-
Theranostics 2020The crosstalk between cardiac microvascular endothelial cells (CMECs) and cardiomyocytes (CMs) has emerged as a key component in the development of, and protection...
The crosstalk between cardiac microvascular endothelial cells (CMECs) and cardiomyocytes (CMs) has emerged as a key component in the development of, and protection against, cardiac diseases. For example, activation of endothelial nitric oxide synthase (eNOS) in CMECs, by therapeutic strategies such as ischemic preconditioning, plays a critical role in the protection against myocardial ischemia/reperfusion (I/R) injury. However, much less is known about the signals produced by CMs that are able to regulate CMEC biology. Here we uncovered one such mechanism using Tongxinluo (TXL), a traditional Chinese medicine, that alleviates myocardial ischemia/reperfusion (I/R) injury by activating CMEC eNOS. The aim of our study is to identify the signals produced by CMs that can regulate CMEC biology during I/R. and settings of ischemia-reperfusion were used in our study, with the protective signaling pathways activated in CMECs identified using genetic inhibition (p70s6k1 siRNA, miR-145-5p mimics, etc.), chemical inhibitors (the eNOS inhibitor, L-NNA, and the small extracellular vesicles (sEVs) inhibitor, GW4869) and Western blot analyses. TritonX-100 at a dose of 0.125% was utilized to inactivate the eNOS activity in endothelium to investigate the role of CMEC-derived eNOS in TXL-induced cardioprotection. We found that while CMEC-derived eNOS activity was required for the cardioprotection of TXL, activation of eNOS in CMECs by TXL did not occur directly. Instead, eNOS activation in CMECs required a crosstalk between CMs and CMECs through the uptake of CM-derived sEVs. We further demonstrate that TXL induced CM-sEVs contain increased levels of Long Intergenic Non-Protein Coding RNA, Regulator Of Reprogramming (Linc-ROR). Upon uptake into CMECs, linc-ROR downregulates its target miR-145-5p leading to activation of the eNOS pathway by facilitating the expression of p70s6k1 in these cells. The activation of CMEC-derived eNOS works to increase survival in both the CMECs and the CMs themselves. These data uncover a mechanism by which the crosstalk between CMs and CMECs leads to the increased survival of the heart after I/R injury and point to a new therapeutic target for the blunting of myocardial I/R injury.
Topics: Aniline Compounds; Animals; Benzylidene Compounds; Cardiotonic Agents; Cell Communication; Cells, Cultured; Coronary Vessels; Disease Models, Animal; Drugs, Chinese Herbal; Endothelial Cells; Endothelium, Vascular; Extracellular Vesicles; Humans; Isolated Heart Preparation; Male; Microvessels; Myocardial Reperfusion Injury; Myocytes, Cardiac; Nitric Oxide Synthase Type III; Nitroarginine; Rats; Signal Transduction
PubMed: 33052245
DOI: 10.7150/thno.43163 -
Turkish Journal of Medical Sciences Jun 2022Hydroxychloroquine (HCQ) is an antimalarial that is widely used in the management of rheumatoid arthritis and other autoimmune diseases. In this study, we aimed to...
BACKGROUND
Hydroxychloroquine (HCQ) is an antimalarial that is widely used in the management of rheumatoid arthritis and other autoimmune diseases. In this study, we aimed to examine the vascular effects of HCQ on rat aorta (RA).
METHODS
The RA rings were suspended in isolated organ baths and tension was recorded isometrically. HCQ-induced relaxations were tested in the presence of the nitric oxide synthase inhibitor, nitro-L-arginine methyl ester (L-NAME, 100 mM); the cyclooxygenase enzyme inhibitor, indomethacin (10 mM); the calcium (Ca2+) ion channel blocker, nilvadipine (10 μM); and the K+ ion channel inhibitors, tetraethylammonium (1 mM), glibenclamide (10 mM), 4-aminopyridine (1 mM), and barium chloride (30 mM). The effect of HCQ on Ca2+ channels was examined using Ca2+-free Krebs solution, and adding calcium chloride (CaCl2 , 10-5- 10-2 M) cumulatively to baths incubated with HCQ.
