RESUMO
The Coronavirus Disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) and its sublineages pose a new challenge to healthcare systems worldwide due to its ability to efficiently spread in immunized populations and its resistance to currently available therapies. COVID-19, although targeting primarily the respiratory system, is also now well established that later affects every organ in the body. Most importantly, despite the available therapy and vaccine-elicited protection, the long-term consequences of viral infection in breakthrough and asymptomatic individuals are areas of concern. In the past two years, investigators accumulated evidence on how the virus triggers our immune system and the molecular signals involved in the cross-talk between immune cells and structural cells in the pulmonary vasculature to drive pathological lung complications such as endothelial dysfunction and thrombosis. In the review, we emphasize recent updates on the pathophysiological inflammatory and immune responses associated with SARS-CoV-2 infection and their potential long-term consequences that may consequently lead to the development of pulmonary vascular diseases.
Assuntos
COVID-19 , Coinfecção , Humanos , SARS-CoV-2 , Pulmão , Reações CruzadasRESUMO
Reduced expression and/or activity of Kv1.5 channels (encoded by KCNA5) is a common hallmark in human or experimental pulmonary arterial hypertension (PAH). Likewise, genetic variants in KCNA5 have been found in patients with PAH, but their functional consequences and potential impact on the disease are largely unknown. Herein, this study aimed to characterize the functional consequences of seven KCNA5 variants found in a cohort of patients with PAH. Potassium currents were recorded by patch-clamp technique in HEK293 cells transfected with wild-type or mutant Kv1.5 cDNA. Flow cytometry, Western blot, and confocal microscopy techniques were used for measuring protein expression and cell apoptosis in HEK293 and human pulmonary artery smooth muscle cells. KCNA5 variants (namely, Arg184Pro and Gly384Arg) found in patients with PAH resulted in a clear loss of potassium channel function as assessed by electrophysiological and molecular modeling analyses. The Arg184Pro variant also resulted in a pronounced reduction of Kv1.5 expression. Transfection with Arg184Pro or Gly384Arg variants decreased apoptosis of human pulmonary artery smooth muscle cells compared with the wild-type cells, demonstrating that KCNA5 dysfunction in both variants affects cell viability. Thus, in addition to affecting channel activity, both variants were associated with impaired apoptosis, a crucial process linked to the disease. The estimated prevalence of dysfunctional KCNA5 variants in the PAH population analyzed was around 1%. The data indicate that some KCNA5 variants found in patients with PAH have critical consequences for channel function, supporting the idea that KCNA5 pathogenic variants may be a causative or contributing factor for PAH.
Assuntos
Hipertensão Pulmonar , Hipertensão Arterial Pulmonar , Humanos , Hipertensão Arterial Pulmonar/metabolismo , Células HEK293 , Hipertensão Pulmonar/metabolismo , Canal de Potássio Kv1.5/genética , Canal de Potássio Kv1.5/metabolismo , Hipertensão Pulmonar Primária Familiar/metabolismo , Artéria Pulmonar/patologiaRESUMO
Objective: Atheromatous fibrous caps are produced by smooth muscle cells (SMCs) that are recruited to the subendothelial space. We tested whether the recruitment mechanisms are the same as in embryonic artery development, which relies prominently on Notch signaling to form the subendothelial medial SMC layers. Approach and Results: Notch elements were expressed in regions of fibrous cap in human and mouse plaques. To assess the causal role of Notch signaling in cap formation, we studied atherosclerosis in mice where the Notch pathway was inactivated in SMCs by conditional knockout of the essential effector transcription factor RBPJ (recombination signal-binding protein for immunoglobulin kappa J region). The recruitment of cap SMCs was significantly reduced without major effects on plaque size. Lineage tracing revealed the accumulation of SMC-derived plaque cells in the cap region was unaltered but that Notch-defective cells failed to re-acquire the SMC phenotype in the cap. Conversely, to analyze whether the loss of Notch signaling is required for SMC-derived cells to accumulate in atherogenesis, we studied atherosclerosis in mice with constitutive activation of Notch signaling in SMCs achieved by conditional expression of the Notch intracellular domain. Forced Notch signaling inhibited the ability of medial SMCs to contribute to plaque cells, including both cap SMCs and osteochondrogenic cells, and significantly reduced atherosclerosis development. Conclusions: Sequential loss and gain of Notch signaling is needed to build the cap SMC population. The shared mechanisms with embryonic arterial media assembly suggest that the cap forms as a neo-media that restores the connection between endothelium and subendothelial SMCs, transiently disrupted in early atherogenesis.
