Regulation of protein l-isoaspartyl methyltransferase by cell–matrix interactions: involvement of integrin αvβ3, PI 3-kinase, and the proteasome

2006 ◽  
Vol 84 (5) ◽  
pp. 684-694 ◽  
Author(s):  
Julie Lanthier ◽  
Richard R. Desrosiers
Keyword(s):  
Endothelial Cells ◽  
Molecular Mechanisms ◽  
Cell Attachment ◽  
Proteasome Inhibitors ◽  
Integrin Αvβ3 ◽  
Pi3k Pathway ◽  
Cell Aging ◽  
Cell Detachment ◽  
Aortic Endothelial Cells ◽  
Isoaspartyl Methyltransferase

The enzyme l-isoaspartyl methyltransferase (PIMT) is known to repair damaged proteins that have accumulated abnormal aspartyl residues during cell aging. However, little is known about the mechanisms involved in the regulation of PIMT expression. Here we report that PIMT expression in bovine aortic endothelial cells is regulated by cell detachment and readhesion to a substratum. During cell detachment, the PIMT level was rapidly and strongly increased and correlated with a stimulation of protein synthesis. Aside from endothelial cells, PIMT levels were also regulated by cell adhesion in various cancer cell lines. The upregulation of PIMT expression could be prevented by an anti-αvβ3 antibody (LM609) or by a cyclic RGD peptide (XJ735) specific to integrin αvβ3, indicating that this integrin was likely involved in PIMT regulation. Moreover, we found that PIMT expression returned to the basal level when cells were replated on a substratum after detachment, though downregulation of PIMT expression could be partly prevented by the PI3K inhibitors LY294002 and wortmannin, as well as by the proteasome inhibitors MG-132, lactacystin, and β-lactone. These findings support the assumption that the PIMT level was downregulated by proteasomal degradation, involving the PI3K pathway, during cell attachment. This study reports new insights on the molecular mechanisms responsible for the regulation of PIMT expression in cells. The regulation of PIMT level upon cell-substratum contact suggests a potential role for PIMT in biological processes such as wound healing, cell migration, and tumor metastasis dissemination.

scholarly journals Gangliosides Contribute to Vascular Insulin Resistance

2019 ◽  
Vol 20 (8) ◽  
pp. 1819 ◽  
Author(s):  
Norihiko Sasaki ◽  
Yoko Itakura ◽  
Masashi Toyoda
Keyword(s):  
Insulin Resistance ◽  
Endothelial Cells ◽  
Vascular Function ◽  
Molecular Mechanisms ◽  
The Elderly ◽  
Necrosis Factor Alpha ◽  
Aortic Endothelial Cells ◽  
Inflammatory Conditions ◽  
Factor Alpha

Insulin in physiological concentrations is important to maintain vascular function. Moreover, vascular insulin resistance contributes to vascular impairment. In the elderly, other factors including hypertension, dyslipidemia, and chronic inflammation amplify senescence of vascular endothelial and smooth muscle cells. In turn, senescence increases the risk for vascular-related diseases such as arteriosclerosis, diabetes, and Alzheimer’s disease. Recently, it was found that GM1 ganglioside, one of the glycolipids localized on the cell membrane, mediates vascular insulin resistance by promoting senescence and/or inflammatory stimulation. First, it was shown that increased GM1 levels associated with aging/senescence contribute to insulin resistance in human aortic endothelial cells (HAECs). Second, the expression levels of gangliosides were monitored in HAECs treated with different concentrations of tumor necrosis factor-alpha (TNFα) for different time intervals to mimic in vivo acute or chronic inflammatory conditions. Third, the levels of insulin signaling-related molecules were monitored in HAECs after TNFα treatment with or without inhibitors of ganglioside synthesis. In this review, we summarize the molecular mechanisms of insulin resistance in aged/senescent and TNFα-stimulated endothelial cells mediated by gangliosides and highlight the possible roles of gangliosides in vascular insulin resistance-related diseases.

