Vascular biology
Abstract
Upon arterial injury, endothelial denudation leads to platelet activation, and delivery of multiple agents (e.g. TXA2, PDGF) promoting VSMC dedifferentiation, and proliferation, in injury repair (intimal hyperplasia). Resolution of vessel injury repair, and prevention of excessive repair (switching VSMC back to a differentiated quiescent state) is a poorly understood process. We now report that internalization of activated platelets by VSMCs promotes resolution of arterial injury by switching on VSMC quiescence. Ex vivo and in vivo studies using lineage tracing reporter mice (PF4-Cre x mTmG) demonstrated uptake of green platelets by red vascular smooth muscle cells upon arterial wire injury. Genome-wide miRNA sequencing of VSMCs co-cultured with activated platelets identified significant increases in platelet-derived miR-223. miR-223 appears to directly target PDGFRβ (in VSMCs) reversing the injury-induced dedifferentiation. Upon arterial injury platelet miR-223 knockout mice exhibit increased intimal hyperplasia, whereas miR-223 mimics reduced intimal hyperplasia. Diabetic mice with reduced expression of miR-223, exhibited enhanced VSMC dedifferentiation, proliferation, and increased intimal hyperplasia. Horizontal transfer of platelet-derived miRNAs into VSMCs provide a novel mechanism for regulating VSMC phenotypic switching. Platelets thus play a dual role in vascular injury repair, initiating an immediate repair process, and concurrently, a delayed process to prevent excessive repair.
Authors
Zhi Zeng, Luoxing Xia, Xuejiao Fan, Allison C. Ostriker, Timur Yarovinsky, Meiling Su, Yuan Zhang, Xiangwen Peng, Xie Yi, Lei Pi, Xiaoqiong Gu, Sookja Kim Chung, Kathleen A. Martin, Renjing Liu, John Hwa, Wai Ho Tang
Abstract
Local flow patterns determine the uneven distribution of atherosclerotic lesions. This research aims to elucidate the mechanism of regulation of nuclear translocation of Yes-associated protein (YAP) under oscillatory shear stress (OSS) in the atheroprone phenotype of endothelial cells (ECs). We report here that OSS led to tyrosine phosphorylation and strong, continuous nuclear translocation of YAP in ECs that is dependent on integrin α5β1 activation. YAP overexpression in ECs blunted the anti-atheroprone effect of an integrin-α5β1 blocking peptide (ATN161) in Apoe-/- mice. Activation of integrin α5β1 induced tyrosine, but not serine, phosphorylation of YAP in ECs. Blockage of integrin α5β1 with ATN161 abolished the phosphorylation of YAP at Y357 induced by OSS. Mechanistic studies showed that c-Abl inhibitor attenuated the integrin α5β1-induced YAP tyrosine phosphorylation. Furthermore, the phosphorylation of c-Abl and YAPY357 was significantly increased in ECs in atherosclerotic vessels of mice and in human plaques vs. normal vessels. Finally, bosutinib, a tyrosine kinase inhibitor, markedly reduced the level of YAPY357 and the development of atherosclerosis in Apoe-/- mice. The c-Abl/YAPY357 pathway serves as a mechanism for the activation of integrin α5β1 and the atherogenic phenotype of ECs in response to OSS, and provides a potential therapeutic strategy for atherogenesis.
