The chromatin-remodeling enzyme CHD4 maintains vascular integrity at mid-gestation; however, it is unknown whether this enzyme contributes to later blood vessel or lymphatic vessel development. Here, we addressed this issue in mice harboring a deletion of
Patrick L. Crosswhite, Joanna J. Podsiadlowska, Carol D. Curtis, Siqi Gao, Lijun Xia, R. Sathish Srinivasan, Courtney T. Griffin
The blood-brain barrier (BBB) protects the brain from toxic substances within the peripheral circulation. It maintains brain homeostasis and is a hurdle for drug delivery to the CNS to treat neurodegenerative diseases, including Alzheimer’s disease and brain tumors. The drug efflux transporter P-glycoprotein (P-gp) is highly expressed on brain endothelial cells and blocks the entry of most drugs delivered to the brain. Here, we show that activation of the A2A adenosine receptor (AR) with an FDA-approved A2A AR agonist (Lexiscan) rapidly and potently decreased P-gp expression and function in a time-dependent and reversible manner. We demonstrate that downmodulation of P-gp expression and function coincided with chemotherapeutic drug accumulation in brains of WT mice and in primary mouse and human brain endothelial cells, which serve as in vitro BBB models. Lexiscan also potently downregulated the expression of BCRP1, an efflux transporter that is highly expressed in the CNS vasculature and other tissues. Finally, we determined that multiple pathways, including MMP9 cleavage and ubiquitinylation, mediated P-gp downmodulation. Based on these data, we propose that A2A AR activation on BBB endothelial cells offers a therapeutic window that can be fine-tuned for drug delivery to the brain and has potential as a CNS drug-delivery technology.
Do-Geun Kim, Margaret S. Bynoe
Lymphatic collecting vessels direct lymph into and from lymph nodes (LNs) and can become hyperpermeable as the result of a previous infection. Enhanced permeability has been implicated in compromised immunity due to reduced flow of lymph and immune cells to LNs, which are the primary site of antigen presentation to T cells. Presently, very little is known about the molecular signals that affect lymphatic collecting vessel permeability. Here, we have shown that lymphatic collecting vessel permeability is controlled by CCR7 and that the chronic hyperpermeability of collecting vessels observed in
Stoyan Ivanov, Joshua P. Scallan, Ki-Wook Kim, Kathrin Werth, Michael W. Johnson, Brian T. Saunders, Peter L. Wang, Emma L. Kuan, Adam C. Straub, Melissa Ouhachi, Erica G. Weinstein, Jesse W. Williams, Carlos Briseño, Marco Colonna, Brant E. Isakson, Emmanuel L. Gautier, Reinhold Förster, Michael J. Davis, Bernd H. Zinselmeyer, Gwendalyn J. Randolph
Abdominal aortic aneurysm (AAA) is a major cause of morbidity and mortality; however, the mechanisms that are involved in disease initiation and progression are incompletely understood. Extracellular matrix proteins play an integral role in modulating vascular homeostasis in health and disease. Here, we determined that the expression of the matricellular protein CCN3 is strongly reduced in rodent AAA models, including angiotensin II–induced AAA and elastase perfusion–stimulated AAA. CCN3 levels were also reduced in human AAA biopsies compared with those in controls. In murine models of induced AAA, germline deletion of
Chao Zhang, Dustin van der Voort, Hong Shi, Rongli Zhang, Yulan Qing, Shuichi Hiraoka, Minoru Takemoto, Koutaro Yokote, Joseph V. Moxon, Paul Norman, Laure Rittié, Helena Kuivaniemi, G. Brandon Atkins, Stanton L. Gerson, Guo-Ping Shi, Jonathan Golledge, Nianguo Dong, Bernard Perbal, Domenick A. Prosdocimo, Zhiyong Lin
Vascular calcification is a common feature of major cardiovascular diseases. Extracellular vesicles participate in the formation of microcalcifications that are implicated in atherosclerotic plaque rupture; however, the mechanisms that regulate formation of calcifying extracellular vesicles remain obscure. Here, we have demonstrated that sortilin is a key regulator of smooth muscle cell (SMC) calcification via its recruitment to extracellular vesicles. Sortilin localized to calcifying vessels in human and mouse atheromata and participated in formation of microcalcifications in SMC culture. Sortilin regulated the loading of the calcification protein tissue nonspecific alkaline phosphatase (TNAP) into extracellular vesicles, thereby conferring its calcification potential. Furthermore, SMC calcification required Rab11-dependent trafficking and FAM20C/casein kinase 2–dependent C-terminal phosphorylation of sortilin. In a murine model,