Regulation of endothelial NO synthase expression by cyclosporin A in bovine aortic endothelial cells

1996 ◽  
Vol 271 (3) ◽  
pp. H1072-H1078 ◽  
Author(s):  
S. Lopez-Ongil ◽  
M. Saura ◽  
D. Rodriguez-Puyol ◽  
M. Rodriguez-Puyol ◽  
S. Lamas
Keyword(s):  
Endothelial Cells ◽  
Cyclosporin A ◽  
Molecular Mechanisms ◽  
Actinomycin D ◽  
No Synthase ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Threefold Increase ◽  
Vasoactive Factors ◽  
Recent Attention

The introduction of cyclosporin A (CsA) is considered a cornerstone advance in immunosuppression. However, serious side effects such as hypertension have fostered an important body of research regarding their pathophysiology. Although the participation of several vasoactive factors in the hypertensive response has been described, recent attention has focused on endothelium-derived vasoactive factors, and several reports describe an overproduction of endothelin-1 and a deficient endothelium-dependent vasodilation. In the case of the latter, no definitive clues for the precise molecular mechanisms have been provided. We demonstrate that endothelial cells in culture synthesize more NO in the presence of CsA for 24 h in a concentration-response manner. This augmentation is correlated with a threefold increase in the endothelial constitutive NO synthase (ecNOS) transcript, which is time dependent and maximal at 24 h. The CsA-induced increase in ecNOS mRNA expression was blocked by actinomycin D but unaltered by cycloheximide. Levels of ecNOS protein were also enhanced by CsA after 24 h. These data establish that NO synthesis is moderately enhanced in endothelial cells exposed to CsA for long periods of time and describe a new mode of regulating ecNOS gene expression.

Prostacyclin synthesis in irradiated endothelial cells cultured from bovine aorta

1985 ◽  
Vol 58 (2) ◽  
pp. 592-597 ◽  
Author(s):  
D. B. Rubin ◽  
E. A. Drab ◽  
C. H. Ts'ao ◽  
D. Gardner ◽  
W. F. Ward
Keyword(s):  
Endothelial Cells ◽  
Culture Media ◽  
Cell Detachment ◽  
Aortic Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Ldh Activity ◽  
Platelet Adherence ◽  
Subendothelial Matrix ◽  
60Co Gamma Rays ◽  
And Control

Confluent monolayers of bovine aortic endothelial cells were examined 2–72 h after exposure to 0.5–5.0 Gy of 60Co gamma-rays. Accumulation of prostacyclin [PGI2, measured as 6-ketoprostaglandin F1 alpha (6-keto-PGF1 alpha)] in the culture media and PGI2 production stimulated by exogenous arachidonate were correlated with cell detachment and release of lactate dehydrogenase (LDH) activity. Platelet adherence to irradiated and control monolayers also was studied. There were simultaneous time- and dose-dependent increases in cell detachment and in the titers of 6-keto-PGF1 alpha and LDH activity in the culture medium. These changes were evident between 4 and 8 h after 5 Gy or at 24 h after 0.5 Gy. Four hours after 5 Gy, both adherent and detached endothelial cells showed a twofold increase in PGI2 production during a 15-min incubation with arachidonate (10 microM). However, by 72 h this increase was less significant. The accumulation of 6-keto-PGF1 alpha appeared to be related to cell destruction, but radiation also stimulated PGI2 synthesis independent of cell detachment. There was an increased platelet interaction with irradiated monolayers, as a result of platelet adherence to subendothelial matrix exposed after cell detachment. However, irradiation did not alter the nonadherent property of the endothelial cell surface toward platelets.

CYCLOSPORIN A (CyA) INDUCED ENDOTHELIAL CELL INJURY

1987 ◽  
Author(s):  
C Zoja ◽  
L Furci ◽  
F Ghilardi ◽  
P Zilio ◽  
A Benigni ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Injury ◽  
Cell Damage ◽  
Chronic Administration ◽  
Endothelial Damage ◽  
Vascular Damage ◽  
Cell Detachment ◽  
Aortic Endothelial Cells ◽  
Specific Increase ◽  
Marked Cell

The chronic administration of CyA to animals and humans to prevent graft rejection may induce renal arteriolar damage resembling hemolytic uremic syndrome (HUS). This is a syndrome of vascular damage with thrombotic occlusions of the microcirculation. Endothelial damage is considered the first event in the pathogenetic cascade leading to HUS. We have used bovine aortic endothelial cells in culture to address the issue of CyA-induced arteriolar damage. CyA-induced a time (1-24 hours) and dose (1-50 μM) dependent cell damage. CyA-induced injury was characterized by an early cell detachment followed by lysis as documented by the increase in LDH and Cr release. 1 μM CyA did not induce cell detachment and lysis was evident only after prolonged incubations. 10 and 50 μM CyA both induced marked cell detachment and lysis: lysis started 3 hours after incubation of endothelial cells with CyA and was maximal at the end of 24 hour incubation (LDH release, percent specific increase over control values: 10 μM CyA, 47%; 50 μM CyA, 70%; 51 Cr release, percent specific increase over control values: 10 μM CyA, 28%; 50 μM CyA, 34%). CyA-induced injury was associated with dose- and time-depedent increase in prostacyclin (PGI2) and thromboxane A2 (TxA2) release by endothelial cells exposed to 10 and 50 μM CyA. CyA-induced generation of PGI2 and TxA2 was inhibited when the incubations were carried-on in the presence of acetyl salicilic acid (500 μM). These studies indicate that CyA exerts a direct toxic effect on endothelial cells and might help to understand the pathogenesis of CyA-induced vascular damage.