RESULTS
Removing the endothelium resulted in less relaxation of RA rings compared to endothelium-intact rings (p < 0.05). The effect of endothelium was supported by using L-NAME where HCQ produced-vasorelaxation was decreased (p < 0.05). The contraction of vascular rings was inhibited to a significant degree following the addition of CaCl2 , PE, or KCl on HCQ-incubated RA rings (p < 0.05). The incubation of the RA rings with the Ca2+ channel blocker, the K+ channel blockers, and the COX inhibitor, indomethacin did not significantly affect vascular relaxation induced by HCQ.
DISCUSSION
HCQ produced relaxation of RA rings. The relaxation mechanism differs according to the concentration of HCQ. At con-centrations of 10-6 and 10-5 M, the relaxation is endothelium-dependent and mediated by NO. We strongly suggest that Ca2+ channel inhibition is involved at concentrations of 10-5 and 10-4 M, as well as NO.
Topics: Rats; Animals; NG-Nitroarginine Methyl Ester; Hydroxychloroquine; Calcium Chloride; Endothelium; Indomethacin; Aorta; Endothelium, Vascular; Vasodilator Agents; Dose-Response Relationship, Drug
PubMed: 36326331
DOI: 10.55730/1300-0144.5382 -
Journal of the American Heart... Aug 2023Background The mechanisms determining vascular tone are still not completely understood, even though it is a significant factor in blood pressure management. Many...
Background The mechanisms determining vascular tone are still not completely understood, even though it is a significant factor in blood pressure management. Many circulating proteins have a significant impact on controlling vascular tone. Progranulin displays anti-inflammatory effects and has been extensively studied in neurodegenerative illnesses. We investigated whether progranulin sustains the vascular tone that helps regulate blood pressure. Methods and Results We used male and female C57BL6/J wild type (progranulin) and B6(Cg)-Grn/J (progranulin) to understand the impact of progranulin on vascular contractility and blood pressure. We found that progranulin mice display elevated blood pressure followed by hypercontractility to noradrenaline in mesenteric arteries, which is restored by supplementing the mice with recombinant progranulin. In ex vivo experiments, recombinant progranulin attenuated the vascular contractility to noradrenaline in male and female progranulin arteries, which was blunted by blocking EphrinA2 or Sortilin1. To understand the mechanisms whereby progranulin evokes anticontractile effects, we inhibited endothelial factors. N(gamma)-nitro-L-arginine methyl ester (nitric oxide synthase inhibitor) prevented the progranulin effects, whereas indomethacin (cyclooxygenase inhibitor) affected only the contractility in arteries incubated with vehicle, indicating that progranulin increases nitric oxide and decreases contractile prostanoids. Finally, recombinant progranulin induced endothelial nitric oxide synthase phosphorylation and nitric oxide production in isolated mesenteric endothelial cells. Conclusions Circulating progranulin regulates vascular tone and blood pressure via EphrinA2 and Sortilin1 receptors and endothelial nitric oxide synthase activation. Collectively, our data suggest that deficiency in progranulin is a cardiovascular risk factor and that progranulin might be a new therapeutic avenue to treat high blood pressure.
