altered gene expression and CF activation. Furthermore, we demonstrate that a quiescent phenotype may be restored in βIV-spectrin deficient fibroblasts by expressing a βIV-spectrin fragment including the STAT3-binding domain or through pharmacological STAT3 inhibition. We find that in vivo STAT3 inhibition abrogates fibrosis and cardiac dysfunction in the setting of global βIV-spectrin deficiency. Finally, we demonstrate that fibroblast-specific deletion of βIV-spectrin is sufficient to induce fibrosis and decreased cardiac function. We propose that the βIV-spectrin/STAT3 complex is a determinant of fibroblast phenotype and fibrosis, with implications for remodeling response in cardiovascular disease.
Authors
Nehal J. Patel, Drew M. Nassal, Amara D. Greer-Short, Sathya D. Unudurthi, Benjamin W. Scandling, Daniel Gratz, Xianyao Xu, Anuradha Kalyanasundaram, Vadim V. Fedorov, Federica Accornero, Peter J. Mohler, Keith J. Gooch, Thomas J. Hund
Abstract
Filoviruses of the genus Ebolavirus include five species with marked differences in their ability to cause disease in humans. From the highly virulent Ebola virus to the seemingly nonpathogenic Reston virus, case-fatality rates can range between 0-90%. In order to understand the molecular basis of these differences it is imperative to establish disease models that recapitulate human disease as faithfully as possible. Non-human primates are the gold-standard models for filovirus pathogenesis, but comparative studies are skewed by the fact that Reston virus infection can be lethal for NHP. Here we have used HLA-A2 transgenic, NOD-scid-interleukin 2γ receptor knockout (NSG-A2) mice reconstituted with human hematopoiesis to compare Ebola virus and Reston virus pathogenesis in a human-like environment. While significantly less pathogenic than Ebola virus, Reston virus killed 20% of infected mice, a finding that was linked to exacerbated inflammation and viral replication in the liver. In addition, ‘humanized’ mice recapitulated the case-fatality ratios of different Ebolavirus species in humans. Our findings point out at humanized mice as a putative model to test the pathogenicity of newly discovered filoviruses, and warrants further investigations on Reston virus pathogenesis in humans.
Authors
Beatriz Escudero-Pérez, Paula Ruibal, Monika Rottstegge, Anja Lüdtke, Julia R. Port, Kristin Hartmann, Sergio Gómez-Medina, Juergen Müller-Guhl, Emily V. Nelson, Susanne Krasemann, Estefanía Rodríguez, César Muñoz-Fontela
Abstract
Severe asthma with fungal sensitization (SAFS) defines a subset of human asthmatics with allergy to one or more fungal species and difficult to control asthma. We have reported that human asthmatics sensitized to fungi have worse lung function and a higher degree of atopy, which was associated with higher IL-1RA levels in bronchoalveolar lavage fluid. IL-1RA further demonstrated a significant negative association with bronchial hyperresponsiveness to methacholine. Here, we show that IL-1α and IL-1β are elevated in both bronchoalveolar lavage fluid and sputum from human asthmatics sensitized to fungi, implicating an association with IL-1α, IL-1β or IL-1RA in fungal asthma severity. In an experimental model of fungal-associated allergic airway inflammation, we demonstrate that IL-1R1 signaling promotes type 1 (IFN-γ, CXCL9, CXCL10) and type 17 (IL-17A, IL-22) responses that were associated with neutrophilic inflammation and increased airway hyperreactivity. Each of these were exacerbated in the absence of IL-1RA. Administration of human recombinant IL-1RA (Kineret/Anakinra) during fungal-associated allergic airway inflammation improved airway hyperreactivity and lowered type 1 and type 17 responses. Taken together, these data suggest that IL-1 receptor signaling contributes to fungal asthma severity via immunopathogenic type 1 and type 17 responses and can be targeted for improving allergic asthma severity.
Authors
Matthew S. Godwin, Kristen M. Reeder, Jaleesa M. Garth, Jonathan P. Blackburn, MaryJane Jones, Zhihong Yu, Sadis Matalon, Annette T. Hastie, Deborah A. Meyers, Chad Steele
Abstract
Lymphatic malformations (LMs) are congenital, non-neoplastic vascular malformations associated with post-zygotic activating PIK3CA mutations. The mutation spectrum within LMs is narrow, with the majority having one of three “hotspot” mutations. Despite this relative genetic homogeneity, clinical presentations differ dramatically. We used molecular inversion probes and droplet digital polymerase chain reaction to perform deep, targeted sequencing of PIK3CA in 271 affected and unaffected tissue samples from 81 individuals with isolated LMs and retrospectively collected clinical data. Pathogenic PIK3CA mutations were identified in affected LM tissue in 64 individuals (79%) with isolated LMs, with variant allele fractions (VAFs) ranging from 0.1 to 13%. Initial analyses revealed no correlation between VAF and phenotype variables. Recognizing that different mutations activate PI3K to varying degrees, we developed a metric, the genotype-adjusted VAF (GVAF), to account for differences in mutation strength, and found significantly higher GVAFs in LMs with more severe clinical characteristics including orofacial location or microcystic structure. In addition to providing insight into LM pathogenesis, we believe GVAF may have broad applicability for genotype-phenotype analyses in mosaic disorders.
Authors
Kaitlyn Zenner, Chi Vicky Cheng, Dana M. Jensen, Andrew E. Timms, Giridhar Shivaram, Randall Bly, Sheila Ganti, Kathryn B. Whitlock, William B. Dobyns, Jonathan Perkins, James T. Bennett
Abstract
miR-511-3p, encoded by CD206/Mrc1, was demonstrated to reduce allergic inflammation and promote alternative (M2) macrophage polarization. Here, we sought to elucidate the fundamental mechanism by which miR-511-3p attenuates allergic inflammation and promotes macrophage polarization. Compared with wild-type mice, the allergen-challenged Mrc1-/- mice showed increased airway hyper-responsiveness (AHR) and inflammation. However, this increased AHR and inflammation were significantly attenuated when these mice were pre-transduced with adeno-associated virus (AAV)-miR-511-3p. Gene expression profiling of macrophages identified Ccl2 as one of the major genes that was highly expressed in M2 macrophages but antagonized by miR-511-3p. The interaction between miR-511-3p and Ccl2 was confirmed by in silico analysis and mRNA-miRNA pull-down assay. Further evidence for the inhibition of Ccl2 by miR-511-3p was given by reduced levels of Ccl2 in supernatants of miR-511-3p transduced macrophages and in bronchoalveolar lavage fluids of AAV-miR-511-3p-infected Mrc1-/- mice. Mechanistically, we demonstrated that Ccl2 promotes M1 macrophage polarization by activating RhoA signaling through Ccr2. The interaction between Ccr2 and RhoA was also supported by co-immunoprecipitation assay. Importantly, inhibition of RhoA signaling suppressed cockroach allergen-induced AHR and lung inflammation. These findings suggest a novel mechanism by which miR-511-3p regulates allergic inflammation and macrophage polarization by targeting Ccl2 and its downstream Ccr2/RhoA axis.
Authors
Danh C. Do, Jie Mu, Xia Ke, Karan Sachdeva, Zili Qin, Mei Wan, Faoud T. Ishmael, Peisong Gao
