Dynamic Regulation of Vascular Myosin Light Chain (MYL9) with Injury and Aging

Aging-associated changes in the cardiovascular system increase the risk for disease development and lead to profound alterations in vascular reactivity and stiffness. Elucidating the molecular response of arteries to injury and age will help understand the exaggerated remodeling of aging vessels.We studied the gene expression profile in a model of mechanical vascular injury in the iliac artery of aging (22 months old) and young rats (4 months old). We investigated aging-related variations in gene expression at 30 min, 3 d and 7 d post injury. We found that the Myosin Light Chain gene (MYL9) was the only gene differentially expressed in the aged versus young injured arteries at all time points studied, peaking at day 3 after injury (4.6 fold upregulation (p<0.05) in the smooth muscle cell layers. We confirmed this finding on an aging aortic microarray experiment available through NCBI's GEO database. We found that Myl9 was consistently upregulated with age in healthy rat aortas. To determine the arterial localization of Myl9 with age and injury, we performed immunohistochemistry for Myl9 in rat iliac arteries and found that in healthy and injured (30 days post injury) arteries, Myl9 expression increased with age in the endothelial layers.The consistent upregulation of the myosin light chain protein (Myl9) with age and injury in arterial tissue draws attention to the increased vascular permeability and to the age-caused predisposition to arterial constriction after balloon angioplasty

Activation of Serine One-Carbon Metabolism by Calcineurin A beta 1 Reduces Myocardial Hypertrophy and Improves Ventricular Function

Background In response to pressure overload, the heart develops ventricular hypertrophy that progressively decompensates and leads to heart failure. This pathological hypertrophy is mediated, among others, by the phosphatase calcineurin and is characterized by metabolic changes that impair energy production by mitochondria. Objectives The authors aimed to determine the role of the calcineurin splicing variant CnAβ1 in the context of cardiac hypertrophy and its mechanism of action. Methods Transgenic mice overexpressing CnAβ1 specifically in cardiomyocytes and mice lacking the unique C-terminal domain in CnAβ1 (CnAβ1Δi12 mice) were used. Pressure overload hypertrophy was induced by transaortic constriction. Cardiac function was measured by echocardiography. Mice were characterized using various molecular analyses. Results In contrast to other calcineurin isoforms, the authors show here that cardiac-specific overexpression of CnAβ1 in transgenic mice reduces cardiac hypertrophy and improves cardiac function. This effect is mediated by activation of serine and one-carbon metabolism, and the production of antioxidant mediators that prevent mitochondrial protein oxidation and preserve ATP production. The induction of enzymes involved in this metabolic pathway by CnAβ1 is dependent on mTOR activity. Inhibition of serine and one-carbon metabolism blocks the beneficial effects of CnAβ1. CnAβ1Δi12 mice show increased cardiac hypertrophy and declined contractility. Conclusions The metabolic reprogramming induced by CnAβ1 redefines the role of calcineurin in the heart and shows for the first time that activation of the serine and one-carbon pathway has beneficial effects on cardiac hypertrophy and function, paving the way for new therapeutic approaches

Activation of Serine One-Carbon Metabolism by Calcineurin A beta 1 Reduces Myocardial Hypertrophy and Improves Ventricular Function

BACKGROUND In response to pressure overload, the heart develops ventricular hypertrophy that progressively decompensates and leads to heart failure. This pathological hypertrophy is mediated, among others, by the phosphatase calcineurin and is characterized by metabolic changes that impair energy production by mitochondria. OBJECTIVES The authors aimed to determine the role of the calcineurin splicing variant CnA beta 1 in the context of cardiac hypertrophy and its mechanism of action. METHODS Transgenic mice overexpressing CnAb1 specifically in cardiomyocytes and mice lacking the unique C-terminal domain in CnA beta 1 (CnA beta 1(Delta i12) mice) were used. Pressure overload hypertrophy was induced by transaortic constriction. Cardiac function was measured by echocardiography. Mice were characterized using various molecular analyses. RESULTS In contrast to other calcineurin isoforms, the authors show here that cardiac-specific overexpression of CnA beta 1 in transgenic mice reduces cardiac hypertrophy and improves cardiac function. This effect is mediated by activation of serine and one-carbon metabolism, and the production of antioxidant mediators that prevent mitochondrial protein oxidation and preserve ATP production. The induction of enzymes involved in this metabolic pathway by CnAb1 is dependent on mTOR activity. Inhibition of serine and one-carbon metabolism blocks the beneficial effects of CnA beta 1. CnA beta 1(Delta i12) mice show increased cardiac hypertrophy and declined contractility. CONCLUSIONS The metabolic reprogramming induced by CnAb1 redefines the role of calcineurin in the heart and shows for the first time that activation of the serine and one-carbon pathway has beneficial effects on cardiac hypertrophy and function, paving the way for new therapeutic approaches. (J Am Coll Cardiol 2018; 71: 654-67) (C) 2018 The Authors. Published by Elsevier on behalf of the American College of Cardiology Foundation. This is an open access article under the CC BY-NC-ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/).This work was supported by grants from the European Union (CardioNeT-ITN-289600 and CardioNext-608027 to Dr. Lara-Pezzi; Meet-ITN-317433 to Dr. Enriquez; UE0/MCA1108 to Dr. Acin-Perez), from the Spanish Ministry of Economy and Competitiveness (SAF2015-65722-R and SAF2012-31451 to Dr. Lara-Pezzi; SAF2015-71521-REDC, BFU2013-50448, and SAF2012-32776 to Dr. Enriquez; RyC-2011-07826 to Dr. Acin-Perez; BIO2012-37926 and BIO2015-67580-P to Dr. Vazquez), from the Spanish Carlos III Institute of Health (CPII14/00027 to Dr. Lara-Pezzi; RD12/0042/066 to Drs. Garcia-Pavia and Lara-Pezzi), from the Regional Government of Madrid (2010-BMD-2321 ``Fibroteam´´ to Dr. Lara-Pezzi; 2011-BMD-2402 ``Mitolab´´ to Dr. Enriquez) and the FIS-ISCIII (PRB2-IPT13/0001 and RD12/0042/0056-RIC-RETICS to Dr. Vazquez). This work was also supported by the Plan Estatal de IthornDthornI 2013-2016-European Regional Development Fund (FEDER) ``A way of making Europe,´´ Spain. The CNIC is supported by the Spanish Ministry of Economy and Competitiveness and by the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (MINECO award SEV-2015-0505). Drs. Vazquez and Garcia-Pavia have served as consultants for VL39. All other authors have reported that they have no relationships relevant to the contents of this paper to disclose. Drs. Padron-Barthe, Villalba-Orero, and Gomez-Salinero contributed equally to this work and are joint first authors. Robyn Shaw, MD, PhD, served as Guest Editor for this paper.S