Assuntos
Aterosclerose/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Placa Aterosclerótica , Receptores Notch/metabolismo , Túnica Média/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Artérias/metabolismo , Artérias/patologia , Aterosclerose/genética , Aterosclerose/patologia , Linhagem da Célula , Células Cultivadas , Progressão da Doença , Fibrose , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Fenótipo , Ratos , Receptores Notch/genética , Transdução de Sinais , Túnica Média/patologiaRESUMO
Vitamin D (VitD) receptor regulates the expression of several genes involved in signaling pathways affected in pulmonary hypertension (PH). VitD deficiency is highly prevalent in PH, and low levels are associated with poor prognosis. We investigated if VitD deficiency may predispose to or exacerbate PH. Male Wistar rats were fed with a standard or a VitD-free diet for 5 wk. Next, rats were further divided into controls or PH, which was induced by a single dose of Su-5416 (20 mg/kg) and exposure to hypoxia (10% O2) for 2 wk. VitD deficiency had no effect on pulmonary pressure in normoxic rats, indicating that, by itself, it does not trigger PH. However, it induced several moderate but significant changes characteristic of PH in the pulmonary arteries, such as increased muscularization, endothelial dysfunction, increased survivin, and reduced bone morphogenetic protein (Bmp) 4, Bmp6, DNA damage-inducible transcript 4, and K+ two-pore domain channel subfamily K member 3 (Kcnk3) expression. Myocytes isolated from pulmonary arteries from VitD-deficient rats had a reduced whole voltage-dependent potassium current density and acid-sensitive (TASK-like) potassium currents. In rats with PH induced by Su-5416 plus hypoxia, VitD-free diet induced a modest increase in pulmonary pressure, worsened endothelial function, increased the hyperreactivity to serotonin, arterial muscularization, decreased total and TASK-1 potassium currents, and further depolarized the pulmonary artery smooth muscle cell membrane. In human pulmonary artery smooth muscle cells from controls and patients with PH, the active form of VitD calcitriol significantly increased KCNK3 mRNA expression. Altogether, these data strongly suggest that the deficit in VitD induces pulmonary vascular dysfunction.
Assuntos
Hipertensão Pulmonar/metabolismo , Músculo Liso Vascular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Deficiência de Vitamina D/metabolismo , Animais , Humanos , Pulmão/metabolismo , Pulmão/fisiopatologia , Masculino , Potenciais da Membrana/fisiologia , Músculo Liso Vascular/fisiopatologia , Miócitos de Músculo Liso/metabolismo , Ratos Wistar , Vitamina D/metabolismoRESUMO
KEY POINTS: The expression of miR-1 is increased in lungs from the Hyp/Su5416 PAH rat model. Pulmonary artery smooth muscle cells from this animal model are more depolarized and show decreased expression and activity of voltage-dependent potassium channel (Kv)1.5. miR-1 directly targets Kv1.5 channels, reduces Kv1.5 activity and induces membrane depolarization. Antagomir-1 prevents Kv1.5 channel downregulation and the depolarization induced by hypoxia/Su5416 exposition. ABSTRACT: Impairment of the voltage-dependent potassium channel (Kv) plays a central role in the development of cardiovascular diseases, including pulmonary arterial hypertension (PAH). MicroRNAs are non-coding RNAs that regulate gene expression by binding to the 3'-untranslated region region of specific mRNAs. The present study aimed to analyse the effects of miR-1 on Kv channel function in pulmonary arteries (PA). Kv channel activity was studied in PA from healthy animals transfected with miR-1 or scrambled-miR. Kv currents were studied using the whole-cell configuration of the patch clamp technique. The characterization of the Kv1.5 currents was performed with the selective inhibitor DPO-1. miR-1 expression was increased and Kv1.5 channels were decreased in lungs from a rat model of PAH induced by hypoxia and Su5416. miR-1 transfection increased cell capacitance, reduced Kv1.5 currents and induced membrane depolarization in isolated pulmonary artery smooth muscle cells. A luciferase reporter assay indicated that KCNA5, which encodes Kv1.5 channels, is a direct target gene of miR-1. Incubation of PA with Su5416 and hypoxia (3% O2 ) increased miR-1 and induced a decline in Kv1.5 currents, which was prevented by antagomiR-1. In conclusion, these data indicate that miR-1 induces pulmonary artery smooth muscle cell hypertrophy and reduces the activity and expression of Kv channels, suggesting a pathophysiological role in PAH.