Topics: Male; Female; Mice; Animals; Nitric Oxide Synthase Type III; Blood Pressure; Progranulins; Nitric Oxide; Endothelial Cells; NG-Nitroarginine Methyl Ester; Mesenteric Arteries; Endothelium, Vascular; Norepinephrine
PubMed: 37581395
DOI: 10.1161/JAHA.123.030353 -
Physiology (Bethesda, Md.) Jan 2024The review deals with the release of endothelium-derived dopamine and 6-nitrodopamine (6-ND) and its effects on isolated vascular tissues and isolated hearts. Basal... (Review)
Review
The review deals with the release of endothelium-derived dopamine and 6-nitrodopamine (6-ND) and its effects on isolated vascular tissues and isolated hearts. Basal release of both dopamine and 6-ND is present in human isolated umbilical cord vessels, human popliteal vessels, nonhuman primate vessels, and reptilia aortas. The 6-ND basal release was significantly reduced when the tissues were treated with -nitro-l-arginine methyl ester and virtually abolished when the endothelium was mechanically removed. 6-Nitrodopamine is a potent vasodilator, and the mechanism of action responsible for this effect is the antagonism of dopamine D-like receptors. As a vasodilator, 6-ND constitutes a novel mechanism by which nitric oxide modulates vascular tone. The basal release of 6-ND was substantially decreased in endothelial nitric oxide synthase knockout (eNOS) mice and not altered in neuronal nitric oxide synthase knockout (nNOS) mice, indicating a nonneurogenic source for 6-ND in the heart. Indeed, in rat isolated right atrium, the release of 6-ND was not affected when the atria were treated with tetrodotoxin. In the rat isolated right atrium, 6-ND is the most potent endogenous positive chronotropic agent, and in Langendorff's heart preparation, it is the most potent endogenous positive inotropic agent. The positive chronotropic and inotropic effects of 6-ND are antagonized by β-adrenoceptor antagonists at concentrations that do not affect the effects induced by noradrenaline, adrenaline, and dopamine, indicating that blockade of the 6-ND receptor is the major modulator of heart chronotropism and inotropism. The review proposes that endothelium-derived catecholamines may constitute a major mechanism for control of vascular tone and heart functions, in contrast to the overrated role attributed to the autonomic nervous system.
Topics: Humans; Rats; Mice; Animals; Dopamine; Nitric Oxide Synthase; Enzyme Inhibitors; NG-Nitroarginine Methyl Ester; Cardiovascular System; Vasodilator Agents; Nitric Oxide; Endothelium
PubMed: 37874898
DOI: 10.1152/physiol.00020.2023 -
The European Journal of Neuroscience Feb 2020The gaseous free radical, nitric oxide (NO) acts as a ubiquitous neuromodulator, contributing to synaptic plasticity in a complex way that can involve either long term... (Review)
Review
The gaseous free radical, nitric oxide (NO) acts as a ubiquitous neuromodulator, contributing to synaptic plasticity in a complex way that can involve either long term potentiation or depression. It is produced by neuronal nitric oxide synthase (nNOS) which is presynaptically expressed and also located postsynaptically in the membrane and cytoplasm of a subpopulation of each major neuronal type in the ventral cochlear nucleus (VCN). We have used iontophoresis in vivo to study the effect of the NOS inhibitor L-NAME (L-NG-Nitroarginine methyl ester) and the NO donors SIN-1 (3-Morpholinosydnonimine hydrochloride) and SNOG (S-Nitrosoglutathione) on VCN units under urethane anaesthesia. Collectively, both donors produced increases and decreases in driven and spontaneous firing rates of some neurones. Inhibition of endogenous NO production with L-NAME evoked a consistent increase in driven firing rates in 18% of units without much effect on spontaneous rate. This reduction of gain produced by endogenous NO was mirrored when studying the effect of L-NAME on NMDA(N-Methyl-D-aspartic acid)-evoked excitation, with 30% of units showing enhanced NMDA-evoked excitation during L-NAME application (reduced NO levels). Approximately 25% of neurones contain nNOS and the NO produced can modulate the firing rate of the main principal cells: medium stellates (choppers), large stellates (onset responses) and bushy cells (primary-like responses). The main endogenous role of NO seems to be to partly suppress driven firing rates associated with NMDA channel activity but there is scope for it to increase neural gain if there were a pathological increase in its production following hearing loss.