Aorta in Pathologies May Function as an Immune Organ by Upregulating Secretomes for Immune and Vascular Cell Activation, Differentiation and Trans-Differentiation-Early Secretomes may Serve as Drivers for Trained Immunity

To determine whether aorta becomes immune organ in pathologies, we performed transcriptomic analyses of six types of secretomic genes (SGs) in aorta and vascular cells and made the following findings: 1) 53.7% out of 21,306 human protein genes are classified into six secretomes, namely, canonical, caspase 1, caspase 4, exosome, Weibel-Palade body, and autophagy; 2) Atherosclerosis (AS), chronic kidney disease (CKD) and abdominal aortic aneurysm (AAA) modulate six secretomes in aortas; and Middle East Respiratory Syndrome Coronavirus (MERS-CoV, COVID-19 homologous) infected endothelial cells (ECs) and angiotensin-II (Ang-II) treated vascular smooth muscle cells (VSMCs) modulate six secretomes; 3) AS aortas upregulate T and B cell immune SGs; CKD aortas upregulate SGs for cardiac hypertrophy, and hepatic fibrosis; and AAA aorta upregulate SGs for neuromuscular signaling and protein catabolism; 4) Ang-II induced AAA, canonical, caspase 4, and exosome SGs have two expression peaks of high (day 7)-low (day 14)-high (day 28) patterns; 5) Elastase induced AAA aortas have more inflammatory/immune pathways than that of Ang-II induced AAA aortas; 6) Most disease-upregulated cytokines in aorta may be secreted via canonical and exosome secretomes; 7) Canonical and caspase 1 SGs play roles at early MERS-CoV infected ECs whereas caspase 4 and exosome SGs play roles in late/chronic phases; and the early upregulated canonical and caspase 1 SGs may function as drivers for trained immunity (innate immune memory); 8) Venous ECs from arteriovenous fistula (AVF) upregulate SGs in five secretomes; and 9) Increased some of 101 trained immunity genes and decreased trained tolerance regulator IRG1 participate in upregulations of SGs in atherosclerotic, Ang-II induced AAA and CKD aortas, and MERS-CoV infected ECs, but less in SGs upregulated in AVF ECs. IL-1 family cytokines, HIF1α, SET7 and mTOR, ROS regulators NRF2 and NOX2 partially regulate trained immunity genes; and NRF2 plays roles in downregulating SGs more than that of NOX2 in upregulating SGs. These results provide novel insights on the roles of aorta as immune organ in upregulating secretomes and driving immune and vascular cell differentiations in COVID-19, cardiovascular diseases, inflammations, transplantations, autoimmune diseases and cancers

Integrin CD11b activation drives anti-tumor innate immunity

Myeloid cells are recruited to damaged tissues where they can resolve infections and tumor growth or stimulate wound healing and tumor progression. Recruitment of these cells is regulated by integrins, a family of adhesion receptors that includes integrin CD11b. Here we report that, unexpectedly, integrin CD11b does not regulate myeloid cell recruitment to tumors but instead controls myeloid cell polarization and tumor growth. CD11b activation promotes pro-inflammatory macrophage polarization by stimulating expression of microRNA Let7a. In contrast, inhibition of CD11b prevents Let7a expression and induces cMyc expression, leading to immune suppressive macrophage polarization, vascular maturation, and accelerated tumor growth. Pharmacological activation of CD11b with a small molecule agonist, Leukadherin 1 (LA1), promotes pro-inflammatory macrophage polarization and suppresses tumor growth in animal models of murine and human cancer. These studies identify CD11b as negative regulator of immune suppression and a target for cancer immune therapy