Assuntos
Hipertensão Pulmonar/metabolismo , Canal de Potássio Kv1.5/metabolismo , MicroRNAs/metabolismo , Artéria Pulmonar/metabolismo , Potenciais de Ação , Animais , Células COS , Hipóxia Celular , Chlorocebus aethiops , Regulação para Baixo , Hipertensão Pulmonar/etiologia , Indóis/toxicidade , Canal de Potássio Kv1.5/genética , Masculino , MicroRNAs/genética , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/fisiologia , Bloqueadores dos Canais de Potássio/farmacologia , Artéria Pulmonar/efeitos dos fármacos , Artéria Pulmonar/fisiopatologia , Pirróis/toxicidade , Ratos , Ratos WistarRESUMO
Human immunodeficiency virus (HIV) infection is an established risk factor for pulmonary arterial hypertension (PAH); however, the pathogenesis of HIV-related PAH remains unclear. Since K+ channel dysfunction is a common marker in most forms of PAH, our aim was to analyze whether the expression of HIV proteins is associated with impairment of K+ channel function in the pulmonary vascular bed. HIV transgenic mice (Tg26) expressing seven of the nine HIV viral proteins and wild-type (WT) mice were used. Hemodynamic assessment was performed by echocardiography and catheterization. Vascular reactivity was studied in endothelium-intact pulmonary arteries. K+ currents were recorded in freshly isolated pulmonary artery smooth muscle cells (PASMC) using the patch-clamp technique. Gene expression was assessed using quantitative RT-PCR. PASMC from Tg26 mice had reduced K+ currents and were more depolarized than those from WT. Whereas voltage-gated K+ channel 1.5 (Kv1.5) currents were preserved, pH-sensitive noninactivating background currents ( IKN) were nearly abolished in PASMC from Tg26 mice. Tg26 mice had reduced lung expression of Kv7.1 and Kv7.4 channels and decreased responses to the Kv7.1 channel activator L-364,373 assessed by vascular reactivity and patch-clamp experimental approaches. Although we found pulmonary vascular remodeling and endothelial dysfunction in Tg26 mice, this was not accompanied by changes in hemodynamic parameters. In conclusion, the expression of HIV proteins in vivo impairs pH-sensitive IKN and Kv7 currents. This negative impact of HIV proteins in K+ channels was not sufficient to induce PAH, at least in mice, but may play a permissive or accessory role in the pathophysiology of HIV-associated PAH.
Assuntos
HIV-1/genética , Proteínas do Vírus da Imunodeficiência Humana/metabolismo , Hipertrofia Ventricular Direita/patologia , Músculo Liso Vascular/patologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Artéria Pulmonar/patologia , Transgenes/fisiologia , Animais , Infecções por HIV/genética , Infecções por HIV/metabolismo , Infecções por HIV/virologia , Proteínas do Vírus da Imunodeficiência Humana/genética , Humanos , Hipertrofia Ventricular Direita/metabolismo , Masculino , Potenciais da Membrana , Camundongos , Camundongos Transgênicos , Músculo Liso Vascular/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Artéria Pulmonar/metabolismo , VasoconstriçãoRESUMO
BACKGROUND: Pulmonary hypertension (PH) is frequently observed in patients with acute respiratory distress syndrome (ARDS) and it is associated with an increased risk of mortality. Both acid sphingomyelinase (aSMase) activity and interleukin 6 (IL-6) levels are increased in patients with sepsis and correlate with worst outcomes, but their role in pulmonary vascular dysfunction pathogenesis has not yet been elucidated. Therefore, the aim of this study was to determine the potential contribution of aSMase and IL-6 in the pulmonary vascular dysfunction induced by lipopolysaccharide (LPS). METHODS: Rat or human pulmonary arteries (PAs) or their cultured smooth muscle cells (SMCs) were exposed to LPS, SMase or IL-6 in the absence or presence of a range of pharmacological inhibitors. The effects of aSMase inhibition in vivo with D609 on pulmonary arterial pressure and inflammation were assessed following intratracheal administration of LPS. RESULTS: LPS increased ceramide and IL-6 production in rat pulmonary artery smooth muscle cells (PASMCs) and inhibited pulmonary vasoconstriction induced by phenylephrine or hypoxia (HPV), induced endothelial dysfunction and potentiated the contractile responses to serotonin. Exogenous SMase and IL-6 mimicked the effects of LPS on endothelial dysfunction, HPV failure and hyperresponsiveness to serotonin in PA; whereas blockade of aSMase or IL-6 prevented LPS-induced effects. Finally, administration of the aSMase inhibitor D609 limited the development of endotoxin-induced PH and ventilation-perfusion mismatch. The protective effects of D609 were validated in isolated human PAs. CONCLUSIONS: Our data indicate that aSMase and IL-6 are not simply biomarkers of poor outcomes but pathogenic mediators of pulmonary vascular dysfunction in ARDS secondary to Gram-negative infections.