Claudia Goettsch, Joshua D. Hutcheson, Masanori Aikawa, Hiroshi Iwata, Tan Pham, Anders Nykjaer, Mads Kjolby, Maximilian Rogers, Thomas Michel, Manabu Shibasaki, Sumihiko Hagita, Rafael Kramann, Daniel J. Rader, Peter Libby, Sasha A. Singh, Elena Aikawa
The ascending thoracic aorta is designed to withstand biomechanical forces from pulsatile blood. Thoracic aortic aneurysms and acute aortic dissections (TAADs) occur as a result of genetically triggered defects in aortic structure and a dysfunctional response to these forces. Here, we describe mutations in the forkhead transcription factor
Shao-Qing Kuang, Olga Medina-Martinez, Dong-chuan Guo, Limin Gong, Ellen S. Regalado, Corey L. Reynolds, Catherine Boileau, Guillaume Jondeau, Siddharth K. Prakash, Callie S. Kwartler, Lawrence Yang Zhu, Andrew M. Peters, Xue-Yan Duan, National Registry of Genetically Triggered Thoracic Aortic Aneurysms and Cardiovascular Conditions (GenTAC) Investigators, National Heart, Lung, and Blood Institute (NHLBI) Grand Opportunity (GO) Exome Sequencing Project (ESP), Michael J. Bamshad, Jay Shendure, Debbie A. Nickerson, Regie L. Santos-Cortez, Xiurong Dong, Suzanne M. Leal, Mark W. Majesky, Eric C. Swindell, Milan Jamrich, Dianna M. Milewicz
The small intestine is a dynamic and complex organ that is characterized by constant epithelium turnover and crosstalk among various cell types and the microbiota. Lymphatic capillaries of the small intestine, called lacteals, play key roles in dietary fat absorption and the gut immune response; however, little is known about the molecular regulation of lacteal function. Here, we performed a high-resolution analysis of the small intestinal stroma and determined that lacteals reside in a permanent regenerative, proliferative state that is distinct from embryonic lymphangiogenesis or quiescent lymphatic vessels observed in other tissues. We further demonstrated that this continuous regeneration process is mediated by Notch signaling and that the expression of the Notch ligand delta-like 4 (DLL4) in lacteals requires activation of VEGFR3 and VEGFR2. Moreover, genetic inactivation of
Jeremiah Bernier-Latmani, Christophe Cisarovsky, Cansaran Saygili Demir, Marine Bruand, Muriel Jaquet, Suzel Davanture, Simone Ragusa, Stefanie Siegert, Olivier Dormond, Rui Benedito, Freddy Radtke, Sanjiv A. Luther, Tatiana V. Petrova
Cellular metabolism is increasingly recognized as a controller of immune cell fate and function. MicroRNA-33 (miR-33) regulates cellular lipid metabolism and represses genes involved in cholesterol efflux, HDL biogenesis, and fatty acid oxidation. Here, we determined that miR-33–mediated disruption of the balance of aerobic glycolysis and mitochondrial oxidative phosphorylation instructs macrophage inflammatory polarization and shapes innate and adaptive immune responses. Macrophage-specific
Mireille Ouimet, Hasini N. Ediriweera, U. Mahesh Gundra, Frederick J. Sheedy, Bhama Ramkhelawon, Susan B. Hutchison, Kaitlyn Rinehold, Coen van Solingen, Morgan D. Fullerton, Katharine Cecchini, Katey J. Rayner, Gregory R. Steinberg, Phillip D. Zamore, Edward A. Fisher, P’ng Loke, Kathryn J. Moore
The molecular mechanisms responsible for the development and progression of atherosclerotic lesions have not been fully established. Here, we investigated the role played by endothelial-to-mesenchymal transition (EndMT) and its key regulator FGF receptor 1 (FGFR1) in atherosclerosis. In cultured human endothelial cells, both inflammatory cytokines and oscillatory shear stress reduced endothelial FGFR1 expression and activated TGF-β signaling. We further explored the link between disrupted FGF endothelial signaling and progression of atherosclerosis by introducing endothelial-specific deletion of FGF receptor substrate 2 α (