Authors
Bochuan Li, Jinlong He, Huizhen Lv, Yajin Liu, Xue Lv, Chenghu Zhang, Yi Zhu, Ding Ai
Abstract
The aortic root is the predominant site for development of aneurysm caused by heterozygous loss-of-function mutations in positive effectors of the transforming growth factor-β (TGF-β) pathway. Using a mouse model of Loeys-Dietz syndrome (LDS) that carries a heterozygous kinase-inactivating mutation in TGF-β receptor I, we found that the effects of this mutation depend on the lineage of origin of vascular smooth muscle cells (VSMCs). Secondary heart field–derived (SHF-derived), but not neighboring cardiac neural crest–derived (CNC-derived), VSMCs showed impaired Smad2/3 activation in response to TGF-β, increased expression of angiotensin II (AngII) type 1 receptor (Agtr1a), enhanced responsiveness to AngII, and higher expression of TGF-β ligands. The preserved TGF-β signaling potential in CNC-derived VSMCs associated, in vivo, with increased Smad2/3 phosphorylation. CNC-, but not SHF-specific, deletion of Smad2 preserved aortic wall architecture and reduced aortic dilation in this mouse model of LDS. Taken together, these data suggest that aortic root aneurysm predisposition in this LDS mouse model depends both on defective Smad signaling in SHF-derived VSMCs and excessive Smad signaling in CNC-derived VSMCs. This work highlights the importance of considering the regional microenvironment and specifically lineage-dependent variation in the vulnerability to mutations in the development and testing of pathogenic models for aortic aneurysm.
Authors
Elena Gallo MacFarlane, Sarah J. Parker, Joseph Y. Shin, Shira G. Ziegler, Tyler J. Creamer, Rustam Bagirzadeh, Djahida Bedja, Yichun Chen, Juan F. Calderon, Katherine Weissler, Pamela A. Frischmeyer-Guerrerio, Mark E. Lindsay, Jennifer P. Habashi, Harry C. Dietz
Abstract
Non-coding RNAs are emerging as important players in gene regulation and disease pathogeneses. Here, we show that a previously uncharacterized long non-coding RNA, NEXN-AS1, modulates the expression of the actin-binding protein NEXN and that NEXN exerts a protective role against atherosclerosis. An expression microarray analysis showed that the expression of both NEXN-AS1 and NEXN were reduced in human atherosclerotic plaques. In vitro experiments revealed that NEXN-AS1 interacted with the chromatin remodeler BAZ1A and the 5′-flanking region of the NEXN gene, and upregulated NEXN expression. Augmentation of NEXN-AS1 expression inhibited toll-like receptor-4 oligomerization and NFκB activity, downregulated the expression of adhesion molecules and inflammatory cytokines by endothelial cells, and suppressed monocyte adhesion to endothelial cells. These inhibitory effects of NEXN-AS1 were abolished by knockdown of NEXN. In vivo experiments of ApoE knockout mice fed a Western high-fat diet demonstrated that NEXN deficiency promoted atherosclerosis and increased macrophage abundance in atherosclerotic lesions, with heightened expression of adhesion molecules and inflammatory cytokines, whereas augmented NEXN expression deterred atherosclerosis. A group of patients with coronary artery disease were found to have lower blood NEXN levels than healthy individuals. These results indicate that NEXN-AS1 and NEXN represent potential therapeutic targets in atherosclerosis related diseases.