Assuntos
Hipertensão Pulmonar/metabolismo , Interleucina-6/metabolismo , Interleucina-6/farmacologia , Artéria Pulmonar/metabolismo , Esfingomielina Fosfodiesterase/metabolismo , Esfingomielina Fosfodiesterase/farmacologia , Animais , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Células Cultivadas , Ceramidases/metabolismo , Modelos Animais de Doenças , Imunofluorescência , Humanos , Interleucina-1beta/metabolismo , Lipopolissacarídeos , Masculino , Músculo Liso Vascular/citologia , Norbornanos , Ratos , Ratos Wistar , Tiocarbamatos , Tionas/farmacologia , Vasoconstrição/efeitos dos fármacosRESUMO
PPARß/δ activation protects against endothelial dysfunction in diabetic models. Elevated glucose is known to impair cAMP-induced relaxation and Kv channel function in coronary arteries (CA). Herein, we aimed to analyse the possible protective effects of the PPARß/δ agonist GW0742 on the hyperglycaemic-induced impairment of cAMP-induced relaxation and Kv channel function in rat CA. As compared with low glucose (LG), incubation under high glucose (HG) conditions attenuated the relaxation induced by the adenylate cyclase activator forskolin in CA and this was prevented by GW0742. The protective effect of GW0742 was supressed by a PPARß/δ antagonist. In myocytes isolated from CA under LG, forskolin enhanced Kv currents and induced hyperpolarization. In contrast, when CA were incubated with HG, Kv currents were diminished and the electrophysiological effects of forskolin were abolished. These deleterious effects were prevented by GW0742. The protective effects of GW0742 on forskolin-induced relaxation and Kv channel function were confirmed in CA from type-1 diabetic rats. In addition, the differences in the relaxation induced by forskolin in CA incubated under LG, HG or HG + GW0742 were abolished by the Kv7 channel inhibitor XE991. Accordingly, GW0742 prevented the down-regulation of Kv7 channels induced by HG. Finally, the preventive effect of GW0742 on oxidative stress and cAMP-induced relaxation were overcome by the pyruvate dehydrogenase kinase 4 (PDK4) inhibitor dichloroacetate (DCA). Our results reveal that the PPARß/δ agonist GW0742 prevents the impairment of the cAMP-mediated relaxation in CA under HG. This protective effect was associated with induction of PDK4, attenuation of oxidative stress and preservation of Kv7 channel function.
Assuntos
Vasos Coronários/metabolismo , AMP Cíclico/metabolismo , Hiperglicemia/metabolismo , Canal de Potássio KCNQ1/metabolismo , PPAR delta/metabolismo , PPAR beta/metabolismo , Animais , Vasos Coronários/efeitos dos fármacos , Vasos Coronários/fisiopatologia , Diabetes Mellitus Experimental , Humanos , Hiperglicemia/genética , Canal de Potássio KCNQ1/genética , Masculino , PPAR delta/genética , PPAR beta/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Piruvato Desidrogenase Quinase de Transferência de Acetil , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Tiazóis/administração & dosagem , Vasodilatação/efeitos dos fármacosRESUMO
BACKGROUND: We hypothesized that treatment with quercetin could result in improved hemodynamics, lung inflammatory parameters and mortality in a rat model of hemorrhagic shock. METHODS: Rats were anesthetized (80 mg/kg ketamine plus 8 mg/kg xylazine i.p.). The protocol included laparotomy for 15 min (trauma), hemorrhagic shock (blood withdrawal to reduce the mean arterial pressure to 35 mmHg) for 75 min and resuscitation by re-infusion of all the shed blood plus lactate Ringer for 90 min. Intravenous quercetin (50 mg/kg) or vehicle were administered during resuscitation. RESULTS: There was a trend for increased survival 84.6% (11/13) in the treated group vs. the shock group 68.4% (13/19, p > 0.05 Kaplan-Meier). Quercetin fully prevented the development of lung edema. The activity of aSMase was increased in the shock group compared to the sham group and the quercetin prevented this effect. However, other inflammatory markers such as myeloperoxidase activity, interleukin-6 in plasma or bronchoalveolar fluid were similar in the sham and shock groups. We found no bacterial DNA in plasma in these animals. CONCLUSIONS: Quercetin partially prevented the changes in blood pressure and lung injury in shock associated to hemorrhage and reperfusion.
