Recent molecular discoveries in angiogenesis and antiangiogenic therapies in cancer

• Text
• PDF

Abstract

Four decades ago, angiogenesis was recognized as a therapeutic target for blocking cancer growth. Because of its importance, VEGF has been at the center stage of antiangiogenic therapy. Now, several years after FDA approval of an anti-VEGF antibody as the first antiangiogenic agent, many patients with cancer and ocular neovascularization have benefited from VEGF-targeted therapy; however, this anticancer strategy is challenged by insufficient efficacy, intrinsic refractoriness, and resistance. Here, we examine recent discoveries of new mechanisms underlying angiogenesis, discuss successes and challenges of current antiangiogenic therapy, and highlight emerging antiangiogenic paradigms.

Authors

Jonathan Welti, Sonja Loges, Stefanie Dimmeler, Peter Carmeliet

A conversation with John Oates

• Text
• PDF

Abstract

Authors

Ushma S. Neill

A tribute to Emil Frei III

• Text
• PDF

Abstract

Authors

Edward J. Benz Jr., David G. Nathan

Collagenases and cracks in the plaque

• Text
• PDF

Abstract

The core of an atheromatous plaque contains lipids, macrophages, and cellular debris, typically covered by a fibrous cap that separates the thrombogenic core from the blood. Rupture of the fibrous cap causes most fatal myocardial infarctions. Interstitial collagen confers tensile strength on the cap, as it does in skin and tendons. In 1994, Peter Libby and colleagues demonstrated overexpression of collagenolytic enzymes in atheromatous plaques and implicated MMPs in the destabilization of these lesions.

Authors

Peter Libby

Rapamycin, anti-aging, and avoiding the fate of Tithonus

• Text
• PDF

Abstract

The discovery that rapamycin increased the lifespan of mice was recognized by Science as one of the top 10 scientific breakthroughs of 2009. In addition to increasing lifespan, Neff and colleagues show that while rapamycin improves several functions/pathologies that change with age, it has little effect on the majority of the physiological and structural parameters they evaluated. What do these data tell us about the ability of rapamycin to delay aging and improve quality of life, i.e., prevent the fate of Tithonus?

Authors

Arlan Richardson

The future of molecular chaperones and beyond

• Text
• PDF

Abstract

Protection of hair cells by HSP70 released by supporting cells is reported by May et al. in this issue of the JCI. Their findings suggest a new way to reduce ototoxicity from therapeutic medications and raise larger questions about the role and integration of heat shock proteins in non–cell-autonomous responses to stress. Increasing evidence suggests an important role for extracellular heat shock proteins in both the nervous system and the immune system. The work also suggests that defective chaperones could cause ear disease and supports the potential use of chaperone therapeutics.

Authors

Rona G. Giffard, Alberto J.L. Macario, Everly Conway de Macario

Chronic THC intake modifies fundamental cerebellar functions

• Text
• PDF

Abstract

Delta9-tetrahydrocannabinol (THC), the principal bioactive component in the Cannabis plant, is truly a captivating drug. Acute and chronic THC intake produces a spectrum of biological effects ranging from transient psychotropic effects to prolonged medicinal benefits, many of which have been fostered for centuries by our society. In the July 2013 issue of the JCI, Cutando et al. combined mouse genetics with classic mouse behavioral analysis to deepen our understanding of the physiological consequence of subchronic THC intake on eyeblink reflexes, a fundamental neuronal adaptive response, revealing that this regimen leads to downregulation of the cannabinoid CB₁ receptor (referred to as CB1 in the Cutando et al. article) in cerebellar stress fibers and the activation of microglia, raising provocative new questions about the safety profile of regimented THC intake.

Authors

Nephi Stella

Colon cancer progression is driven by APEX1-mediated upregulation of Jagged

• Text
• PDF

Abstract

Aberrant expression of apurinic-apyrimidinic endonuclease–1 (APEX1) has been reported in numerous human solid tumors and is positively correlated with cancer progression; however, the role of APEX1 in tumor progression is poorly defined. Here, we show that APEX1 contributes to aggressive colon cancer behavior and functions as an upstream activator in the Jagged1/Notch signaling pathway. APEX1 overexpression or knockdown in human colon cancer cell lines induced profound changes in malignant properties such as cell proliferation, anchorage-independent growth, migration, invasion, and angiogenesis in vitro and in tumor formation and metastasis in mouse xenograft models. These oncogenic effects of APEX1 were mediated by the upregulation of Jagged1, a major Notch ligand. Furthermore, APEX1 expression was associated with Jagged1 in various colon cancer cell lines and in tissues from colon cancer patients. This finding identifies APEX1 as a positive regulator of Jagged1/Notch activity and suggests that it is a potential therapeutic target in colon cancers that exhibit high levels of Jagged1/Notch signaling.

Authors

Mi-Hwa Kim, Hong-Beum Kim, Sang Pil Yoon, Sung-Chul Lim, Man Jin Cha, Young Jin Jeon, Sang Gon Park, In-Youb Chang, Ho Jin You

MerTK inhibition in tumor leukocytes decreases tumor growth and metastasis

• Text
• PDF

Abstract

MerTK, a receptor tyrosine kinase (RTK) of the TYRO3/AXL/MerTK family, is expressed in myeloid lineage cells in which it acts to suppress proinflammatory cytokines following ingestion of apoptotic material. Using syngeneic mouse models of breast cancer, melanoma, and colon cancer, we found that tumors grew slowly and were poorly metastatic in MerTK^–/– mice. Transplantation of MerTK^–/– bone marrow, but not wild-type bone marrow, into lethally irradiated MMTV-PyVmT mice (a model of metastatic breast cancer) decreased tumor growth and altered cytokine production by tumor CD11b⁺ cells. Although MerTK expression was not required for tumor infiltration by leukocytes, MerTK^–/– leukocytes exhibited lower tumor cell–induced expression of wound healing cytokines, e.g., IL-10 and growth arrest-specific 6 (GAS6), and enhanced expression of acute inflammatory cytokines, e.g., IL-12 and IL-6. Intratumoral CD8⁺ T lymphocyte numbers were higher and lymphocyte proliferation was increased in tumor-bearing MerTK^–/– mice compared with tumor-bearing wild-type mice. Antibody-mediated CD8⁺ T lymphocyte depletion restored tumor growth in MerTK^–/– mice. These data demonstrate that MerTK signaling in tumor-associated CD11b⁺ leukocytes promotes tumor growth by dampening acute inflammatory cytokines while inducing wound healing cytokines. These results suggest that inhibition of MerTK in the tumor microenvironment may have clinical benefit, stimulating antitumor immune responses or enhancing immunotherapeutic strategies.

Authors

Rebecca S. Cook, Kristen M. Jacobsen, Anne M. Wofford, Deborah DeRyckere, Jamie Stanford, Anne L. Prieto, Elizabeth Redente, Melissa Sandahl, Debra M. Hunter, Karen E. Strunk, Douglas K. Graham, H. Shelton Earp III

ARHGDIA mutations cause nephrotic syndrome via defective RHO GTPase signaling

• Text
• PDF

Abstract

Nephrotic syndrome (NS) is divided into steroid-sensitive (SSNS) and -resistant (SRNS) variants. SRNS causes end-stage kidney disease, which cannot be cured. While the disease mechanisms of NS are not well understood, genetic mapping studies suggest a multitude of unknown single-gene causes. We combined homozygosity mapping with whole-exome resequencing and identified an ARHGDIA mutation that causes SRNS. We demonstrated that ARHGDIA is in a complex with RHO GTPases and is prominently expressed in podocytes of rat glomeruli. ARHGDIA mutations (R120X and G173V) from individuals with SRNS abrogated interaction with RHO GTPases and increased active GTP-bound RAC1 and CDC42, but not RHOA, indicating that RAC1 and CDC42 are more relevant to the pathogenesis of this SRNS variant than RHOA. Moreover, the mutations enhanced migration of cultured human podocytes; however, enhanced migration was reversed by treatment with RAC1 inhibitors. The nephrotic phenotype was recapitulated in arhgdia-deficient zebrafish. RAC1 inhibitors were partially effective in ameliorating arhgdia-associated defects. These findings identify a single-gene cause of NS and reveal that RHO GTPase signaling is a pathogenic mediator of SRNS.