Topics: Animals; Cochlear Nucleus; Enzyme Inhibitors; Guinea Pigs; NG-Nitroarginine Methyl Ester; Neurons; Nitric Oxide; Nitric Oxide Donors
PubMed: 31494975
DOI: 10.1111/ejn.14572 -
American Journal of Physiology. Heart... Mar 2016Mitochondrial dysfunction has been implicated as a cause of energy deprivation in heart failure (HF). Herein, we tested individual and combined effects of two pathogenic...
Mitochondrial dysfunction has been implicated as a cause of energy deprivation in heart failure (HF). Herein, we tested individual and combined effects of two pathogenic factors of nonischemic HF, inhibition of nitric oxide synthesis [with l-N(G)-nitroarginine methyl ester (l-NAME)] and hypertension [with angiotensin II (AngII)], on myocardial mitochondrial function, oxidative stress, and metabolic gene expression. l-NAME and AngII were administered individually and in combination to mice for 5 wk. Although all treatments increased blood pressure and reduced cardiac contractile function, the l-NAME + AngII group was associated with the most severe HF, as characterized by edema, hypertrophy, oxidative stress, increased expression of Nppa and Nppb, and decreased expression of Atp2a2 and Camk2b. l-NAME + AngII-treated mice exhibited robust deterioration of cardiac mitochondrial function, as observed by reduced respiratory control ratios in subsarcolemmal mitochondria and reduced state 3 levels in interfibrillar mitochondria for complex I but not for complex II substrates. Cardiac myofibrils showed reduced ADP-supported and oligomycin-inhibited oxygen consumption. Mitochondrial functional impairment was accompanied by reduced mitochondrial DNA content and activities of pyruvate dehydrogenase and complex I but increased H2O2 production and tissue protein carbonyls in hearts from AngII and l-NAME + AngII groups. Microarray analyses revealed the majority of the gene changes attributed to the l-NAME + AngII group. Pathway analyses indicated significant changes in metabolic pathways, such as oxidative phosphorylation, mitochondrial function, cardiac hypertrophy, and fatty acid metabolism in l-NAME + AngII hearts. We conclude that l-NAME + AngII is associated with impaired mitochondrial respiratory function and increased oxidative stress compared with either l-NAME or AngII alone, resulting in nonischemic HF.
Topics: Angiotensin II; Animals; Atrial Natriuretic Factor; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cardiomegaly; DNA, Mitochondrial; Electron Transport Complex I; Electron Transport Complex II; Enzyme Inhibitors; Gene Expression; Heart; Heart Failure; Hydrogen Peroxide; Mice; Mitochondria, Heart; Myocardium; NG-Nitroarginine Methyl Ester; Natriuretic Peptide, Brain; Natriuretic Peptide, C-Type; Nitric Oxide; Oxidative Stress; Protein Precursors; Pyruvate Dehydrogenase Complex; Reverse Transcriptase Polymerase Chain Reaction; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Vasoconstrictor Agents
PubMed: 26747502
DOI: 10.1152/ajpheart.00746.2015 -
PloS One 2022The endothelium maintains and regulates vascular homeostasis mainly by balancing interplay between vasorelaxation and vasoconstriction via regulating Nitric Oxide (NO)...
The endothelium maintains and regulates vascular homeostasis mainly by balancing interplay between vasorelaxation and vasoconstriction via regulating Nitric Oxide (NO) availability. Endothelial nitric oxide synthase (eNOS) is one of three NOS isoforms that catalyses the synthesis of NO to regulate endothelial function. However, eNOS's role in the regulation of endothelial function, such as cell proliferation and migration remain unclear. To gain a better understanding, we genetically knocked down eNOS in cultured endothelial cells using sieNOS and evaluated cell proliferation, migration and also tube forming potential in vitro. To our surprise, loss of eNOS significantly induced endothelial cell proliferation, which was associated with significant downregulation of both cell cycle inhibitor p21 and cell proliferation antigen Ki-67. Knockdown of eNOS induced cell migration but inhibited formation of tube-like structures in vitro. Mechanistically, loss of eNOS was associated with activation of MAPK/ERK and inhibition of PI3-K/AKT signaling pathway. On the contrary, pharmacologic inhibition of eNOS by inhibitors L-NAME or L-NMMA, inhibited cell proliferation. Genetic and pharmacologic inhibition of eNOS, both promoted endothelial cell migration but inhibited tube-forming potential. Our findings confirm that eNOS regulate endothelial function by inversely controlling endothelial cell proliferation and migration, and by directly regulating its tube-forming potential. Differential results obtained following pharmacologic versus genetic inhibition of eNOS indicates a more complex mechanism behind eNOS regulation and activity in endothelial cells, warranting further investigation.