Pei-Yu Chen, Lingfeng Qin, Nicolas Baeyens, Guangxin Li, Titilayo Afolabi, Madhusudhan Budatha, George Tellides, Martin A. Schwartz, Michael Simons
Recent evidence indicates that saturated fatty acid–induced (SFA-induced) lipotoxicity contributes to the pathogenesis of cardiovascular and metabolic diseases; however, the molecular mechanisms that underlie SFA-induced lipotoxicity remain unclear. Here, we have shown that repression of stearoyl-CoA desaturase (SCD) enzymes, which regulate the intracellular balance of SFAs and unsaturated FAs, and the subsequent accumulation of SFAs in vascular smooth muscle cells (VSMCs), are characteristic events in the development of vascular calcification. We evaluated whether SMC-specific inhibition of SCD and the resulting SFA accumulation plays a causative role in the pathogenesis of vascular calcification and generated mice with SMC-specific deletion of both
Masashi Masuda, Shinobu Miyazaki-Anzai, Audrey L. Keenan, Kayo Okamura, Jessica Kendrick, Michel Chonchol, Stefan Offermanns, James M. Ntambi, Makoto Kuro-o, Makoto Miyazaki
Lacteals are lymphatic vessels located at the center of each intestinal villus and provide essential transport routes for lipids and other lipophilic molecules. However, it is unclear how absorbed molecules are transported through the lacteal. Here, we used reporter mice that express GFP under the control of the lymphatic-specific promoter
Kibaek Choe, Jeon Yeob Jang, Intae Park, Yeseul Kim, Soyeon Ahn, Dae-Young Park, Young-Kwon Hong, Kari Alitalo, Gou Young Koh, Pilhan Kim
Biomechanical forces, such as fluid shear stress, govern multiple aspects of endothelial cell biology. In blood vessels, disturbed flow is associated with vascular diseases, such as atherosclerosis, and promotes endothelial cell proliferation and apoptosis. Here, we identified an important role for disturbed flow in lymphatic vessels, in which it cooperates with the transcription factor FOXC2 to ensure lifelong stability of the lymphatic vasculature. In cultured lymphatic endothelial cells,
Amélie Sabine, Esther Bovay, Cansaran Saygili Demir, Wataru Kimura, Muriel Jaquet, Yan Agalarov, Nadine Zangger, Joshua P. Scallan, Werner Graber, Elgin Gulpinar, Brenda R. Kwak, Taija Mäkinen, Inés Martinez-Corral, Sagrario Ortega, Mauro Delorenzi, Friedemann Kiefer, Michael J. Davis, Valentin Djonov, Naoyuki Miura, Tatiana V. Petrova
Heterozygous germline mutations in the zinc finger transcription factor
Jan Kazenwadel, Kelly L. Betterman, Chan-Eng Chong, Philippa H. Stokes, Young K. Lee, Genevieve A. Secker, Yan Agalarov, Cansaran Saygili Demir, David M. Lawrence, Drew L. Sutton, Sebastien P. Tabruyn, Naoyuki Miura, Marjo Salminen, Tatiana V. Petrova, Jacqueline M. Matthews, Christopher N. Hahn, Hamish S. Scott, Natasha L. Harvey
Fluid shear forces have established roles in blood vascular development and function, but whether such forces similarly influence the low-flow lymphatic system is unknown. It has been difficult to test the contribution of fluid forces in vivo because mechanical or genetic perturbations that alter flow often have direct effects on vessel growth. Here, we investigated the functional role of flow in lymphatic vessel development using mice deficient for the platelet-specific receptor C-type lectin–like receptor 2 (CLEC2) as blood backfills the lymphatic network and blocks lymph flow in these animals. CLEC2-deficient animals exhibited normal growth of the primary mesenteric lymphatic plexus but failed to form valves in these vessels or remodel them into a structured, hierarchical network. Smooth muscle cell coverage (SMC coverage) of CLEC2-deficient lymphatic vessels was both premature and excessive, a phenotype identical to that observed with loss of the lymphatic endothelial transcription factor FOXC2. In vitro evaluation of lymphatic endothelial cells (LECs) revealed that low, reversing shear stress is sufficient to induce expression of genes required for lymphatic valve development and identified GATA2 as an upstream transcriptional regulator of FOXC2 and the lymphatic valve genetic program. These studies reveal that lymph flow initiates and regulates many of the key steps in collecting lymphatic vessel maturation and development.