Authors
Yan-Wei Hu, Feng-Xia Guo, Yuan-Jun Xu, Pan Li, Zhi-Feng Lu, David G. McVey, Lei Zheng, Qian Wang, John H. Ye, Chun-Min Kang, Shao-Guo Wu, Jing-Jing Zhao, Xin Ma, Zhen Yang, Fu-Chun Fang, Yu-Rong Qiu, Bang-Ming Xu, Lei Xiao, Qian Wu, Li-Mei Wu, Li Ding, Tom R. Webb, Nilesh J. Samani, Shu Ye
Abstract
BACKGROUND.l-Carnitine, an abundant nutrient in red meat, accelerates atherosclerosis in mice via gut microbiota–dependent formation of trimethylamine (TMA) and trimethylamine N-oxide (TMAO) via a multistep pathway involving an atherogenic intermediate, γ-butyrobetaine (γBB). The contribution of γBB in gut microbiota–dependent l-carnitine metabolism in humans is unknown. METHODS. Omnivores and vegans/vegetarians ingested deuterium-labeled l-carnitine (d3-l-carnitine) or γBB (d9-γBB), and both plasma metabolites and fecal polymicrobial transformations were examined at baseline, following oral antibiotics, or following chronic (≥2 months) l-carnitine supplementation. Human fecal commensals capable of performing each step of the l-carnitine→γBB→TMA transformation were identified. RESULTS. Studies with oral d3-l-carnitine or d9-γBB before versus after antibiotic exposure revealed gut microbiota contribution to the initial 2 steps in a metaorganismal l-carnitine→γBB→TMA→TMAO pathway in subjects. Moreover, a striking increase in d3-TMAO generation was observed in omnivores over vegans/vegetarians (>20-fold; P = 0.001) following oral d3-l-carnitine ingestion, whereas fasting endogenous plasma l-carnitine and γBB levels were similar in vegans/vegetarians (n = 32) versus omnivores (n = 40). Fecal metabolic transformation studies, and oral isotope tracer studies before versus after chronic l-carnitine supplementation, revealed that omnivores and vegans/vegetarians alike rapidly converted carnitine to γBB, whereas the second gut microbial transformation, γBB→TMA, was diet inducible (l-carnitine, omnivorous). Extensive anaerobic subculturing of human feces identified no single commensal capable of l-carnitine→TMA transformation, multiple community members that converted l-carnitine to γBB, and only 1 Clostridiales bacterium, Emergencia timonensis, that converted γBB to TMA. In coculture, E. timonensis promoted the complete l-carnitine→TMA transformation. CONCLUSION. In humans, dietary l-carnitine is converted into the atherosclerosis- and thrombosis-promoting metabolite TMAO via 2 sequential gut microbiota–dependent transformations: (a) initial rapid generation of the atherogenic intermediate γBB, followed by (b) transformation into TMA via low-abundance microbiota in omnivores, and to a markedly lower extent, in vegans/vegetarians. Gut microbiota γBB→TMA/TMAO transformation is induced by omnivorous dietary patterns and chronic l-carnitine exposure. TRIAL REGISTRATION. ClinicalTrials.gov NCT01731236. FUNDING. NIH and Office of Dietary Supplements grants HL103866, HL126827, and DK106000, and the Leducq Foundation.
Authors
Robert A. Koeth, Betzabe Rachel Lam-Galvez, Jennifer Kirsop, Zeneng Wang, Bruce S. Levison, Xiaodong Gu, Matthew F. Copeland, David Bartlett, David B. Cody, Hong J. Dai, Miranda K. Culley, Xinmin S. Li, Xiaoming Fu, Yuping Wu, Lin Li, Joseph A. DiDonato, W.H. Wilson Tang, Jose Carlos Garcia-Garcia, Stanley L. Hazen
Abstract
Activation of the type 1 angiotensin II receptor (AT1) triggers proinflammatory signaling through pathways independent of classical Gq signaling that regulate vascular homeostasis. Here, we report that the AT1 receptor preformed a heteromeric complex with the receptor for advanced glycation endproducts (RAGE). Activation of the AT1 receptor by angiotensin II (Ang II) triggered transactivation of the cytosolic tail of RAGE and NF-κB–driven proinflammatory gene expression independently of the liberation of RAGE ligands or the ligand-binding ectodomain of RAGE. The importance of this transactivation pathway was demonstrated by our finding that adverse proinflammatory signaling events induced by AT1 receptor activation were attenuated when RAGE was deleted or transactivation of its cytosolic tail was inhibited. At the same time, classical homeostatic Gq signaling pathways were unaffected by RAGE deletion or inhibition. These data position RAGE transactivation by the AT1 receptor as a target for vasculoprotective interventions. As proof of concept, we showed that treatment with the mutant RAGE peptide S391A-RAGE362–404 was able to inhibit transactivation of RAGE and attenuate Ang II–dependent inflammation and atherogenesis. Furthermore, treatment with WT RAGE362–404 restored Ang II–dependent atherogenesis in Ager/Apoe-KO mice, without restoring ligand-mediated signaling via RAGE, suggesting that the major effector of RAGE activation was its transactivation.