Authors

Heon Yung Gee, Pawaree Saisawat, Shazia Ashraf, Toby W. Hurd, Virginia Vega-Warner, Humphrey Fang, Bodo B. Beck, Olivier Gribouval, Weibin Zhou, Katrina A. Diaz, Sivakumar Natarajan, Roger C. Wiggins, Svjetlana Lovric, Gil Chernin, Dominik S. Schoeb, Bugsu Ovunc, Yaacov Frishberg, Neveen A. Soliman, Hanan M. Fathy, Heike Goebel, Julia Hoefele, Lutz T. Weber, Jeffrey W. Innis, Christian Faul, Zhe Han, Joseph Washburn, Corinne Antignac, Shawn Levy, Edgar A. Otto, Friedhelm Hildebrandt

Whole body correction of mucopolysaccharidosis IIIA by intracerebrospinal fluid gene therapy

• Text
• PDF

Abstract

For most lysosomal storage diseases (LSDs) affecting the CNS, there is currently no cure. The BBB, which limits the bioavailability of drugs administered systemically, and the short half-life of lysosomal enzymes, hamper the development of effective therapies. Mucopolysaccharidosis type IIIA (MPS IIIA) is an autosomic recessive LSD caused by a deficiency in sulfamidase, a sulfatase involved in the stepwise degradation of glycosaminoglycan (GAG) heparan sulfate. Here, we demonstrate that intracerebrospinal fluid (intra-CSF) administration of serotype 9 adenoassociated viral vectors (AAV9s) encoding sulfamidase corrects both CNS and somatic pathology in MPS IIIA mice. Following vector administration, enzymatic activity increased throughout the brain and in serum, leading to whole body correction of GAG accumulation and lysosomal pathology, normalization of behavioral deficits, and prolonged survival. To test this strategy in a larger animal, we treated beagle dogs using intracisternal or intracerebroventricular delivery. Administration of sulfamidase-encoding AAV9 resulted in transgenic expression throughout the CNS and liver and increased sulfamidase activity in CSF. High-titer serum antibodies against AAV9 only partially blocked CSF-mediated gene transfer to the brains of dogs. Consistently, anti-AAV antibody titers were lower in CSF than in serum collected from healthy and MPS IIIA–affected children. These results support the clinical translation of this approach for the treatment of MPS IIIA and other LSDs with CNS involvement.

Authors

Virginia Haurigot, Sara Marcó, Albert Ribera, Miguel Garcia, Albert Ruzo, Pilar Villacampa, Eduard Ayuso, Sònia Añor, Anna Andaluz, Mercedes Pineda, Gemma García-Fructuoso, Maria Molas, Luca Maggioni, Sergio Muñoz, Sandra Motas, Jesús Ruberte, Federico Mingozzi, Martí Pumarola, Fatima Bosch

Rapamycin extends murine lifespan but has limited effects on aging

• Text
• PDF

Abstract

Aging is a major risk factor for a large number of disorders and functional impairments. Therapeutic targeting of the aging process may therefore represent an innovative strategy in the quest for novel and broadly effective treatments against age-related diseases. The recent report of lifespan extension in mice treated with the FDA-approved mTOR inhibitor rapamycin represented the first demonstration of pharmacological extension of maximal lifespan in mammals. Longevity effects of rapamycin may, however, be due to rapamycin’s effects on specific life-limiting pathologies, such as cancers, and it remains unclear if this compound actually slows the rate of aging in mammals. Here, we present results from a comprehensive, large-scale assessment of a wide range of structural and functional aging phenotypes, which we performed to determine whether rapamycin slows the rate of aging in male C57BL/6J mice. While rapamycin did extend lifespan, it ameliorated few studied aging phenotypes. A subset of aging traits appeared to be rescued by rapamycin. Rapamycin, however, had similar effects on many of these traits in young animals, indicating that these effects were not due to a modulation of aging, but rather related to aging-independent drug effects. Therefore, our data largely dissociate rapamycin’s longevity effects from effects on aging itself.

Authors

Frauke Neff, Diana Flores-Dominguez, Devon P. Ryan, Marion Horsch, Susanne Schröder, Thure Adler, Luciana Caminha Afonso, Juan Antonio Aguilar-Pimentel, Lore Becker, Lillian Garrett, Wolfgang Hans, Moritz M. Hettich, Richard Holtmeier, Sabine M. Hölter, Kristin Moreth, Cornelia Prehn, Oliver Puk, Ildikó Rácz, Birgit Rathkolb, Jan Rozman, Beatrix Naton, Rainer Ordemann, Jerzy Adamski, Johannes Beckers, Raffi Bekeredjian, Dirk H. Busch, Gerhard Ehninger, Jochen Graw, Heinz Höfler, Martin Klingenspor, Thomas Klopstock, Markus Ollert, Jörg Stypmann, Eckhard Wolf, Wolfgang Wurst, Andreas Zimmer, Helmut Fuchs, Valérie Gailus-Durner, Martin Hrabe de Angelis, Dan Ehninger

Developmental differences in IFN signaling affect GATA1s-induced megakaryocyte hyperproliferation

• Text
• PDF

Abstract

About 10% of Down syndrome (DS) infants are born with a transient myeloproliferative disorder (DS-TMD) that spontaneously resolves within the first few months of life. About 20%–30% of these infants subsequently develop acute megakaryoblastic leukemia (DS-AMKL). Somatic mutations leading to the exclusive production of a short GATA1 isoform (GATA1s) occur in all cases of DS-TMD and DS-AMKL. Mice engineered to exclusively produce GATA1s have marked megakaryocytic progenitor (MkP) hyperproliferation during early fetal liver (FL) hematopoiesis, but not during postnatal BM hematopoiesis, mirroring the spontaneous resolution of DS-TMD. The mechanisms that underlie these developmental stage–specific effects are incompletely understood. Here, we report a striking upregulation of type I IFN–responsive gene expression in prospectively isolated mouse BM- versus FL-derived MkPs. Exogenous IFN-α markedly reduced the hyperproliferation FL-derived MkPs of GATA1s mice in vitro. Conversely, deletion of the α/β IFN receptor 1 (Ifnar1) gene or injection of neutralizing IFN-α/β antibodies increased the proliferation of BM-derived MkPs of GATA1s mice beyond the initial postnatal period. We also found that these differences existed in human FL versus BM megakaryocytes and that primary DS-TMD cells expressed type I IFN–responsive genes. We propose that increased type I IFN signaling contributes to the developmental stage–specific effects of GATA1s and possibly the spontaneous resolution of DS-TMD.

Authors

Andrew J. Woo, Karen Wieland, Hui Huang, Thomas E. Akie, Taylor Piers, Jonghwan Kim, Alan B. Cantor

Inactivation of specific β cell transcription factors in type 2 diabetes

• Text
• PDF

Abstract

Type 2 diabetes (T2DM) commonly arises from islet β cell failure and insulin resistance. Here, we examined the sensitivity of key islet-enriched transcription factors to oxidative stress, a condition associated with β cell dysfunction in both type 1 diabetes (T1DM) and T2DM. Hydrogen peroxide treatment of β cell lines induced cytoplasmic translocation of MAFA and NKX6.1. In parallel, the ability of nuclear PDX1 to bind endogenous target gene promoters was also dramatically reduced, whereas the activity of other key β cell transcriptional regulators was unaffected. MAFA levels were reduced, followed by a reduction in NKX6.1 upon development of hyperglycemia in db/db mice, a T2DM model. Transgenic expression of the glutathione peroxidase-1 antioxidant enzyme (GPX1) in db/db islet β cells restored nuclear MAFA, nuclear NKX6.1, and β cell function in vivo. Notably, the selective decrease in MAFA, NKX6.1, and PDX1 expression was found in human T2DM islets. MAFB, a MAFA-related transcription factor expressed in human β cells, was also severely compromised. We propose that MAFA, MAFB, NKX6.1, and PDX1 activity provides a gauge of islet β cell function, with loss of MAFA (and/or MAFB) representing an early indicator of β cell inactivity and the subsequent deficit of more impactful NKX6.1 (and/or PDX1) resulting in overt dysfunction associated with T2DM.