Assuntos
Quercetina/uso terapêutico , Choque Hemorrágico/tratamento farmacológico , Choque Traumático/tratamento farmacológico , Animais , Pressão Arterial/efeitos dos fármacos , Biomarcadores/sangue , Edema/prevenção & controle , Hemodinâmica , Inflamação/complicações , Inflamação/tratamento farmacológico , Interleucina-6/química , Soluções Isotônicas/uso terapêutico , Masculino , Peroxidase/química , Edema Pulmonar/complicações , Edema Pulmonar/tratamento farmacológico , Ratos , Ratos Wistar , Reperfusão , Ressuscitação , Lactato de Ringer , Choque Hemorrágico/complicações , Choque Traumático/complicaçõesRESUMO
OBJECTIVE: Voltage-dependent K(+) (Kv) channels from the Kv7 family are expressed in blood vessels and contribute to cardiovascular physiology. Although Kv7 channel blockers trigger muscle contractions, Kv7 activators act as vasorelaxants. Kv7.1 and Kv7.5 are expressed in many vessels. Kv7.1 is under intense investigation because Kv7.1 blockers fail to modulate smooth muscle reactivity. In this study, we analyzed whether Kv7.1 and Kv7.5 may form functional heterotetrameric channels increasing the channel diversity in vascular smooth muscles. APPROACH AND RESULTS: Kv7.1 and Kv7.5 currents elicited in arterial myocytes, oocyte, and mammalian expression systems suggest the formation of heterotetrameric complexes. Kv7.1/Kv7.5 heteromers, exhibiting different pharmacological characteristics, participate in the arterial tone. Kv7.1/Kv7.5 associations were confirmed by coimmunoprecipitation, fluorescence resonance energy transfer, and fluorescence recovery after photobleaching experiments. Kv7.1/Kv7.5 heterotetramers were highly retained at the endoplasmic reticulum. Studies in HEK-293 cells, heart, brain, and smooth and skeletal muscles demonstrated that the predominant presence of Kv7.5 stimulates release of Kv7.1/Kv7.5 oligomers out of lipid raft microdomains. Electrophysiological studies supported that KCNE1 and KCNE3 regulatory subunits further increased the channel diversity. Finally, the analysis of rat isolated myocytes and human blood vessels demonstrated that Kv7.1 and Kv7.5 exhibited a differential expression, which may lead to channel diversity. CONCLUSIONS: Kv7.1 and Kv7.5 form heterotetrameric channels increasing the diversity of structures which fine-tune blood vessel reactivity. Because the lipid raft localization of ion channels is crucial for cardiovascular physiology, Kv7.1/Kv7.5 heteromers provide efficient spatial and temporal regulation of smooth muscle function. Our results shed light on the debate about the contribution of Kv7 channels to vasoconstriction and hypertension.
Assuntos
Canais de Potássio KCNQ/metabolismo , Canal de Potássio KCNQ1/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Potássio/metabolismo , Animais , Células COS , Chlorocebus aethiops , Células HEK293 , Humanos , Canais de Potássio KCNQ/química , Canais de Potássio KCNQ/efeitos dos fármacos , Canais de Potássio KCNQ/genética , Canal de Potássio KCNQ1/química , Canal de Potássio KCNQ1/efeitos dos fármacos , Canal de Potássio KCNQ1/genética , Microdomínios da Membrana/metabolismo , Potenciais da Membrana , Músculo Liso Vascular/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Estrutura Quaternária de Proteína , Ratos , Transfecção , XenopusRESUMO
An effective pulmonary hypertension (PH) treatment should combine antiproliferative and vasodilator effects. We characterized a wide-range of drugs comparing their anti-proliferative vs vasodilator effects in human and rat pulmonary artery smooth muscle cells (PASMC). Key findings: 1) Approved PH drugs (PDE5 inhibitors, sGC stimulators and PGI2 agonists) are preferential vasodilators. 2) cGMP stimulators were more effective in cells derived from hypertensive rats. 3) Nifedipine acted equally as vasodilator and antiproliferative. 4) quercetin and imatinib were potent dual vasodilator/antiproliferative drugs. 5) Tacrolimus and levosimendan lacked antiproliferative effects. 6) Forskolin, pinacidil and hydroxyfasudil were more effective as antiproliferative in human cells.
Assuntos
Proliferação de Células , Hipertensão Pulmonar , Miócitos de Músculo Liso , Artéria Pulmonar , Vasodilatadores , Animais , Humanos , Proliferação de Células/efeitos dos fármacos , Artéria Pulmonar/efeitos dos fármacos , Artéria Pulmonar/fisiopatologia , Artéria Pulmonar/metabolismo , Vasodilatadores/farmacologia , Hipertensão Pulmonar/tratamento farmacológico , Hipertensão Pulmonar/fisiopatologia , Hipertensão Pulmonar/patologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Células Cultivadas , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/fisiopatologia , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Masculino , Ratos , Anti-Hipertensivos/farmacologia , Vasodilatação/efeitos dos fármacosRESUMO
Drugs that lower plasma apolipoprotein B (ApoB)-containing lipoproteins are central to treating advanced atherosclerosis and provide partial protection against clinical events. Previous research showed that lowering ApoB-containing lipoproteins stops plaque inflammation, but how these drugs affect the heterogeneous population of plaque cells derived from smooth muscle cells (SMCs) is unknown. SMC-derived cells are the main cellular component of atherosclerotic lesions and the source of structural components that determine the size of plaques and their propensity to rupture and trigger thrombosis, the proximate cause of heart attack and stroke. Using lineage tracing and single-cell techniques to investigate the full SMC-derived cellular compartment in progressing and regressing plaques in mice, here we show that lowering ApoB-containing lipoproteins reduces nuclear factor kappa-light-chain-enhancer of activated B cells signaling in SMC-derived fibromyocytes and chondromyocytes and leads to depletion of these abundant cell types from plaques. These results uncover an important mechanism through which cholesterol-lowering drugs can achieve plaque regression.