Authors
Raelene J. Pickering, Christos Tikellis, Carlos J. Rosado, Despina Tsorotes, Alexandra Dimitropoulos, Monique Smith, Olivier Huet, Ruth M. Seeber, Rekhati Abhayawardana, Elizabeth K.M. Johnstone, Jonathan Golledge, Yutang Wang, Karin A. Jandeleit-Dahm, Mark E. Cooper, Kevin D.G. Pfleger, Merlin C. Thomas
Abstract
Tumor cure with conventional fractionated radiotherapy is 65%, dependent on tumor cell-autonomous gradual buildup of DNA double strand break (DSB) misrepair. Here we report single dose radiotherapy (SDRT), a disruptive technique that ablates >90% of human cancers, operates a distinct dual-target mechanism, linking acid sphingomyelinase (ASMase)-mediated microvascular perfusion defects to DNA unrepair in tumor cells to confer tumor cell lethality. ASMase-mediated microcirculatory vasoconstriction post-SDRT conferred an ischemic stress response within parenchymal tumor cells, with reactive oxygen species triggering the evolutionarily conserved SUMO Stress Response, specifically depleting chromatin-associated free SUMO3. Whereas SUMO3, but not SUMO2, was indispensible for homology-directed repair (HDR) of DSBs, HDR loss-of-function post-SDRT yielded DSB unrepair, chromosomal aberrations and tumor clonogen demise. Vasoconstriction blockade with the endothelin-1 inhibitor BQ-123, or ROS scavenging post-SDRT using peroxiredoxin-6 overexpression or the SOD-mimetic tempol, prevented chromatin SUMO3 depletion, HDR loss-of-function and SDRT tumor ablation. We also provide evidence of mouse to human translation of this biology in a randomized clinical trial, showing 24Gy SDRT, but not 3x9Gy fractionation, coupled early tumor ischemia/reperfusion to human cancer ablation. The SDRT biology provides opportunities for mechanism-based selective tumor radiosensitization via accessing SDRT/ASMase signaling, as current studies indicate this pathway is tractable to pharmacologic intervention.
Authors
Sahra Bodo, Cecile Campagne, Tin Htwe Thin, Daniel S. Higginson, H. Alberto Vargas, Guoqiang Hua, John D. Fuller, Ellen Ackerstaff, James Russell, Zhigang Zhang, Stefan Klingler, HyungJoon Cho, Matthew G. Kaag, Yousef Mazaheri, Andreas Rimner, Katia Manova-Todorova, Boris Epel, Joan Zatcky, Cristian R. Cleary, Shyam S. Rao, Yoshiya Yamada, Michael J. Zelefsky, Howard J. Halpern, Jason A. Koutcher, Carlos Cordon-Cardo, Carlo Greco, Adriana Haimovitz-Friedman, Evis Sala, Simon N. Powell, Richard Kolesnick, Zvi Fuks
Abstract
Emerging evidence indicates that angiopoietin-2 (Angpt2), a well-recognized vascular destabilizing factor, is a biomarker of poor outcome in ischemic heart disease. However, its precise role in postischemic cardiovascular remodeling is poorly understood. Here, we show that Angpt2 plays multifaceted roles in the exacerbation of cardiac hypoxia and inflammation after myocardial ischemia. Angpt2 was highly expressed in endothelial cells at the infarct border zone after myocardial infarction (MI) or ischemia/reperfusion injury in mice. In the acute phase of MI, endothelial-derived Angpt2 antagonized Angpt1/Tie2 signaling, which was greatly involved in pericyte detachment, vascular leakage, increased adhesion molecular expression, degradation of the glycocalyx and extracellular matrix, and enhanced neutrophil infiltration and hypoxia in the infarct border area. In the chronic remodeling phase after MI, endothelial- and macrophage-derived Angpt2 continuously promoted abnormal vascular remodeling and proinflammatory macrophage polarization through integrin α5β1 signaling, worsening cardiac hypoxia and inflammation. Accordingly, inhibition of Angpt2 either by gene deletion or using an anti-Angpt2 blocking antibody substantially alleviated these pathological findings and ameliorated postischemic cardiovascular remodeling. Blockade of Angpt2 thus has potential as a therapeutic option for ischemic heart failure.