Daniel T. Sweet, Juan M. Jiménez, Jeremy Chang, Paul R. Hess, Patricia Mericko-Ishizuka, Jianxin Fu, Lijun Xia, Peter F. Davies, Mark L. Kahn
Hereditary angioedema type III (HAEIII) is a rare inherited swelling disorder that is associated with point mutations in the gene encoding the plasma protease factor XII (FXII). Here, we demonstrate that HAEIII-associated mutant FXII, derived either from HAEIII patients or recombinantly produced, is defective in mucin-type Thr309-linked glycosylation. Loss of glycosylation led to increased contact-mediated autoactivation of zymogen FXII, resulting in excessive activation of the bradykinin-forming kallikrein-kinin pathway. In contrast, both FXII-driven coagulation and the ability of C1-esterase inhibitor to bind and inhibit activated FXII were not affected by the mutation. Intravital laser-scanning microscopy revealed that, compared with control animals, both
Jenny Björkqvist, Steven de Maat, Urs Lewandrowski, Antonio Di Gennaro, Chris Oschatz, Kai Schönig, Markus M. Nöthen, Christian Drouet, Hal Braley, Marc W. Nolte, Albert Sickmann, Con Panousis, Coen Maas, Thomas Renné
Elevated blood pressure is a key risk factor for developing cardiovascular diseases. Blood pressure is largely determined by vasodilatory mediators, such as nitric oxide (NO), that are released from the endothelium in response to fluid shear stress exerted by the flowing blood. Previous work has identified several mechanotransduction signaling processes that are involved in fluid shear stress–induced endothelial effects, but how fluid shear stress initiates the response is poorly understood. Here, we evaluated human and bovine endothelial cells and found that the purinergic receptor P2Y2 and the G proteins Gq/G11 mediate fluid shear stress–induced endothelial responses, including [Ca2+]i transients, activation of the endothelial NO synthase (eNOS), phosphorylation of PECAM-1 and VEGFR-2, as well as activation of SRC and AKT. In response to fluid shear stress, endothelial cells released ATP, which activates the purinergic P2Y2 receptor. Mice with induced endothelium-specific P2Y2 or Gq/G11 deficiency lacked flow-induced vasodilation and developed hypertension that was accompanied by reduced eNOS activation. Together, our data identify P2Y2 and Gq/G11 as a critical endothelial mechanosignaling pathway that is upstream of previously described mechanotransduction processes and demonstrate that P2Y2 and Gq/G11 are required for basal endothelial NO formation, vascular tone, and blood pressure.