Assuntos
Aterosclerose , Modelos Animais de Doenças , Miócitos de Músculo Liso , Placa Aterosclerótica , Animais , Placa Aterosclerótica/patologia , Placa Aterosclerótica/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/patologia , Miócitos de Músculo Liso/metabolismo , Aterosclerose/patologia , Aterosclerose/tratamento farmacológico , Aterosclerose/metabolismo , Condrócitos/efeitos dos fármacos , Condrócitos/patologia , Condrócitos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Anticolesterolemiantes/farmacologia , Anticolesterolemiantes/uso terapêutico , Masculino , Colesterol/metabolismo , Colesterol/sangue , Camundongos , Doenças da Aorta/patologia , Doenças da Aorta/metabolismo , Análise de Célula Única , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/patologia , Músculo Liso Vascular/metabolismo , NF-kappa B/metabolismoRESUMO
Background: Despite increasing evidence suggesting that pulmonary arterial hypertension (PAH) is a complex disease involving vasoconstriction, thrombosis, inflammation, metabolic dysregulation and vascular proliferation, all the drugs approved for PAH mainly act as vasodilating agents. Since excessive TGF-ß signaling is believed to be a critical factor in pulmonary vascular remodeling, we hypothesized that blocking TGFß-activated kinase 1 (TAK-1), alone or in combination with a vasodilator therapy (i.e., riociguat) could achieve a greater therapeutic benefit. Methods: PAH was induced in male Wistar rats by a single injection of the VEGF receptor antagonist SU5416 (20 mg/kg) followed by exposure to hypoxia (10%O2) for 21 days. Two weeks after SU5416 administration, vehicle, riociguat (3 mg/kg/day), the TAK-1 inhibitor 5Z-7-oxozeaenol (OXO, 3 mg/kg/day), or both drugs combined were administered for 7 days. Metabolic profiling of right ventricle (RV), lung tissues and PA smooth muscle cells (PASMCs) extracts were performed by magnetic resonance spectroscopy, and the differences between groups analyzed by multivariate statistical methods. Results: In vitro, riociguat induced potent vasodilator effects in isolated pulmonary arteries (PA) with negligible antiproliferative effects and metabolic changes in PASMCs. In contrast, 5Z-7-oxozeaenol effectively inhibited the proliferation of PASMCs characterized by a broad metabolic reprogramming but had no acute vasodilator effects. In vivo, treatment with riociguat partially reduced the increase in pulmonary arterial pressure (PAP), RV hypertrophy (RVH), and pulmonary vascular remodeling, attenuated the dysregulation of inosine, glucose, creatine and phosphocholine (PC) in RV and fully abolished the increase in lung IL-1ß expression. By contrast, 5Z-7-oxozeaenol significantly reduced pulmonary vascular remodeling and attenuated the metabolic shifts of glucose and PC in RV but had no effects on PAP or RVH. Importantly, combined therapy had an additive effect on pulmonary vascular remodeling and induced a significant metabolic effect over taurine, amino acids, glycolysis, and TCA cycle metabolism via glycine-serine-threonine metabolism. However, it did not improve the effects induced by riociguat alone on pulmonary pressure or RV remodeling. None of the treatments attenuated pulmonary endothelial dysfunction and hyperresponsiveness to serotonin in isolated PA. Conclusion: Our results suggest that inhibition of TAK-1 induces antiproliferative effects and its addition to short-term vasodilator therapy enhances the beneficial effects on pulmonary vascular remodeling and RV metabolic reprogramming in experimental PAH.
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[This corrects the article DOI: 10.3389/fphar.2023.1021535.].
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KV7 channels exert a pivotal role regulating vascular tone in several vascular beds. In this context, KV7 channel agonists represent an attractive strategy for the treatment of pulmonary arterial hypertension (PAH). Therefore, in this study, we have explored the pulmonary vascular effects of the novel KV7 channel agonist URO-K10. Consequently, the vasodilator and electrophysiological effects of URO-K10 were tested in rat and human pulmonary arteries (PA) and PA smooth muscle cells (PASMC) using myography and patch-clamp techniques. Protein expression was also determined by Western blot. Morpholino-induced knockdown of KCNE4 was assessed in isolated PA. PASMC proliferation was measured by BrdU incorporation assay. In summary, our data show that URO-K10 is a more effective relaxant of PA than the classical KV7 activators retigabine and flupirtine. URO-K10 enhanced KV currents in PASMC and its electrophysiological and relaxant effects were inhibited by the KV7 channel blocker XE991. The effects of URO-K10 were confirmed in human PA. URO-K10 also exhibited antiproliferative effects in human PASMC. Unlike retigabine and flupirtine, URO-K10-induced pulmonary vasodilation was not affected by morpholino-induced knockdown of the KCNE4 regulatory subunit. Noteworthy, the pulmonary vasodilator efficacy of this compound was considerably increased under conditions mimicking the ionic remodelling (as an in vitro model of PAH) and in PA from monocrotaline-induced pulmonary hypertensive rats. Taking all together, URO-K10 behaves as a KCNE4-independent KV7 channel activator with much increased pulmonary vascular effects compared to classical KV7 channel activators. Our study identifies a promising new drug in the context of PAH.