Authors

Shuangli Guo, Chunhua Dai, Min Guo, Brandon Taylor, Jamie S. Harmon, Maike Sander, R. Paul Robertson, Alvin C. Powers, Roland Stein

β-globin gene transfer to human bone marrow for sickle cell disease

• Text
• PDF

Abstract

Autologous hematopoietic stem cell gene therapy is an approach to treating sickle cell disease (SCD) patients that may result in lower morbidity than allogeneic transplantation. We examined the potential of a lentiviral vector (LV) (CCL-βAS3-FB) encoding a human hemoglobin (HBB) gene engineered to impede sickle hemoglobin polymerization (HBBAS3) to transduce human BM CD34⁺ cells from SCD donors and prevent sickling of red blood cells produced by in vitro differentiation. The CCL-βAS3-FB LV transduced BM CD34⁺ cells from either healthy or SCD donors at similar levels, based on quantitative PCR and colony-forming unit progenitor analysis. Consistent expression of HBBAS3 mRNA and HbAS3 protein compromised a fourth of the total β-globin–like transcripts and hemoglobin (Hb) tetramers. Upon deoxygenation, a lower percentage of HBBAS3-transduced red blood cells exhibited sickling compared with mock-transduced cells from sickle donors. Transduced BM CD34⁺ cells were transplanted into immunodeficient mice, and the human cells recovered after 2–3 months were cultured for erythroid differentiation, which showed levels of HBBAS3 mRNA similar to those seen in the CD34⁺ cells that were directly differentiated in vitro. These results demonstrate that the CCL-βAS3-FB LV is capable of efficient transfer and consistent expression of an effective anti-sickling β-globin gene in human SCD BM CD34⁺ progenitor cells, improving physiologic parameters of the resulting red blood cells.

Authors

Zulema Romero, Fabrizia Urbinati, Sabine Geiger, Aaron R. Cooper, Jennifer Wherley, Michael L. Kaufman, Roger P. Hollis, Rafael Ruiz de Assin, Shantha Senadheera, Arineh Sahagian, Xiangyang Jin, Alyse Gellis, Xiaoyan Wang, David Gjertson, Satiro DeOliveira, Pamela Kempert, Sally Shupien, Hisham Abdel-Azim, Mark C. Walters, Herbert J. Meiselman, Rosalinda B. Wenby, Theresa Gruber, Victor Marder, Thomas D. Coates, Donald B. Kohn

Gray platelet syndrome and defective thrombo-inflammation in Nbeal2-deficient mice

• Text
• PDF

Abstract

Platelets are anuclear organelle-rich cell fragments derived from bone marrow megakaryocytes (MKs) that safeguard vascular integrity. The major platelet organelles, α-granules, release proteins that participate in thrombus formation and hemostasis. Proteins stored in α-granules are also thought to play a role in inflammation and wound healing, but their functional significance in vivo is unknown. Mutations in NBEAL2 have been linked to gray platelet syndrome (GPS), a rare bleeding disorder characterized by macrothrombocytopenia, with platelets lacking α-granules. Here we show that Nbeal2-knockout mice display the characteristics of human GPS, with defective α-granule biogenesis in MKs and their absence from platelets. Nbeal2 deficiency did not affect MK differentiation and proplatelet formation in vitro or platelet life span in vivo. Nbeal2-deficient platelets displayed impaired adhesion, aggregation, and coagulant activity ex vivo that translated into defective arterial thrombus formation and protection from thrombo-inflammatory brain infarction following focal cerebral ischemia. In a model of excisional skin wound repair, Nbeal2-deficient mice exhibited impaired development of functional granulation tissue due to severely reduced differentiation of myofibroblasts in the absence of α-granule secretion. This study demonstrates that platelet α-granule constituents are critically required not only for hemostasis but also thrombosis, acute thrombo-inflammatory disease states, and tissue reconstitution after injury.

Authors

Carsten Deppermann, Deya Cherpokova, Paquita Nurden, Jan-Niklas Schulz, Ina Thielmann, Peter Kraft, Timo Vögtle, Christoph Kleinschnitz, Sebastian Dütting, Georg Krohne, Sabine A. Eming, Alan T. Nurden, Beate Eckes, Guido Stoll, David Stegner, Bernhard Nieswandt

Immunoglobulin-like domain containing receptor 1 mediates fat-stimulated cholecystokinin secretion

• Text
• PDF

Abstract

Cholecystokinin (CCK) is a satiety hormone produced by discrete enteroendocrine cells scattered among absorptive cells of the small intestine. CCK is released into blood following a meal; however, the mechanisms inducing hormone secretion are largely unknown. Ingested fat is the major stimulant of CCK secretion. We recently identified a novel member of the lipoprotein remnant receptor family known as immunoglobulin-like domain containing receptor 1 (ILDR1) in intestinal CCK cells and postulated that this receptor conveyed the signal for fat-stimulated CCK secretion. In the intestine, ILDR1 is expressed exclusively in CCK cells. Orogastric administration of fatty acids elevated blood levels of CCK in wild-type mice but not Ildr1-deficient mice, although the CCK secretory response to trypsin inhibitor was retained. The uptake of fluorescently labeled lipoproteins in ILDR1-transfected CHO cells and release of CCK from isolated intestinal cells required a unique combination of fatty acid plus HDL. CCK secretion secondary to ILDR1 activation was associated with increased [Ca²⁺]_i, consistent with regulated hormone release. These findings demonstrate that ILDR1 regulates CCK release through a mechanism dependent on fatty acids and lipoproteins and that absorbed fatty acids regulate gastrointestinal hormone secretion.

Authors

Rashmi Chandra, Yu Wang, Rafiq A. Shahid, Steven R. Vigna, Neil J. Freedman, Rodger A. Liddle

Cross-species malaria immunity induced by chemically attenuated parasites

• Text
• PDF

Abstract

Vaccine development for the blood stages of malaria has focused on the induction of antibodies to parasite surface antigens, most of which are highly polymorphic. An alternate strategy has evolved from observations that low-density infections can induce antibody-independent immunity to different strains. To test this strategy, we treated parasitized red blood cells from the rodent parasite Plasmodium chabaudi with seco-cyclopropyl pyrrolo indole analogs. These drugs irreversibly alkylate parasite DNA, blocking their ability to replicate. After administration in mice, DNA from the vaccine could be detected in the blood for over 110 days and a single vaccination induced profound immunity to different malaria parasite species. Immunity was mediated by CD4⁺ T cells and was dependent on the red blood cell membrane remaining intact. The human parasite, Plasmodium falciparum, could also be attenuated by treatment with seco-cyclopropyl pyrrolo indole analogs. These data demonstrate that vaccination with chemically attenuated parasites induces protective immunity and provide a compelling rationale for testing a blood-stage parasite-based vaccine targeting human Plasmodium species.

Authors

Michael F. Good, Jennifer M. Reiman, I. Bibiana Rodriguez, Koichi Ito, Stephanie K. Yanow, Ibrahim M. El-Deeb, Michael R. Batzloff, Danielle I. Stanisic, Christian Engwerda, Terry Spithill, Stephen L. Hoffman, Moses Lee, Virginia McPhun

Lipocalin 2 deactivates macrophages and worsens pneumococcal pneumonia outcomes

• Text
• PDF

Abstract

Macrophages play a key role in responding to pathogens and initiate an inflammatory response to combat microbe multiplication. Deactivation of macrophages facilitates resolution of the inflammatory response. Deactivated macrophages are characterized by an immunosuppressive phenotype, but the lack of unique markers that can reliably identify these cells explains the poorly defined biological role of this macrophage subset. We identified lipocalin 2 (LCN2) as both a marker of deactivated macrophages and a macrophage deactivator. We show that LCN2 attenuated the early inflammatory response and impaired bacterial clearance, leading to impaired survival of mice suffering from pneumococcal pneumonia. LCN2 induced IL-10 formation by macrophages, skewing macrophage polarization in a STAT3-dependent manner. Pulmonary LCN2 levels were tremendously elevated during bacterial pneumonia in humans, and high LCN2 levels were indicative of a detrimental outcome from pneumonia with Gram-positive bacteria. Our data emphasize the importance of macrophage deactivation for the outcome of pneumococcal infections and highlight the role of LCN2 and IL-10 as determinants of macrophage performance in the respiratory tract.