Authors
Seung-Jun Lee, Choong-kun Lee, Seok Kang, Intae Park, Yoo Hyung Kim, Seo Ki Kim, Seon Pyo Hong, Hosung Bae, Yulong He, Yoshiaki Kubota, Gou Young Koh
Abstract
Arteriolar endothelial cell–expressed (EC-expressed) α-globin binds endothelial NOS (eNOS) and degrades its enzymatic product, NO, via dioxygenation, thereby lessening the vasodilatory effects of NO on nearby vascular smooth muscle. Although this reaction potentially affects vascular physiology, the mechanisms that regulate α-globin expression and dioxygenase activity in ECs are unknown. Without β-globin, α-globin is unstable and cytotoxic, particularly in its oxidized form, which is generated by dioxygenation and recycled via endogenous reductases. We show that the molecular chaperone α-hemoglobin–stabilizing protein (AHSP) promotes arteriolar α-globin expression in vivo and facilitates its reduction by eNOS. In Ahsp−/− mice, EC α-globin was decreased by 70%. Ahsp−/− and Hba1−/− mice exhibited similar evidence of increased vascular NO signaling, including arteriolar dilation, blunted α1-adrenergic vasoconstriction, and reduced blood pressure. Purified α-globin bound eNOS or AHSP, but not both together. In ECs in culture, eNOS or AHSP enhanced α-globin expression posttranscriptionally. However, only AHSP prevented oxidized α-globin precipitation in solution. Finally, eNOS reduced AHSP-bound α-globin approximately 6-fold faster than did the major erythrocyte hemoglobin reductases (cytochrome B5 reductase plus cytochrome B5). Our data support a model whereby redox-sensitive shuttling of EC α-globin between AHSP and eNOS regulates EC NO degradation and vascular tone.
Authors
Christophe Lechauve, Joshua T. Butcher, Abdullah Freiwan, Lauren A. Biwer, Julia M. Keith, Miranda E. Good, Hans Ackerman, Heather S. Tillman, Laurent Kiger, Brant E. Isakson, Mitchell J. Weiss
Abstract
The M2 isoform of pyruvate kinase (PKM2) is highly expressed in most cancer cells, and has been studied extensively as a driver of oncogenic metabolism. In contrast, the role of PKM2 in nontransformed cells is little studied, and nearly nothing is known of its role, if any, in quiescent cells. We show here that endothelial cells express PKM2 almost exclusively over PKM1. In proliferating endothelial cells, PKM2 is required to suppress p53 and maintain cell cycle progression. In sharp contrast, PKM2 has a strikingly different role in quiescent endothelial cells, where inhibition of PKM2 leads to degeneration of tight junctions and barrier function. Mechanistically, PKM2 regulates barrier function independently of its canonical activity as a pyruvate kinase. Instead, PKM2 suppresses NF-kB and its downstream target, the vascular permeability factor angiopoietin 2. As a consequence, loss of endothelial cell PKM2 in vivo predisposes mice to VEGF-induced vascular leak, and to severe bacteremia and death in response to sepsis. Together, these data demonstrate new roles of PKM2 in quiescent cells, and highlight the need for caution in developing cancer therapies that target PKM2.
Authors
Boa Kim, Cholsoon Jang, Harita Dharaneeswaran, Jian Li, Mohit Bhide, Steven Yang, Kristina Li, Zolt Arany
Copyright © 2019 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)