Assuntos
Canais de Potássio KCNQ , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Animais , Humanos , Ratos , Canais de Potássio KCNQ/genética , Morfolinos , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Vasodilatadores/farmacologiaRESUMO
Background: Aim of this study was to investigate immune cells and subsets in different stages of human coronary artery disease with a novel multiplex immunohistochemistry (mIHC) technique. Methods: Human left anterior descending coronary artery specimens were analyzed: eccentric intimal thickening (N = 11), pathological intimal thickening (N = 10), fibroatheroma (N = 9), and fibrous plaque (N = 9). Eccentric intimal thickening was considered normal, and pathological intimal thickening, fibroatheroma, and fibrous plaque were considered diseased coronary arteries. Two mIHC panels, consisting of six and five primary antibodies, autofluoresence, and DAPI, were used to detect adaptive and innate immune cells. Via semi-automated analysis, (sub)types of immune cells in whole plaques and specific plaque regions were quantified. Results: Increased numbers of CD3+ T cells (P < 0.001), CD20+ B cells (P = 0.013), CD68+ macrophages (P = 0.003), CD15+ neutrophils (P = 0.017), and CD31+ endothelial cells (P = 0.024) were identified in intimas of diseased coronary arteries compared to normal. Subset analyses of T cells and macrophages showed that diseased coronary arteries contained an abundance of CD3+CD8- non-cytotoxic T cells and CD68+CD206- non-M2-like macrophages. Proportions of CD3+CD45RO+ memory T cells were similar to normal coronary arteries. Among pathological intimal thickening, fibroatheroma, and fibrous plaque, all immune cell numbers and subsets were similar. Conclusions: The type of immune response does not differ substantially between different stages of plaque development and may provide context for mechanistic research into immune cell function in atherosclerosis. We provide the first comprehensive map of immune cell subtypes across plaque types in coronary arteries demonstrating the potential of mIHC for vascular research.
RESUMO
Although type 1 and type 2 diabetes are strongly associated with systemic cardiovascular morbidity, the relationship with pulmonary vascular disease had been almost disregarded until recent epidemiological data revealed that diabetes might be a risk factor for pulmonary hypertension. Recent experimental studies suggest that diabetes induces changes in lung function insufficient to elevate pulmonary pressure. The aim of this study was to assess the effects of diabetes on the sensitivity to other risk factors for pulmonary hypertension. We therefore analysed the effects of the combination of diabetes with exposure to moderate hypoxia on classical markers of pulmonary hypertension. Control (saline-treated) and diabetic (70 mg kg(-1) streptozotocin-treated) male Wistar-Kyoto rats were followed for 4 weeks and exposed to normoxia or moderate normobaric hypoxia (14%) for another 2 weeks. Hypoxia, but not diabetes, strongly reduced voltage-gated potassium currents, whereas diabetes, but not hypoxia, induced pulmonary artery endothelial dysfunction. Both factors independently induced pulmonary vascular remodelling and downregulated the lung bone morphogenetic protein receptor type 2. However, diabetes, but not hypoxia, induced pulmonary infiltration of macrophages, which was markedly increased when both factors were combined. Diabetes plus hypoxia induced a modest increase in diastolic and mean pulmonary artery pressure and right ventricular weight, while each of the two factors alone had no significant effect. The pattern of changes in markers of pulmonary hypertension was different for moderate hypoxia and diabetes, with no synergic effect except for macrophage recruitment, and the combination of both factors was required to induce a moderate elevation in pulmonary arterial pressure.