Abstract 927: The Role Of Peroxiredoxins As Mechanosensitive Antioxidants In Endothelial Cells

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Amy Mowbray ◽  
Sang Won Kang ◽  
Sue Goo Rhee ◽  
Hanjoong Jo
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Subcellular Localization ◽  
Molecular Mechanisms ◽  
Immunofluorescent Staining ◽  
Dependent Manner ◽  
Laminar Shear Stress ◽  
Arterial Tree ◽  
Aortic Endothelial Cells ◽  
Stress Dependent

Atherosclerosis is an inflammatory disease occurring primarily in curved or branching regions of the arterial tree where disturbed flow patterns, such as oscillation, exist. We have previously shown that oscillatory shear stress (OS) increases reactive oxygen species (ROS) levels in endothelial cells, while laminar shear stress (LS) reduces ROS compared to static controls. OS stimulation of ROS has been shown to occur in an NADPH oxidase-dependent manner. However, the mechanism by which LS removes ROS remains unclear. Peroxiredoxins (PRX) are a family of antioxidant proteins that have been linked to the prevention of oxidative stress and inflammation, but their role in atherosclerosis is unknown. Here, we hypothesize that shear stress regulates ROS levels in endothelial cells by controlling antioxidant peroxiredoxins. To test this hypothesis, bovine aortic endothelial cells (BAEC) were subjected to static, laminar, and oscillatory fluid flow conditions via cone-and-plate viscometer. Western blot analysis and immunofluorescent staining were used to evaluate the expression and subcellular localization of six known mammalian peroxiredoxins (PRX I-VI). Immunoblots indicated that BAEC express all six isoforms of peroxiredoxin proteins and that LS upregulated PRX I levels significantly compared to static controls and OS. Immunofluorescence also showed a distinct subcellular localization of each PRX: PRX I, II, IV, V and VI in the cytoplasm, PRX I, IV and V in the Golgi, PRX III in the mitochondria, and PRX I in the nucleus. These results indicate that peroxiredoxins are mechanosensitive antioxidants, removing ROS in a subcellular-specific manner. Based on these data, we suggest that peroxiredoxin antioxidants are likely involved in the molecular mechanisms that control shear stress-dependent atherosclerotic plaque development.

Quantification of eNOS mRNA in the canine cardiac vasculature by competitive PCR

2000 ◽  
Vol 278 (2) ◽  
pp. H658-H665 ◽  
Author(s):  
David Fulton ◽  
Andreas Papapetropoulos ◽  
Xiaoping Zhang ◽  
John D. Catravas ◽  
Thomas H. Hintze ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Coronary Artery ◽  
Left Ventricle ◽  
Molecular Mechanisms ◽  
Splenic Vein ◽  
Mrna Levels ◽  
Competitive Pcr ◽  
Aortic Endothelial Cells ◽  
Standard Curve ◽  
Enos Mrna

The goal of the present study was to develop a competitive PCR assay to measure changes in the expression of endothelial nitric oxide synthase (eNOS) mRNA levels throughout the canine vascular tree. A partial sequence of canine eNOS cDNA (1.86 kb), inducible NOS (1.95 kb), and neuronal NOS (1.16 kb) was cultured from canine aortic endothelial cells, LPS-treated canine splenic vein endothelial cells, and from canine left ventricle, respectively. Competitor eNOS cDNA (eNOS-C) was constructed via recombinant PCR. Thus, with the use of a standard curve competitive PCR with eNOS-C, the amount of eNOS mRNA in 500 ng of total RNA was greatest in the circumflex > right coronary artery > left anterior descending coronary artery > aorta. The isolation of coronary microvessels from the left ventricle was associated with an enrichment of endothelial cell markers such as eNOS, von Willebrand factor, and caveolin-1, an observation supported by the detection of up to 15-fold higher levels of eNOS mRNA in coronary microvessels relative to the larger arteries. The ability to quantify changes in eNOS mRNA levels throughout the canine vasculature should provide greater insight into the molecular mechanisms of how this gene is regulated in physiological and pathophysiological states.