Authors

Joanna M. Warszawska, Riem Gawish, Omar Sharif, Stefanie Sigel, Bianca Doninger, Karin Lakovits, Ildiko Mesteri, Manfred Nairz, Louis Boon, Alexander Spiel, Valentin Fuhrmann, Birgit Strobl, Mathias Müller, Peter Schenk, Günter Weiss, Sylvia Knapp

Cerebrovascular degradation of TRKB by MMP9 in the diabetic brain

• Text
• PDF

Abstract

Diabetes elevates the risk for neurological diseases, but little is known about the underlying mechanisms. Brain-derived neurotrophic factor (BDNF) is secreted by microvascular endothelial cells (ECs) in the brain, functioning as a neuroprotectant through the activation of the neurotrophic tyrosine kinase receptor TRKB. In a rat model of streptozotocin-induced hyperglycemia, we found that endothelial activation of MMP9 altered TRKB-dependent trophic pathways by degrading TRKB in neurons. Treatment of brain microvascular ECs with advanced glycation endproducts (AGE), a metabolite commonly elevated in diabetic patients, increased MMP9 activation, similar to in vivo findings. Recombinant human MMP9 degraded the TRKB ectodomain in primary neuronal cultures, suggesting that TRKB could be a substrate for MMP9 proteolysis. Consequently, AGE-conditioned endothelial media with elevated MMP9 activity degraded the TRKB ectodomain and simultaneously disrupted the ability of endothelium to protect neurons against hypoxic injury. Our findings demonstrate that neuronal TRKB trophic function is ablated by MMP9-mediated degradation in the diabetic brain, disrupting cerebrovascular trophic coupling and leaving the brain vulnerable to injury.

Authors

Deepti Navaratna, Xiang Fan, Wendy Leung, Josephine Lok, Shuzhen Guo, Changhong Xing, Xiaoying Wang, Eng H. Lo

Atrial natriuretic peptide is negatively regulated by microRNA-425

• Text
• PDF

Abstract

Numerous common genetic variants have been linked to blood pressure, but no underlying mechanism has been elucidated. Population studies have revealed that the variant rs5068 (A/G) in the 3′ untranslated region of NPPA, the gene encoding atrial natriuretic peptide (ANP), is associated with blood pressure. We selected individuals on the basis of rs5068 genotype (AG vs. AA) and fed them a low- or high-salt diet for 1 week, after which they were challenged with an intravenous saline infusion. On both diets, before and after saline administration, ANP levels were up to 50% higher in AG individuals than in AA individuals, a difference comparable to the changes induced by high-salt diet or saline infusion. In contrast, B-type natriuretic peptide levels did not differ by rs5068 genotype. We identified a microRNA, miR-425, that is expressed in human atria and ventricles and is predicted to bind the sequence spanning rs5068 for the A, but not the G, allele. miR-425 silenced NPPA mRNA in an allele-specific manner, with the G allele conferring resistance to miR-425. This study identifies miR-425 as a regulator of ANP production, raising the possibility that miR-425 antagonists could be used to treat disorders of salt overload, including hypertension and heart failure.

Authors

Pankaj Arora, Connie Wu, Abigail May Khan, Donald B. Bloch, Brandi N. Davis-Dusenbery, Anahita Ghorbani, Ester Spagnolli, Andrew Martinez, Allicia Ryan, Laurel T. Tainsh, Samuel Kim, Jian Rong, Tianxiao Huan, Jane E. Freedman, Daniel Levy, Karen K. Miller, Akiko Hata, Federica del Monte, Sara Vandenwijngaert, Melissa Swinnen, Stefan Janssens, Tara M. Holmes, Emmanuel S. Buys, Kenneth D. Bloch, Christopher Newton-Cheh, Thomas J. Wang

IL-12p70–producing patient DC vaccine elicits Tc1-polarized immunity

• Text
• PDF

Abstract

Background. Systemic administration of IL-12p70 has demonstrated clinical activity in cancer patients, but dose-limiting toxicities have hindered its incorporation in vaccine formulations. Here, we report on the immunological and clinical outcomes upon vaccination with CD40L/IFN-γ–matured, IL-12p70–producing DCs.

Methods. 7 HLA-A*0201⁺ newly diagnosed stage IV melanoma patients were immunized against the gp100 melanoma antigen using autologous peptide-pulsed, CD40L/IFN-γ–matured DCs. PBMCs were taken weekly for immune monitoring by tetramer analysis and functional assays. CT imaging was performed at baseline, week 9, and week 18 for clinical assessment using RECIST.

Results. 6 of 7 treated patients developed sustained T cell immunity to all 3 melanoma gp100 antigen–derived peptides. 3 of the 6 immunological responders developed confirmed clinical responses (1 complete remission >4 years, 2 partial response). Importantly, DC vaccine–derived IL-12p70 levels positively correlated with time to progression (P = 0.019, log-rank), as did T-cytotoxic 1 (Tc1) immunity, as assessed by IFN-γ/IL-13 and IFN-γ/IL-5 ratios (P = 0.035 and P = 0.030, respectively, log-rank). In contrast, a pathway-specific defect in IL-12p35 transcription was identified upon CD40L/IFN-γ activation in clinical nonresponder patient DCs, and gp100-specific T cells from these patients displayed a Tc2 phenotype. Incorporation of TLR3 and TLR8 agonists into the CD40L/IFN-γ activation protocol corrected the IL-12p70 production defect in DCs derived from clinical nonresponder patients.

Conclusion. These findings underscore the essential role of IL-12p70 in the development of therapeutic type 1 antigen–specific CD8⁺ T cell immunity in humans with cancer.

Trial registration. Clinicaltrials.gov NCT00683670.

Funding. Barnes-Jewish Hospital Foundation, Siteman Cancer Frontier Fund, Washington University/JNJ Translational Medicine Award, and NCI (P30 CA91842).

Authors

Beatriz M. Carreno, Michelle Becker-Hapak, Alexander Huang, Megan Chan, Amer Alyasiry, Wen-Rong Lie, Rebecca L. Aft, Lynn A. Cornelius, Kathryn M. Trinkaus, Gerald P. Linette

Cold acclimation recruits human brown fat and increases nonshivering thermogenesis

• Text
• PDF

Abstract

In recent years, it has been shown that humans have active brown adipose tissue (BAT) depots, raising the question of whether activation and recruitment of BAT can be a target to counterbalance the current obesity pandemic. Here, we show that a 10-day cold acclimation protocol in humans increases BAT activity in parallel with an increase in nonshivering thermogenesis (NST). No sex differences in BAT presence and activity were found either before or after cold acclimation. Respiration measurements in permeabilized fibers and isolated mitochondria revealed no significant contribution of skeletal muscle mitochondrial uncoupling to the increased NST. Based on cell-specific markers and on uncoupling protein-1 (characteristic of both BAT and beige/brite cells), this study did not show “browning” of abdominal subcutaneous white adipose tissue upon cold acclimation. The observed physiological acclimation is in line with the subjective changes in temperature sensation; upon cold acclimation, the subjects judged the environment warmer, felt more comfortable in the cold, and reported less shivering. The combined results suggest that a variable indoor environment with frequent cold exposures might be an acceptable and economic manner to increase energy expenditure and may contribute to counteracting the current obesity epidemic.