Assuntos
Diabetes Mellitus Tipo 1/complicações , Hipertensão Pulmonar/etiologia , Hipóxia/complicações , Animais , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/metabolismo , Diabetes Mellitus Experimental/complicações , Regulação para Baixo , Hipertrofia Ventricular Direita/etiologia , Pulmão/patologia , Pulmão/fisiopatologia , Macrófagos/imunologia , Masculino , Artéria Pulmonar/patologia , Artéria Pulmonar/fisiopatologia , Ratos , Ratos Endogâmicos WKY , Doenças Vasculares/etiologiaRESUMO
HIV and Schistosoma infections have been individually associated with pulmonary vascular disease. Co-infection with these pathogens is very common in tropical areas, with an estimate of six million people co-infected worldwide. However, the effects of HIV and Schistosoma co-exposure on the pulmonary vasculature and its impact on the development of pulmonary vascular disease are largely unknown. Here, we have approached these questions by using a non-infectious animal model based on lung embolization of Schistosoma mansoni eggs in HIV-1 transgenic (HIV) mice. Schistosome-exposed HIV mice but not wild-type (Wt) counterparts showed augmented pulmonary arterial pressure associated with markedly suppressed endothelial-dependent vasodilation, increased endothelial remodeling and vessel obliterations, formation of plexiform-like lesions and a higher degree of perivascular fibrosis. In contrast, medial wall muscularization was similarly increased in both types of mice. Moreover, HIV mice displayed an impaired immune response to parasite eggs in the lung, as suggested by decreased pulmonary leukocyte infiltration, small-sized granulomas, and augmented residual egg burden. Notably, vascular changes in co-exposed mice were associated with increased expression of proinflammatory and profibrotic cytokines, including IFN-γ and IL-17A in CD4+ and γδ T cells and IL-13 in myeloid cells. Collectively, our study shows for the first time that combined pulmonary persistence of HIV proteins and Schistosoma eggs, as it may occur in co-infected people, alters the cytokine landscape and targets the vascular endothelium for aggravated pulmonary vascular pathology. Furthermore, it provides an experimental model for the understanding of pulmonary vascular disease associated with HIV and Schistosoma co-morbidity.
Assuntos
Infecções por HIV , Esquistossomose mansoni , Doenças Vasculares , Animais , Citocinas/metabolismo , Infecções por HIV/complicações , Infecções por HIV/patologia , Humanos , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Schistosoma mansoni , Esquistossomose mansoni/complicações , Esquistossomose mansoni/patologia , Doenças Vasculares/patologiaRESUMO
Background: Vitamin D (vitD) deficiency is highly prevalent in patients with pulmonary arterial hypertension (PAH). Moreover, PAH-patients with lower levels of vitD have worse prognosis. We hypothesize that recovering optimal levels of vitD in an animal model of PAH previously depleted of vitD improves the hemodynamics, the endothelial dysfunction and the ionic remodeling. Methods: Male Wistar rats were fed a vitD-free diet for five weeks and then received a single dose of Su5416 (20 mg/Kg) and were exposed to vitD-free diet and chronic hypoxia (10% O2) for three weeks to induce PAH. Following this, vitD deficient rats with PAH were housed in room air and randomly divided into two groups: (a) continued on vitD-free diet or (b) received an oral dose of 100,000 IU/Kg of vitD plus standard diet for three weeks. Hemodynamics, pulmonary vascular remodeling, pulmonary arterial contractility, and K+ currents were analyzed. Results: Recovering optimal levels of vitD improved endothelial function, measured by an increase in the endothelium-dependent vasodilator response to acetylcholine. It also increased the activity of TASK-1 potassium channels. However, vitD supplementation did not reduce pulmonary pressure and did not ameliorate pulmonary vascular remodeling and right ventricle hypertrophy. Conclusions: Altogether, these data suggest that in animals with PAH and severe deficit of vitD, restoring vitD levels to an optimal range partially improves some pathophysiological features of PAH.
Assuntos
Endotélio Vascular/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Proteínas do Tecido Nervoso/metabolismo , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Hipertensão Arterial Pulmonar , Deficiência de Vitamina D , Vitamina D , Animais , Endotélio Vascular/patologia , Masculino , Hipertensão Arterial Pulmonar/tratamento farmacológico , Hipertensão Arterial Pulmonar/metabolismo , Hipertensão Arterial Pulmonar/patologia , Ratos , Ratos Wistar , Vitamina D/farmacocinética , Vitamina D/farmacologia , Deficiência de Vitamina D/tratamento farmacológico , Deficiência de Vitamina D/metabolismo , Deficiência de Vitamina D/patologiaRESUMO
Current approved therapies for pulmonary hypertension (PH) aim to restore the balance between endothelial mediators in the pulmonary circulation. These drugs may exert vasodilator effects on poorly oxygenated vessels. This may lead to the derivation of blood perfusion towards low ventilated alveoli, i.e., producing ventilation-perfusion mismatch, with detrimental effects on gas exchange. The aim of this study is to analyze the oxygen-sensitivity in vitro of 25 drugs currently used or potentially useful for PH. Additionally, the study analyses the effectiveness of these vasodilators in the pulmonary vs the systemic vessels. Vasodilator responses were recorded in pulmonary arteries (PA) and mesenteric arteries (MA) from rats and in human PA in a wire myograph under different oxygen concentrations. None of the studied drugs showed oxygen selectivity, being equally or more effective as vasodilators under conditions of low oxygen as compared to high oxygen levels. The drugs studied showed low pulmonary selectivity, being equally or more effective as vasodilators in systemic than in PA. A similar behavior was observed for the members within each drug family. In conclusion, none of the drugs showed optimal vasodilator profile, which may limit their therapeutic efficacy in PH.