Innate immunity of vascular smooth muscle cells contributes to two-wave inflammation in atherosclerosis, twin-peak inflammation in aortic aneurysms and trans-differentiation potential into 25 cell types

IntroductionVascular smooth muscle cells (VSMCs) are the predominant cell type in the medial layer of the aorta, which plays a critical role in aortic diseases. Innate immunity is the main driving force for cardiovascular diseases. MethodsTo determine the roles of innate immunity in VSMC and aortic pathologies, we performed transcriptome analyses on aortas from ApoE–/– angiotensin II (Ang II)-induced aortic aneurysm (AAA) time course, and ApoE–/– atherosclerosis time course, as well as VSMCs stimulated with danger-associated molecular patterns (DAMPs).ResultsWe made significant findings: 1) 95% and 45% of the upregulated innate immune pathways (UIIPs, based on data of 1226 innate immune genes) in ApoE–/– Ang II-induced AAA at 7 days were different from that of 14 and 28 days, respectively; and AAA showed twin peaks of UIIPs with a major peak at 7 days and a minor peak at 28 days; 2) all the UIIPs in ApoE–/– atherosclerosis at 6 weeks were different from that of 32 and 78 weeks (two waves); 3) analyses of additional 12 lists of innate immune-related genes with 1325 cytokine and chemokine genes, 2022 plasma membrane protein genes, 373 clusters of differentiation (CD) marker genes, 280 nuclear membrane protein genes, 1425 nucleoli protein genes, 6750 nucleoplasm protein genes, 1496 transcription factors (TFs) including 15 pioneer TFs, 164 histone modification enzymes, 102 oxidative cell death genes, 68 necrotic cell death genes, and 47 efferocytosis genes confirmed two-wave inflammation in atherosclerosis and twin-peak inflammation in AAA; 4) DAMPs-stimulated VSMCs were innate immune cells as judged by the upregulation of innate immune genes and genes from 12 additional lists; 5) DAMPs-stimulated VSMCs increased trans-differentiation potential by upregulating not only some of 82 markers of 7 VSMC-plastic cell types, including fibroblast, osteogenic, myofibroblast, macrophage, adipocyte, foam cell, and mesenchymal cell, but also 18 new cell types (out of 79 human cell types with 8065 cell markers); 6) analysis of gene deficient transcriptomes indicated that the antioxidant transcription factor NRF2 suppresses, however, the other five inflammatory transcription factors and master regulators, including AHR, NF-KB, NOX (ROS enzyme), PERK, and SET7 promote the upregulation of twelve lists of innate immune genes in atherosclerosis, AAA, and DAMP-stimulated VSMCs; and 7) both SET7 and trained tolerance-promoting metabolite itaconate contributed to twin-peak upregulation of cytokines in AAA. DiscussionOur findings have provided novel insights on the roles of innate immune responses and nuclear stresses in the development of AAA, atherosclerosis, and VSMC immunology and provided novel therapeutic targets for treating those significant cardiovascular and cerebrovascular diseases

Immunological and Metabolomic Impacts of Administration of Cry1Ab Protein and MON 810 Maize in Mouse

We have investigated the immunological and metabolomic impacts of Cry1Ab administration to mice, either as a purified protein or as the Cry1Ab-expressing genetically modified (GM) MON810 maize. Humoral and cellular specific immune responses induced in BALB/cJ mice after intra-gastric (i.g.) or intra-peritoneal (i.p.) administration of purified Cry1Ab were analyzed and compared with those induced by proteins of various immunogenic and allergic potencies. Possible unintended effects of the genetic modification on the pattern of expression of maize natural allergens were studied using IgE-immunoblot and sera from maize-allergic patients. Mice were experimentally sensitized (i.g. or i.p. route) with protein extracts from GM or non-GM maize, and then anti-maize proteins and anti-Cry1Ab–induced immune responses were analyzed. In parallel, longitudinal metabolomic studies were performed on the urine of mice treated via the i.g. route. Weak immune responses were observed after i.g. administration of the different proteins. Using the i.p. route, a clear Th2 response was observed with the known allergenic proteins, whereas a mixed Th1/Th2 immune response was observed with immunogenic protein not known to be allergenic and with Cry1Ab. This then reflects protein immunogenicity in the BALB/c Th2-biased mouse strain rather than allergenicity. No difference in natural maize allergen profiles was evidenced between MON810 and its non-GM comparator. Immune responses against maize proteins were quantitatively equivalent in mice treated with MON810 vs the non-GM counterpart and no anti-Cry1Ab–specific immune response was detected in mice that received MON810. Metabolomic studies showed a slight “cultivar” effect, which represented less than 1% of the initial metabolic information. Our results confirm the immunogenicity of purified Cry1Ab without evidence of allergenic potential. Immunological and metabolomic studies revealed slight differences in mouse metabolic profiles after i.g. administration of MON810 vs its non-GM counterpart, but no significant unintended effect of the genetic modification on immune responses was seen