ShengPeng Wang, András Iring, Boris Strilic, Julián Albarrán Juárez, Harmandeep Kaur, Kerstin Troidl, Sarah Tonack, Joachim C. Burbiel, Christa E. Müller, Ingrid Fleming, Jon O. Lundberg, Nina Wettschureck, Stefan Offermanns
The ability of cells to detect and respond to nucleotide signals in the local microenvironment is essential for vascular homeostasis. The enzyme ectonucleotide tri(di)phosphohydrolase-1 (ENTPD1, also known as CD39) on the surface of leukocytes and endothelial cells metabolizes locally released, intravascular ATP and ADP, thereby eliminating these prothrombotic and proinflammatory stimuli. Here, we evaluated the contribution of CD39 to atherogenesis in the apolipoprotein E–deficient (ApoE-deficient) mouse model of atherosclerosis. Compared with control ApoE-deficient animals, plaque burden was markedly increased along with circulating markers of platelet activation in
Yogendra Kanthi, Matthew C. Hyman, Hui Liao, Amy E. Baek, Scott H. Visovatti, Nadia R. Sutton, Sascha N. Goonewardena, Mithun K. Neral, Hanjoong Jo, David J. Pinsky
In mammals, the outflow tract (OFT) of the developing heart septates into the base of the pulmonary artery and aorta to guide deoxygenated right ventricular blood into the lungs and oxygenated left ventricular blood into the systemic circulation. Accordingly, defective OFT septation is a life-threatening condition that can occur in both syndromic and nonsyndromic congenital heart disease. Even though studies of genetic mouse models have previously revealed a requirement for VEGF-A, the class 3 semaphorin SEMA3C, and their shared receptor neuropilin 1 (NRP1) in OFT development, the precise mechanism by which these proteins orchestrate OFT septation is not yet understood. Here, we have analyzed a complementary set of ligand-specific and tissue-specific mouse mutants to show that neural crest–derived SEMA3C activates NRP1 in the OFT endothelium. Explant assays combined with gene-expression studies and lineage tracing further demonstrated that this signaling pathway promotes an endothelial-to-mesenchymal transition that supplies cells to the endocardial cushions and repositions cardiac neural crest cells (NCCs) within the OFT, 2 processes that are essential for septal bridge formation. These findings elucidate a mechanism by which NCCs cooperate with endothelial cells in the developing OFT to enable the postnatal separation of the pulmonary and systemic circulation.
Alice Plein, Amélie Calmont, Alessandro Fantin, Laura Denti, Naomi A. Anderson, Peter J. Scambler, Christiana Ruhrberg
Bacterial meningitis is a serious infection of the CNS that results when blood-borne bacteria are able to cross the blood-brain barrier (BBB). Group B
Brandon J. Kim, Bryan M. Hancock, Andres Bermudez, Natasha Del Cid, Efren Reyes, Nina M. van Sorge, Xavier Lauth, Cameron A. Smurthwaite, Brett J. Hilton, Aleksandr Stotland, Anirban Banerjee, John Buchanan, Roland Wolkowicz, David Traver, Kelly S. Doran
Functional interactions between neurons, vasculature, and glia within neurovascular units are critical for maintenance of the retina and other CNS tissues. For example, the architecture of the neurosensory retina is a highly organized structure with alternating layers of neurons and blood vessels that match the metabolic demand of neuronal activity with an appropriate supply of oxygen within perfused blood. Here, using murine genetic models and cell ablation strategies, we have demonstrated that a subset of retinal interneurons, the amacrine and horizontal cells, form neurovascular units with capillaries in 2 of the 3 retinal vascular plexuses. Moreover, we determined that these cells are required for generating and maintaining the intraretinal vasculature through precise regulation of hypoxia-inducible and proangiogenic factors, and that amacrine and horizontal cell dysfunction induces alterations to the intraretinal vasculature and substantial visual deficits. These findings demonstrate that specific retinal interneurons and the intraretinal vasculature are highly interdependent, and loss of either or both elicits profound effects on photoreceptor survival and function.
Yoshihiko Usui, Peter D. Westenskow, Toshihide Kurihara, Edith Aguilar, Susumu Sakimoto, Liliana P. Paris, Carli Wittgrove, Daniel Feitelberg, Mollie S.H. Friedlander, Stacey K. Moreno, Michael I. Dorrell, Martin Friedlander