scholarly journals Autocrine Bradykinin Release Promotes Ischemic Preconditioning-Induced Cytoprotection in Bovine Aortic Endothelial Cells

2020 ◽  
Vol 21 (8) ◽  
pp. 2965 ◽  
Author(s):  
Alessandro Bellis ◽  
Daniela Sorriento ◽  
Antonella Fiordelisi ◽  
Raffaele Izzo ◽  
Junichi Sadoshima ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Death ◽  
Ischemic Preconditioning ◽  
Molecular Mechanisms ◽  
Tissue Kallikrein ◽  
Aortic Endothelial Cells ◽  
Akt Activation ◽  
Bovine Aortic Endothelial Cells ◽  
Hoe 140 ◽  
Aortic Endothelial

The aims of this study were to assess whether ischemic preconditioning (PC) induces bradykinin (Bk) synthesis in bovine aortic endothelial cells (bAECs) and, if so, to explore the molecular mechanisms by which this peptide provides cytoprotection against hypoxia. PC was induced by exposing bAECs to three cycles of 15 min of hypoxia followed by 15 min of reoxygenation. Bk synthesis peaked in correspondence to the early and late phases of PC (10−12 M and 10−11 M, respectively) and was abolished by a selective tissue kallikrein inhibitor, aprotinin. Stimulation with exogenous Bk at concentrations of 10−12 M and 10−11 M reduced the cell death induced by 12 h of hypoxia by 50%. Pretreatment with HOE−140, a Bk receptor 2 (BKR2) inhibitor, in bAECs exposed to 12 h of hypoxia, abrogated the cytoprotective effect of early and late PC, whereas des-Arg-HOE-140, a Bk receptor 1 (BKR1) inhibitor, affected only the late PC. In addition, we found that PC evoked endocytosis and the recycling of BKR2 during both the early and late phases, and that inhibition of these pathways affected PC-mediated cytoprotection. Finally, we evaluated the activation of PKA and Akt in the presence or absence of BKR2 inhibitor. HOE-140 abrogated PKA and Akt activation during both early and late PC. Consistently, BKR2 inhibition abolished cross-talk between PKA and Akt in PC. In bAECs, Bk-synthesis evoked by PC mediates the protection against both apoptotic and necrotic hypoxia-induced cell death in an autocrine manner, by both BKR2- and BKR1-dependent mechanisms.

Expression of cathepsin K is regulated by shear stress in cultured endothelial cells and is increased in endothelium in human atherosclerosis

2007 ◽  
Vol 292 (3) ◽  
pp. H1479-H1486 ◽  
Author(s):  
Manu O. Platt ◽  
Randall F. Ankeny ◽  
Guo-Ping Shi ◽  
Daiana Weiss ◽  
J. D. Vega ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Molecular Mechanisms ◽  
Cathepsin K ◽  
Immunohistochemical Analysis ◽  
Cysteine Proteases ◽  
Laminar Shear Stress ◽  
Gelatinase Activity ◽  
Aortic Endothelial Cells ◽  
Expression And Activity

Cathepsins, the lysosomal cysteine proteases, are involved in vascular remodeling and atherosclerosis. Genetic knockout of cathepsins S and K in mice has shown to reduce atherosclerosis, although the molecular mechanisms remain unclear. Because atherosclerosis preferentially occurs in arteries exposed to disturbed flow conditions, we hypothesized that shear stress would regulate cathepsin K expression and activity in endothelial cells. Mouse aortic endothelial cells (MAEC) exposed to proatherogenic oscillatory shear (OS, ± 5 dyn/cm2 for 1 day) showed significantly higher cathepsin K expression and activity than that of atheroprotective, unidirectional laminar shear stress (LS, 15 dyn/cm2 for 1 day). Western blot and active-site labeling studies showed an active, mature form of cathepsin K in the conditioned medium of MAEC exposed to OS but not in that of LS. Functionally, MAEC exposed to OS significantly increased elastase and gelatinase activity above that of LS. The OS-dependent elastase and gelatinase activities were significantly reduced by knocking down cathepsin K with small-interfering (si) RNA, but not by a nonsilencing siRNA control, suggesting that cathepsin K is a shear-sensitive protease. In addition, immunohistochemical analysis of atherosclerotic human coronary arteries showed a positive correlation between the cathepsin K expression levels in endothelium and elastic lamina integrity. These findings suggest that cathepsin K is a mechanosensitive, extracellular matrix protease that, in turn, may be involved in arterial wall remodeling and atherosclerosis.

scholarly journals Linoleyl hydroperoxide transcriptionally upregulates heme oxygenase-1 gene expression in human renal epithelial and aortic endothelial cells.