Authors

Anouk A.J.J. van der Lans, Joris Hoeks, Boudewijn Brans, Guy H.E.J. Vijgen, Mariëlle G.W. Visser, Maarten J. Vosselman, Jan Hansen, Johanna A. Jörgensen, Jun Wu, Felix M. Mottaghy, Patrick Schrauwen, Wouter D. van Marken Lichtenbelt

Recruited brown adipose tissue as an antiobesity agent in humans

• Text
• PDF

Abstract

Brown adipose tissue (BAT) burns fat to produce heat when the body is exposed to cold and plays a role in energy metabolism. Using fluorodeoxyglucose-positron emission tomography and computed tomography, we previously reported that BAT decreases with age and thereby accelerates age-related accumulation of body fat in humans. Thus, the recruitment of BAT may be effective for body fat reduction. In this study, we examined the effects of repeated stimulation by cold and capsinoids (nonpungent capsaicin analogs) in healthy human subjects with low BAT activity. Acute cold exposure at 19°C for 2 hours increased energy expenditure (EE). Cold-induced increments of EE (CIT) strongly correlated with BAT activity independently of age and fat-free mass. Daily 2-hour cold exposure at 17°C for 6 weeks resulted in a parallel increase in BAT activity and CIT and a concomitant decrease in body fat mass. Changes in BAT activity and body fat mass were negatively correlated. Similarly, daily ingestion of capsinoids for 6 weeks increased CIT. These results demonstrate that human BAT can be recruited even in individuals with decreased BAT activity, thereby contributing to body fat reduction.

Authors

Takeshi Yoneshiro, Sayuri Aita, Mami Matsushita, Takashi Kayahara, Toshimitsu Kameya, Yuko Kawai, Toshihiko Iwanaga, Masayuki Saito

FOXOs attenuate bone formation by suppressing Wnt signaling

• Text
• PDF

Abstract

Wnt/β-catenin/TCF signaling stimulates bone formation and suppresses adipogenesis. The hallmarks of skeletal involution with age, on the other hand, are decreased bone formation and increased bone marrow adiposity. These changes are associated with increased oxidative stress and decreased growth factor production, which activate members of the FOXO family of transcription factors. FOXOs in turn attenuate Wnt/β-catenin signaling by diverting β-catenin from TCF- to FOXO-mediated transcription. We show herein that mice lacking Foxo1, -3, and -4 in bipotential progenitors of osteoblast and adipocytes (expressing Osterix1) exhibited increased osteoblast number and high bone mass that was maintained in old age as well as decreased adiposity in the aged bone marrow. The increased bone mass in the Foxo-deficient mice was accounted for by increased proliferation of osteoprogenitor cells and bone formation resulting from upregulation of Wnt/β-catenin signaling and cyclin D1 expression, but not changes in redox balance. Consistent with this mechanism, β-catenin deletion in Foxo null cells abrogated both the increased cyclin D1 expression and proliferation. The elucidation of a restraining effect of FOXOs on Wnt signaling in bipotential progenitors suggests that FOXO activation by accumulation of age-associated cellular stressors may be a seminal pathogenetic mechanism in the development of involutional osteoporosis.

Authors

Srividhya Iyer, Elena Ambrogini, Shoshana M. Bartell, Li Han, Paula K. Roberson, Rafael de Cabo, Robert L. Jilka, Robert S. Weinstein, Charles A. O’Brien, Stavros C. Manolagas, Maria Almeida

The nucleotide sugar UDP-glucose mobilizes long-term repopulating primitive hematopoietic cells

• Text
• PDF

Abstract

Hematopoietic stem progenitor cells (HSPCs) are present in very small numbers in the circulating blood in steady-state conditions. In response to stress or injury, HSPCs are primed to migrate out of their niche to peripheral blood. Mobilized HSPCs are now commonly used as stem cell sources due to faster engraftment and reduced risk of posttransplant infection. In this study, we demonstrated that a nucleotide sugar, UDP-glucose, which is released into extracellular fluids in response to stress, mediates HSPC mobilization. UDP-glucose–mobilized cells possessed the capacity to achieve long-term repopulation in lethally irradiated animals and the ability to differentiate into multi-lineage blood cells. Compared with G-CSF–mobilized cells, UDP-glucose–mobilized cells preferentially supported long-term repopulation and exhibited lymphoid-biased differentiation, suggesting that UDP-glucose triggers the mobilization of functionally distinct subsets of HSPCs. Furthermore, co-administration of UDP-glucose and G-CSF led to greater HSPC mobilization than G-CSF alone. Administration of the antioxidant agent NAC significantly reduced UDP-glucose–induced mobilization, coinciding with a reduction in RANKL and osteoclastogenesis. These findings provide direct evidence demonstrating a potential role for UDP-glucose in HSPC mobilization and may provide an attractive strategy to improve the yield of stem cells in poor-mobilizing allogeneic or autologous donors.

Authors

Sungho Kook, Joonseok Cho, Sean Bong Lee, Byeong-Chel Lee

Angiopoietin 2 mediates microvascular and hemodynamic alterations in sepsis

• Text
• PDF

Abstract

Septic shock is characterized by increased vascular permeability and hypotension despite increased cardiac output. Numerous vasoactive cytokines are upregulated during sepsis, including angiopoietin 2 (ANG2), which increases vascular permeability. Here we report that mice engineered to inducibly overexpress ANG2 in the endothelium developed sepsis-like hemodynamic alterations, including systemic hypotension, increased cardiac output, and dilatory cardiomyopathy. Conversely, mice with cardiomyocyte-restricted ANG2 overexpression failed to develop hemodynamic alterations. Interestingly, the hemodynamic alterations associated with endothelial-specific overexpression of ANG2 and the loss of capillary-associated pericytes were reversed by intravenous injections of adeno-associated viruses (AAVs) transducing cDNA for angiopoietin 1, a TIE2 ligand that antagonizes ANG2, or AAVs encoding PDGFB, a chemoattractant for pericytes. To confirm the role of ANG2 in sepsis, we i.p. injected LPS into C57BL/6J mice, which rapidly developed hypotension, acute pericyte loss, and increased vascular permeability. Importantly, ANG2 antibody treatment attenuated LPS-induced hemodynamic alterations and reduced the mortality rate at 36 hours from 95% to 61%. These data indicate that ANG2-mediated microvascular disintegration contributes to septic shock and that inhibition of the ANG2/TIE2 interaction during sepsis is a potential therapeutic target.

Authors

Tilman Ziegler, Jan Horstkotte, Claudia Schwab, Vanessa Pfetsch, Karolina Weinmann, Steffen Dietzel, Ina Rohwedder, Rabea Hinkel, Lisa Gross, Seungmin Lee, Junhao Hu, Oliver Soehnlein, Wolfgang M. Franz, Markus Sperandio, Ulrich Pohl, Markus Thomas, Christian Weber, Hellmut G. Augustin, Reinhard Fässler, Urban Deutsch, Christian Kupatt

Neutrophil extracellular traps sequester circulating tumor cells and promote metastasis

• Text
• PDF

Abstract

The majority of patients with cancer undergo at least one surgical procedure as part of their treatment. Severe postsurgical infection is associated with adverse oncologic outcomes; however, the mechanisms underlying this phenomenon are unclear. Emerging evidence suggests that neutrophils, which function as the first line of defense during infections, facilitate cancer progression. Neutrophil extracellular traps (NETs) are extracellular neutrophil-derived DNA webs released in response to inflammatory cues that trap and kill invading pathogens. The role of NETs in cancer progression is entirely unknown. We report that circulating tumor cells become trapped within NETs in vitro under static and dynamic conditions. In a murine model of infection using cecal ligation and puncture, we demonstrated microvascular NET deposition and consequent trapping of circulating lung carcinoma cells within DNA webs. NET trapping was associated with increased formation of hepatic micrometastases at 48 hours and gross metastatic disease burden at 2 weeks following tumor cell injection. These effects were abrogated by NET inhibition with DNAse or a neutrophil elastase inhibitor. These findings implicate NETs in the process of cancer metastasis in the context of systemic infection and identify NETs as potential therapeutic targets.