Topics: Cells, Cultured; Endothelial Cells; Endothelium; Ki-67 Antigen; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase Type III; Protein Isoforms; Proto-Oncogene Proteins c-akt; omega-N-Methylarginine
PubMed: 36149900
DOI: 10.1371/journal.pone.0274487 -
Andrology Nov 2022Rat isolated vas deferens releases 6-nitrodopamine (6-ND), and the spasmogenic activity of this novel catecholamine is significantly reduced by tricyclic compounds such...
BACKGROUND
Rat isolated vas deferens releases 6-nitrodopamine (6-ND), and the spasmogenic activity of this novel catecholamine is significantly reduced by tricyclic compounds such as amitriptyline, desipramine, and carbamazepine and by antagonists of the α -adrenergic receptors such as doxazosin, tamsulosin, and prazosin.
OBJECTIVES
To investigate the liberation of 6-ND by human epididymal vas deferens (HEVDs) and its pharmacological actions.
METHODS
The in vitro liberation of 6-ND, dopamine, noradrenaline, and adrenaline from human vas deferens was evaluated by LC-MS/MS. The contractile effect of the catecholamines in HEVDs was investigated in vitro. The action of tricyclic antidepressants was evaluated on the spasmogenic activity ellicited by the catecholamines and by the electric-field stimulation (EFS). The tissue was also incubated with the inhibitor of nitric oxide (NO) synthase L-NAME and the release of catecholamines and the contractile response to EFS were assessed.
RESULTS
6-ND is the major catecholamine released from human vas deferens and its synthesis/release is inhibited by NO inhibition. The spasmogenic activity elicited by EFS in the human vas deferens was blocked by tricyclic antidepressants only at concentrations that selectively antagonize 6-ND induced contractions of the human vas deferens, without affecting the spasmogenic activity induced by dopamine, noradrenaline, and adrenaline in this tissue. Incubation of the vas deferens with L-NAME reduced both the 6-ND release and the contractions induced by EFS.
DISCUSSION AND CONCLUSION
6-ND should be considered a major endogenous modulator of human vas deferens contractility and possibly plays a pivotal role in the emission process of ejaculation. It offers a novel and shared mechanism of action for tricyclic antidepressants and α -adrenergic receptor antagonists.
Topics: Adrenergic Antagonists; Amitriptyline; Animals; Antidepressive Agents, Tricyclic; Carbamazepine; Chromatography, Liquid; Desipramine; Dopamine; Doxazosin; Epinephrine; Humans; Male; Muscle Contraction; Muscle, Smooth; NG-Nitroarginine Methyl Ester; Nitric Oxide; Norepinephrine; Prazosin; Rats; Receptors, Adrenergic; Tamsulosin; Tandem Mass Spectrometry; Vas Deferens
PubMed: 35934935
DOI: 10.1111/andr.13263 -
Clinical Science (London, England :... Dec 2022Hypertension (HTN) is associated with renal proinflammatory immune cell infiltration and increased sodium retention. We reported previously that renal lymphatic vessels,...
BACKGROUND
Hypertension (HTN) is associated with renal proinflammatory immune cell infiltration and increased sodium retention. We reported previously that renal lymphatic vessels, which are responsible for trafficking immune cells from the interstitial space to draining lymph nodes, increase in density under hypertensive conditions. We also demonstrated that augmenting renal lymphatic density can prevent HTN in mice. Whether renal lymphangiogenesis can treat HTN in mice is unknown. We hypothesized that genetically inducing renal lymphangiogenesis after the establishment of HTN would attenuate HTN in male and female mice from three different HTN models.