1998 ◽  
Vol 9 (11) ◽  
pp. 1990-1997
Author(s):  
A Agarwal ◽  
F Shiraishi ◽  
G A Visner ◽  
H S Nick
Keyword(s):  
Endothelial Cells ◽  
Renal Disease ◽  
Heme Oxygenase ◽  
Molecular Mechanisms ◽  
De Novo ◽  
Oxidized Ldl ◽  
Heme Oxygenase 1 ◽  
Aortic Endothelial Cells ◽  
Mrna Induction ◽  
Aortic Endothelial

Atherogenic lipoproteins such as oxidized LDL are implicated in the pathogenesis of atherosclerosis and renal disease. Fatty acid hydroperoxides and phospholipids such as linoleyl hydroperoxide (LAox or 13-HPODE) and lysophosphatidylcholine (lyso-PC), abundant components of oxidized LDL, mediate the effects of atherogenic lipids. Oxidized LDL has been shown to induce heme oxygenase-1 (HO-1), a microsomal enzyme that is involved in heme detoxification and is a major endogenous source of carbon monoxide. HO-1 is also induced by many other stimuli that shift cellular redox. To identify the constituents and molecular mechanisms of oxidized LDL-mediated HO-1 induction, human renal epithelial cells and aortic endothelial cells were exposed to LAox and lyso-PC. Exposure to LAox (25 microM) showed an approximately 16-fold induction of HO-1 mRNA, whereas exposure to lyso-PC (25 microM) showed only an approximate 2.6-fold increase. Treatment with actinomycin-D (4 microM), a transcriptional inhibitor, as well as nuclear run-on assays, demonstrated that LAox-mediated HO-1 gene induction is dependent on de novo transcription. Cycloheximide did not affect LAox-mediated HO-1 mRNA induction, suggesting that new protein synthesis is not required for transcriptional induction. Transfection of a human HO-1 promoter-reporter gene construct showed that LAox upregulation of HO-1 occurs via mechanisms different from those of known inducers, heme and cadmium. These studies are the first demonstration that LAox induces HO-1 by transcriptional mechanisms and may have implications in the pathogenesis of cell injury in atherosclerosis and progressive renal disease.

scholarly journals Bisphenol A Induces Accelerated Cell Aging in Murine Endothelium

Biomolecules ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 1429
Author(s):  
Rafael Moreno-Gómez-Toledano ◽  
Sandra Sánchez-Esteban ◽  
Alberto Cook ◽  
Marta Mínguez-Moratinos ◽  
Rafael Ramírez-Carracedo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Bisphenol A ◽  
Vascular Endothelium ◽  
Cell Aging ◽  
Western Blots ◽  
Aortic Endothelial Cells ◽  
Response System ◽  
Senescent Phenotype ◽  
Aging Mechanisms ◽  
Protein Response

Bisphenol A (BPA) is a widespread endocrine disruptor affecting many organs and systems. Previous work in our laboratory demonstrated that BPA could induce death due to necroptosis in murine aortic endothelial cells (MAECs). This work aims to evaluate the possible involvement of BPA-induced senescence mechanisms in endothelial cells. The β-Gal assays showed interesting differences in cell senescence at relatively low doses (100 nM and 5 µM). Western blots confirmed that proteins involved in senescence mechanisms, p16 and p21, were overexpressed in the presence of BPA. In addition, the UPR (unfolding protein response) system, which is part of the senescent phenotype, was also explored by Western blot and qPCR, confirming the involvement of the PERK-ATF4-CHOP pathway (related to pathological processes). The endothelium of mice treated with BPA showed an evident increase in the expression of the proteins p16, p21, and CHOP, confirming the results observed in cells. Our results demonstrate that oxidative stress induced by BPA leads to UPR activation and senescence since pretreatment with N-acetylcysteine (NAC) in BPA-treated cells reduced the percentage of senescent cells prevented the overexpression of proteins related to BPA-induced senescence and reduced the activation of the UPR system. The results suggest that BPA participates actively in accelerated cell aging mechanisms, affecting the vascular endothelium and promoting cardiovascular diseases.

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