Authors

Jonathan Cools-Lartigue, Jonathan Spicer, Braedon McDonald, Stephen Gowing, Simon Chow, Betty Giannias, France Bourdeau, Paul Kubes, Lorenzo Ferri

Metastasis-associated PRL-3 induces EGFR activation and addiction in cancer cells

• Text
• PDF

Abstract

Metastasis-associated phosphatase of regenerating liver-3 (PRL-3) has pleiotropic effects in driving cancer progression, yet the signaling mechanisms of PRL-3 are still not fully understood. Here, we provide evidence for PRL-3–induced hyperactivation of EGFR and its downstream signaling cascades in multiple human cancer cell lines. Mechanistically, PRL-3–induced activation of EGFR was attributed primarily to transcriptional downregulation of protein tyrosine phosphatase 1B (PTP1B), an inhibitory phosphatase for EGFR. Functionally, PRL-3–induced hyperactivation of EGFR correlated with increased cell growth, promigratory characteristics, and tumorigenicity. Moreover, PRL-3 induced cellular addiction to EGFR signaling, as evidenced by the pronounced reversion of these oncogenic attributes upon EGFR-specific inhibition. Of clinical significance, we verified elevated PRL-3 expression as a predictive marker for favorable therapeutic response in a heterogeneous colorectal cancer (CRC) patient cohort treated with the clinically approved anti-EGFR antibody cetuximab. The identification of PRL-3–driven EGFR hyperactivation and consequential addiction to EGFR signaling opens new avenues for inhibiting PRL-3–driven cancer progression. We propose that elevated PRL-3 expression is an important clinical predictive biomarker for favorable anti-EGFR cancer therapy.

Authors

Abdul Qader Omer Al-aidaroos, Hiu Fung Yuen, Ke Guo, Shu Dong Zhang, Tae-Hoon Chung, Wee Joo Chng, Qi Zeng

Macrophages regulate corpus luteum development during embryo implantation in mice

• Text
• PDF

Abstract

Macrophages are prominent in the uterus and ovary at conception. Here we utilize the Cd11b-Dtr mouse model of acute macrophage depletion to define the essential role of macrophages in early pregnancy. Macrophage depletion after conception caused embryo implantation arrest associated with diminished plasma progesterone and poor uterine receptivity. Implantation failure was alleviated by administration of bone marrow–derived CD11b⁺F4/80⁺ monocytes/macrophages. In the ovaries of macrophage-depleted mice, corpora lutea were profoundly abnormal, with elevated Ptgs2, Hif1a, and other inflammation and apoptosis genes and with diminished expression of steroidogenesis genes Star, Cyp11a1, and Hsd3b1. Infertility was rescued by exogenous progesterone, which confirmed that uterine refractoriness was fully attributable to the underlying luteal defect. In normally developing corpora lutea, macrophages were intimately juxtaposed with endothelial cells and expressed the proangiogenic marker TIE2. After macrophage depletion, substantial disruption of the luteal microvascular network occurred and was associated with altered ovarian expression of genes that encode vascular endothelial growth factors. These data indicate a critical role for macrophages in supporting the extensive vascular network required for corpus luteum integrity and production of progesterone essential for establishing pregnancy. Our findings raise the prospect that disruption of macrophage-endothelial cell interactions underpinning corpus luteum development contributes to infertility in women in whom luteal insufficiency is implicated.

Authors

Alison S. Care, Kerrilyn R. Diener, Melinda J. Jasper, Hannah M. Brown, Wendy V. Ingman, Sarah A. Robertson

LXRβ/estrogen receptor-α signaling in lipid rafts preserves endothelial integrity

• Text
• PDF

Abstract

Liver X receptors (LXR) are stimulated by cholesterol-derived oxysterols and serve as transcription factors to regulate gene expression in response to alterations in cholesterol. In the present study, we investigated the role of LXRs in vascular endothelial cells (ECs) and discovered that LXRβ has nonnuclear function and stimulates EC migration by activating endothelial NOS (eNOS). This process is mediated by estrogen receptor-α (ERα). LXR activation promoted the direct binding of LXRβ to the ligand-binding domain of ERα and initiated an extranuclear signaling cascade that requires ERα Ser118 phosphorylation by PI3K/AKT. Further studies revealed that LXRβ and ERα are colocalized and functionally coupled in EC plasma membrane caveolae/lipid rafts. In isolated aortic rings, LXR activation of NOS caused relaxation, while in mice, LXR activation stimulated carotid artery reendothelialization via LXRβ- and ERα-dependent processes. These studies demonstrate that LXRβ has nonnuclear function in EC caveolae/lipid rafts that entails crosstalk with ERα, which promotes NO production and maintains endothelial monolayer integrity in vivo.

Authors

Tomonori Ishikawa, Ivan S. Yuhanna, Junko Umetani, Wan-Ru Lee, Kenneth S. Korach, Philip W. Shaul, Michihisa Umetani

Collagen VII plays a dual role in wound healing

• Text
• PDF

Abstract

Although a host of intracellular signals is known to contribute to wound healing, the role of the cell microenvironment in tissue repair remains elusive. Here we employed 2 different mouse models of genetic skin fragility to assess the role of the basement membrane protein collagen VII (COL7A1) in wound healing. COL7A1 secures the attachment of the epidermis to the dermis, and its mutations cause a human skin fragility disorder coined recessive dystrophic epidermolysis bullosa (RDEB) that is associated with a constant wound burden. We show that COL7A1 is instrumental for skin wound closure by 2 interconnected mechanisms. First, COL7A1 was required for re-epithelialization through organization of laminin-332 at the dermal-epidermal junction. Its loss perturbs laminin-332 organization during wound healing, which in turn abrogates strictly polarized expression of integrin α6β4 in basal keratinocytes and negatively impacts the laminin-332/integrin α6β4 signaling axis guiding keratinocyte migration. Second, COL7A1 supported dermal fibroblast migration and regulates their cytokine production in the granulation tissue. These findings, which were validated in human wounds, identify COL7A1 as a critical player in physiological wound healing in humans and mice and may facilitate development of therapeutic strategies not only for RDEB, but also for other chronic wounds.

Authors

Alexander Nyström, Daniela Velati, Venugopal R. Mittapalli, Anja Fritsch, Johannes S. Kern, Leena Bruckner-Tuderman

Medullary thymic epithelial cell depletion leads to autoimmune hepatitis

• Text
• PDF

Abstract

TRAF6, an E3 ubiquitin protein ligase, plays a critical role in T cell tolerance by regulating medullary thymic epithelial cell (mTEC) development. mTECs regulate T cell tolerance by ectopically expressing self-antigens and eliminating autoreactive T cells in the thymus. Here we show that mice with mTEC depletion due to conditional deletion of Traf6 expression in murine thymic epithelial cells (Traf6ΔTEC mice) showed a surprisingly narrow spectrum of autoimmunity affecting the liver. The liver inflammation in Traf6ΔTEC mice exhibited all the histological and immunological characteristics of human autoimmune hepatitis (AIH). The role of T cells in AIH establishment was supported by intrahepatic T cell population changes and AIH development after transfer of liver T cells into immunodeficient mice. Despite a 50% reduction in natural Treg thymic output, peripheral tolerance in Traf6ΔTEC mice was normal, whereas compensatory T regulatory mechanisms were evident in the liver of these animals. These data indicate that mTECs exert a cell-autonomous role in central T cell tolerance and organ-specific autoimmunity, but play a redundant role in peripheral tolerance. These findings also demonstrate that Traf6ΔTEC mice are a relevant model with which to study the pathophysiology of AIH, as well as autoantigen-specific T cell responses and regulatory mechanisms underlying this disease.