METHODS
Mice with inducible kidney-specific overexpression of VEGF-D (KidVD) experience renal lymphangiogenesis upon doxycycline administration. HTN was induced in KidVD+ and KidVD- mice by subcutaneous release of angiotensin II, administration of the nitric oxide synthase inhibitor L-NAME, or consumption of a 4% salt diet following a L-NAME priming and washout period. After a week of HTN stimuli treatment, doxycycline was introduced. Systolic blood pressure (SBP) readings were taken weekly. Kidney function was determined from urine and serum measures. Kidneys were processed for RT-qPCR, flow cytometry, and imaging.
RESULTS
Mice that underwent renal-specific lymphangiogenesis had significantly decreased SBP and renal proinflammatory immune cells. Additionally, renal lymphangiogenesis was associated with a decrease in sodium transporter expression and increased fractional excretion of sodium, indicating improved sodium handling efficiency.
CONCLUSIONS
These findings demonstrate that augmenting renal lymphangiogenesis can treat HTN in male and female mice by improving renal immune cell trafficking and sodium handling.
Topics: Mice; Male; Female; Animals; Lymphangiogenesis; NG-Nitroarginine Methyl Ester; Doxycycline; Hypertension; Kidney; Sodium
PubMed: 36345993
DOI: 10.1042/CS20220547 -
Lipid Emulsion Inhibits Amlodipine-Induced Nitric Oxide-Mediated Vasodilation in Isolated Rat Aorta.International Journal of Molecular... May 2023This study aimed to examine the effect of lipid emulsion on the vasodilation induced by a toxic dose of amlodipine in isolated rat aorta and elucidate its mechanism,...
This study aimed to examine the effect of lipid emulsion on the vasodilation induced by a toxic dose of amlodipine in isolated rat aorta and elucidate its mechanism, with a particular focus on nitric oxide. The effects of endothelial denudation, N-nitro-L-arginvine methyl ester (L-NAME), methylene blue, lipid emulsion, and linolenic acid on the amlodipine-induced vasodilation and amlodipine-induced cyclic guanosine monophosphate (cGMP) production were examined. Furthermore, the effects of lipid emulsion, amlodipine, and PP2, either alone or combined, on endothelial nitric oxide synthase (eNOS), caveolin-1, and Src-kinase phosphorylation were examined. Amlodipine-induced vasodilation was higher in endothelium-intact aorta than in endothelium-denuded aorta. L-NAME, methylene blue, lipid emulsion, and linolenic acid inhibited amlodipine-induced vasodilation and amlodipine-induced cGMP production in the endothelium-intact aorta. Lipid emulsion reversed the increased stimulatory eNOS (Ser1177) phosphorylation and decreased inhibitory eNOS (Thr495) phosphorylation induced via amlodipine. PP2 inhibited stimulatory eNOS, caveolin-1, and Src-kinase phosphorylation induced via amlodipine. Lipid emulsion inhibited amlodipine-induced endothelial intracellular calcium increase. These results suggest that lipid emulsion attenuated the vasodilation induced via amlodipine through inhibiting nitric oxide release in isolated rat aorta, which seems to be mediated via reversal of stimulatory eNOS (Ser1177) phosphorylation and inhibitory eNOS (Thr495) dephosphorylation, which are also induced via amlodipine.
Topics: Fat Emulsions, Intravenous; Nitric Oxide; Vasodilation; Aorta; Female; Animals; In Vitro Techniques; Amlodipine; Vasodilator Agents; NG-Nitroarginine Methyl Ester; Phospholipids; Soybean Oil; Nitric Oxide Synthase Type III
PubMed: 37240087
DOI: 10.3390/ijms24108741