Authors

Anthony J. Bonito, Costica Aloman, M. Isabel Fiel, Nichole M. Danzl, Sungwon Cha, Erica G. Weinstein, Seihwan Jeong, Yongwon Choi, Matthew C. Walsh, Konstantina Alexandropoulos

FGFR2 signaling underlies p63 oncogenic function in squamous cell carcinoma

• Text
• PDF

Abstract

Oncogenic transcription factors drive many human cancers, yet identifying and therapeutically targeting the resulting deregulated pathways has proven difficult. Squamous cell carcinoma (SCC) is a common and lethal human cancer, and relatively little progress has been made in improving outcomes for SCC due to a poor understanding of its underlying molecular pathogenesis. While SCCs typically lack somatic oncogene-activating mutations, they exhibit frequent overexpression of the p53-related transcription factor p63. We developed an in vivo murine tumor model to investigate the function and key transcriptional programs of p63 in SCC. Here, we show that established SCCs are exquisitely dependent on p63, as acute genetic ablation of p63 in advanced, invasive SCC induced rapid and dramatic apoptosis and tumor regression. In vivo genome-wide gene expression analysis identified a tumor-survival program involving p63-regulated FGFR2 signaling that was activated by ligand emanating from abundant tumor-associated stroma. Correspondingly, we demonstrate the therapeutic efficacy of extinguishing this signaling axis in endogenous SCCs using the clinical FGFR2 inhibitor AZD4547. Collectively, these results reveal an unanticipated role for p63-driven paracrine FGFR2 signaling as an addicting pathway in human cancer and suggest a new approach for the treatment of SCC.

Authors

Matthew R. Ramsey, Catherine Wilson, Benjamin Ory, S. Michael Rothenberg, William Faquin, Alea A. Mills, Leif W. Ellisen

A link between FTO, ghrelin, and impaired brain food-cue responsivity

• Text
• PDF

Abstract

Polymorphisms in the fat mass and obesity-associated gene (FTO) are associated with human obesity and obesity-prone behaviors, including increased food intake and a preference for energy-dense foods. FTO demethylates N⁶-methyladenosine, a potential regulatory RNA modification, but the mechanisms by which FTO predisposes humans to obesity remain unclear. In adiposity-matched, normal-weight humans, we showed that subjects homozygous for the FTO “obesity-risk” rs9939609 A allele have dysregulated circulating levels of the orexigenic hormone acyl-ghrelin and attenuated postprandial appetite reduction. Using functional MRI (fMRI) in normal-weight AA and TT humans, we found that the FTO genotype modulates the neural responses to food images in homeostatic and brain reward regions. Furthermore, AA and TT subjects exhibited divergent neural responsiveness to circulating acyl-ghrelin within brain regions that regulate appetite, reward processing, and incentive motivation. In cell models, FTO overexpression reduced ghrelin mRNA N⁶-methyladenosine methylation, concomitantly increasing ghrelin mRNA and peptide levels. Furthermore, peripheral blood cells from AA human subjects exhibited increased FTO mRNA, reduced ghrelin mRNA N⁶-methyladenosine methylation, and increased ghrelin mRNA abundance compared with TT subjects. Our findings show that FTO regulates ghrelin, a key mediator of ingestive behavior, and offer insight into how FTO obesity-risk alleles predispose to increased energy intake and obesity in humans.

Authors

Efthimia Karra, Owen G. O’Daly, Agharul I. Choudhury, Ahmed Yousseif, Steven Millership, Marianne T. Neary, William R. Scott, Keval Chandarana, Sean Manning, Martin E. Hess, Hiroshi Iwakura, Takashi Akamizu, Queensta Millet, Cigdem Gelegen, Megan E. Drew, Sofia Rahman, Julian J. Emmanuel, Steven C.R. Williams, Ulrich U. Rüther, Jens C. Brüning, Dominic J. Withers, Fernando O. Zelaya, Rachel L. Batterham

Epigenetic changes induced by adenosine augmentation therapy prevent epileptogenesis

• Text
• PDF

Abstract

Epigenetic modifications, including changes in DNA methylation, lead to altered gene expression and thus may underlie epileptogenesis via induction of permanent changes in neuronal excitability. Therapies that could inhibit or reverse these changes may be highly effective in halting disease progression. Here we identify an epigenetic function of the brain’s endogenous anticonvulsant adenosine, showing that this compound induces hypomethylation of DNA via biochemical interference with the transmethylation pathway. We show that inhibition of DNA methylation inhibited epileptogenesis in multiple seizure models. Using a rat model of temporal lobe epilepsy, we identified an increase in hippocampal DNA methylation, which correlates with increased DNA methyltransferase activity, disruption of adenosine homeostasis, and spontaneous recurrent seizures. Finally, we used bioengineered silk implants to deliver a defined dose of adenosine over 10 days to the brains of epileptic rats. This transient therapeutic intervention reversed the DNA hypermethylation seen in the epileptic brain, inhibited sprouting of mossy fibers in the hippocampus, and prevented the progression of epilepsy for at least 3 months. These data demonstrate that pathological changes in DNA methylation homeostasis may underlie epileptogenesis and reversal of these epigenetic changes with adenosine augmentation therapy may halt disease progression.

Authors

Rebecca L. Williams-Karnesky, Ursula S. Sandau, Theresa A. Lusardi, Nikki K. Lytle, Joseph M. Farrell, Eleanor M. Pritchard, David L. Kaplan, Detlev Boison

Transcription factor EGR1 directs tendon differentiation and promotes tendon repair

• Text
• PDF

Abstract

Tendon formation and repair rely on specific combinations of transcription factors, growth factors, and mechanical parameters that regulate the production and spatial organization of type I collagen. Here, we investigated the function of the zinc finger transcription factor EGR1 in tendon formation, healing, and repair using rodent animal models and mesenchymal stem cells (MSCs). Adult tendons of Egr1^–/– mice displayed a deficiency in the expression of tendon genes, including Scx, Col1a1, and Col1a2, and were mechanically weaker compared with their WT littermates. EGR1 was recruited to the Col1a1 and Col2a1 promoters in postnatal mouse tendons in vivo. Egr1 was required for the normal gene response following tendon injury in a mouse model of Achilles tendon healing. Forced Egr1 expression programmed MSCs toward the tendon lineage and promoted the formation of in vitro–engineered tendons from MSCs. The application of EGR1-producing MSCs increased the formation of tendon-like tissues in a rat model of Achilles tendon injury. We provide evidence that the ability of EGR1 to promote tendon differentiation is partially mediated by TGF-β2. This study demonstrates EGR1 involvement in adult tendon formation, healing, and repair and identifies Egr1 as a putative target in tendon repair strategies.

Authors

Marie-Justine Guerquin, Benjamin Charvet, Geoffroy Nourissat, Emmanuelle Havis, Olivier Ronsin, Marie-Ange Bonnin, Mathilde Ruggiu, Isabel Olivera-Martinez, Nicolas Robert, Yinhui Lu, Karl E. Kadler, Tristan Baumberger, Levon Doursounian, Francis Berenbaum, Delphine Duprez

Inner ear supporting cells protect hair cells by secreting HSP70

• Text
• PDF

Abstract

Mechanosensory hair cells are the receptor cells of hearing and balance. Hair cells are sensitive to death from exposure to therapeutic drugs with ototoxic side effects, including aminoglycoside antibiotics and cisplatin. We recently showed that the induction of heat shock protein 70 (HSP70) inhibits ototoxic drug–induced hair cell death. Here, we examined the mechanisms underlying the protective effect of HSP70. In response to heat shock, HSP70 was induced in glia-like supporting cells but not in hair cells. Adenovirus-mediated infection of supporting cells with Hsp70 inhibited hair cell death. Coculture with heat-shocked utricles protected nonheat-shocked utricles against hair cell death. When heat-shocked utricles from Hsp70^–/– mice were used in cocultures, protection was abolished in both the heat-shocked utricles and the nonheat-shocked utricles. HSP70 was detected by ELISA in the media surrounding heat-shocked utricles, and depletion of HSP70 from the media abolished the protective effect of heat shock, suggesting that HSP70 is secreted by supporting cells. Together our data indicate that supporting cells mediate the protective effect of HSP70 against hair cell death, and they suggest a major role for supporting cells in determining the fate of hair cells exposed to stress.

Authors

Lindsey A. May, Inga I. Kramarenko, Carlene S. Brandon, Christina Voelkel-Johnson, Soumen Roy, Kristy Truong, Shimon P. Francis, Elyssa L. Monzack, Fu-Shing Lee, Lisa L. Cunningham

CHIP protects against cardiac pressure overload through regulation of AMPK

• Text
• PDF

Abstract

Protein quality control and metabolic homeostasis are integral to maintaining cardiac function during stress; however, little is known about if or how these systems interact. Here we demonstrate that C terminus of HSC70-interacting protein (CHIP), a regulator of protein quality control, influences the metabolic response to pressure overload by direct regulation of the catalytic α subunit of AMPK. Induction of cardiac pressure overload in Chip^–/– mice resulted in robust hypertrophy and decreased cardiac function and energy generation stemming from a failure to activate AMPK. Mechanistically, CHIP promoted LKB1-mediated phosphorylation of AMPK, increased the specific activity of AMPK, and was necessary and sufficient for stress-dependent activation of AMPK. CHIP-dependent effects on AMPK activity were accompanied by conformational changes specific to the α subunit, both in vitro and in vivo, identifying AMPK as the first physiological substrate for CHIP chaperone activity and establishing a link between cardiac proteolytic and metabolic pathways.

Authors

Jonathan C. Schisler, Carrie E. Rubel, Chunlian Zhang, Pamela Lockyer, Douglas M. Cyr, Cam Patterson

FK506 activates BMPR2, rescues endothelial dysfunction, and reverses pulmonary hypertension

• Text
• PDF

Abstract

Dysfunctional bone morphogenetic protein receptor-2 (BMPR2) signaling is implicated in the pathogenesis of pulmonary arterial hypertension (PAH). We used a transcriptional high-throughput luciferase reporter assay to screen 3,756 FDA-approved drugs and bioactive compounds for induction of BMPR2 signaling. The best response was achieved with FK506 (tacrolimus), via a dual mechanism of action as a calcineurin inhibitor that also binds FK-binding protein-12 (FKBP12), a repressor of BMP signaling. FK506 released FKBP12 from type I receptors activin receptor-like kinase 1 (ALK1), ALK2, and ALK3 and activated downstream SMAD1/5 and MAPK signaling and ID1 gene regulation in a manner superior to the calcineurin inhibitor cyclosporine and the FKBP12 ligand rapamycin. In pulmonary artery endothelial cells (ECs) from patients with idiopathic PAH, low-dose FK506 reversed dysfunctional BMPR2 signaling. In mice with conditional Bmpr2 deletion in ECs, low-dose FK506 prevented exaggerated chronic hypoxic PAH associated with induction of EC targets of BMP signaling, such as apelin. Low-dose FK506 also reversed severe PAH in rats with medial hypertrophy following monocrotaline and in rats with neointima formation following VEGF receptor blockade and chronic hypoxia. Our studies indicate that low-dose FK506 could be useful in the treatment of PAH.

Authors

Edda Spiekerkoetter, Xuefei Tian, Jie Cai, Rachel K. Hopper, Deepti Sudheendra, Caiyun G. Li, Nesrine El-Bizri, Hirofumi Sawada, Roxanna Haghighat, Roshelle Chan, Leila Haghighat, Vinicio de Jesus Perez, Lingli Wang, Sushma Reddy, Mingming Zhao, Daniel Bernstein, David E. Solow-Cordero, Philip A. Beachy, Thomas J. Wandless, Peter ten Dijke, Marlene Rabinovitch

Isocitrate ameliorates anemia by suppressing the erythroid iron restriction response

• Text
• PDF

Abstract

The unique sensitivity of early red cell progenitors to iron deprivation, known as the erythroid iron restriction response, serves as a basis for human anemias globally. This response impairs erythropoietin-driven erythropoiesis and underlies erythropoietic repression in iron deficiency anemia. Mechanistically, the erythroid iron restriction response results from inactivation of aconitase enzymes and can be suppressed by providing the aconitase product isocitrate. Recent studies have implicated the erythroid iron restriction response in anemia of chronic disease and inflammation (ACDI), offering new therapeutic avenues for a major clinical problem; however, inflammatory signals may also directly repress erythropoiesis in ACDI. Here, we show that suppression of the erythroid iron restriction response by isocitrate administration corrected anemia and erythropoietic defects in rats with ACDI. In vitro studies demonstrated that erythroid repression by inflammatory signaling is potently modulated by the erythroid iron restriction response in a kinase-dependent pathway involving induction of the erythroid-inhibitory transcription factor PU.1. These results reveal the integration of iron and inflammatory inputs in a therapeutically tractable erythropoietic regulatory circuit.

Authors

Chanté L. Richardson, Lorrie L. Delehanty, Grant C. Bullock, Claudia M. Rival, Kenneth S. Tung, Donald L. Kimpel, Sara Gardenghi, Stefano Rivella, Adam N. Goldfarb

Somitic disruption of GNAS in chick embryos mimics progressive osseous heteroplasia

• Text
• PDF

Abstract

Progressive osseous heteroplasia (POH) is a rare developmental disorder of heterotopic ossification (HO) caused by heterozygous inactivating germline mutations in the paternal allele of the GNAS gene. Interestingly, POH lesions have a bewildering mosaic distribution. Using clinical, radiographic, and photographic documentation, we found that most of the 12 individuals studied had a lesional bias toward one side or the other, even showing exclusive sidedness. Most strikingly, all had a dermomyotomal distribution of HO lesions. We hypothesized that somatic mutations in a progenitor cell of somitic origin may act on a background of germline haploinsufficiency to cause loss of heterozygosity at the GNAS locus and lead to the unilateral distribution of POH lesions. Taking advantage of the chick system, we examined our hypothesis by mimicking loss of heterozygosity of GNAS expression using dominant-negative GNAS that was introduced into a subset of chick somites, the progenitors that give rise to dermis and muscle. We observed rapid ectopic cartilage and bone induction at the axial and lateral positions in a unilateral distribution corresponding to the injected somites, which suggests that blocking GNAS activity in a targeted population of progenitor cells can lead to mosaic ectopic ossification reminiscent of that seen in POH.

Authors

Dana M. Cairns, Robert J. Pignolo, Tomoya Uchimura, Tracy A. Brennan, Carter M. Lindborg, Meiqi Xu, Frederick S. Kaplan, Eileen M. Shore, Li Zeng

BM mesenchymal stromal cell–derived exosomes facilitate multiple myeloma progression

• Text
• PDF

Abstract

Authors

Aldo M. Roccaro, Antonio Sacco, Patricia Maiso, Abdel Kareem Azab, Yu-Tzu Tai, Michaela Reagan, Feda Azab, Ludmila M. Flores, Federico Campigotto, Edie Weller, Kenneth C. Anderson, David T. Scadden, Irene M. Ghobrial

Rare variants in single-minded 1 (SIM1) are associated with severe obesity

• Text
• PDF

Abstract

Authors

Shwetha Ramachandrappa, Anne Raimondo, Anna M.G. Cali, Julia M. Keogh, Elana Henning, Sadia Saeed, Amanda Thompson, Sumedha Garg, Elena G. Bochukova, Soren Brage, Victoria Trowse, Eleanor Wheeler, Adrienne E. Sullivan, Mehul Dattani, Peter E. Clayton, Vipan Datta, John B. Bruning, Nick J. Wareham, Stephen O’Rahilly, Daniel J. Peet, Ines Barroso, Murray L. Whitelaw, I. Sadaf Farooqi

Targeting autophagy potentiates tyrosine kinase inhibitor–induced cell death in Philadelphia chromosome–positive cells, including primary CML stem cells

• Text
• PDF

Abstract

Authors

Cristian Bellodi, Maria Rosa Lidonnici, Ashley Hamilton, G. Vignir Helgason, Angela Rachele Soliera, Mattia Ronchetti, Sara Galavotti, Kenneth W. Young, Tommaso Selmi, Rinat Yacobi, Richard A. Van Etten, Nick Donato, Ann Hunter, David Dinsdale, Elena Tirrò, Paolo Vigneri, Pierluigi Nicotera, Martin J. Dyer, Tessa Holyoake, Paolo Salomoni, Bruno Calabretta

Muscle lipogenesis balances insulin sensitivity and strength through calcium signaling

• Text
• PDF

Abstract

Authors

Katsuhiko Funai, Haowei Song, Li Yin, Irfan J. Lodhi, Xiaochao Wei, Jun Yoshino, Trey Coleman, Clay F. Semenkovich