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1.
Molecules ; 26(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203711

RESUMO

Inflammation and stiffness in the arteries is referred to as vascular calcification. This process is a prevalent yet poorly understood consequence of cardiovascular disease and diabetes mellitus, comorbidities with few treatments clinically available. Because this is an active process similar to bone formation, it is hypothesized that osteoclasts (OCs), bone-resorbing cells in the body, could potentially work to reverse existing calcification by resorbing bone material. The receptor activator of nuclear kappa B-ligand (RANKL) is a molecule responsible for triggering a response in monocytes and macrophages that allows them to differentiate into functional OCs. In this study, OC and RANKL delivery were employed to determine whether calcification could be attenuated. OCs were either delivered via direct injection, collagen/alginate microbeads, or collagen gel application, while RANKL was delivered via injection, through either a porcine subdermal model or aortic injury model. While in vitro results yielded a decrease in calcification using OC therapy, in vivo delivery mechanisms did not provide control or regulation to keep cells localized long enough to induce calcification reduction. However, these results do provide context and direction for the future of OC therapy, revealing necessary steps for this treatment to effectively reduce calcification in vivo. The discrepancy between in vivo and in vitro success for OC therapy points to the need for a more stable and time-controlled delivery mechanism that will allow OCs not only to remain at the site of calcification, but also to be regulated so that they are healthy and functioning normally when introduced to diseased tissue.


Assuntos
Terapia Baseada em Transplante de Células e Tecidos/métodos , Osteoclastos/fisiologia , Calcificação Vascular/terapia , Animais , Reabsorção Óssea/metabolismo , Proteínas de Transporte/farmacologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Elastina/metabolismo , Elastina/fisiologia , Macrófagos/metabolismo , Masculino , Glicoproteínas de Membrana , Monócitos/metabolismo , Miócitos de Músculo Liso/metabolismo , Osteoclastos/metabolismo , Osteogênese/efeitos dos fármacos , Ligante RANK/metabolismo , Ligante RANK/farmacologia , Ratos , Ratos Sprague-Dawley , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Suínos , Calcificação Vascular/metabolismo
2.
Molecules ; 26(12)2021 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-34200914

RESUMO

Flavonoids are a group of secondary metabolites derived from plant-based foods, and they offer many health benefits in different stages of several diseases. This review will focus on their effects on ion channels expressed in vascular smooth muscle during atherosclerosis. Since ion channels can be regulated by redox potential, it is expected that during the onset of oxidative stress-related diseases, ion channels present changes in their conductive activity, impacting the progression of the disease. A typical oxidative stress-related condition is atherosclerosis, which involves the dysfunction of vascular smooth muscle. We aim to present the state of the art on how redox potential affects vascular smooth muscle ion channel function and summarize if the benefits observed in this disease by using flavonoids involve restoring the ion channel activity.


Assuntos
Aterosclerose/tratamento farmacológico , Flavonoides/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Animais , Aterosclerose/metabolismo , Progressão da Doença , Humanos , Canais Iônicos/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Oxirredução/efeitos dos fármacos
3.
Zhonghua Jie He He Hu Xi Za Zhi ; 44(6): 537-542, 2021 Jun 12.
Artigo em Chinês | MEDLINE | ID: mdl-34102715

RESUMO

Objective: To investigate the effect of VDR gene silencing on proliferation of airway smooth muscle cells (ASMCs) and elucidate the role of NF-κB. Methods: A recombinant lentiviral vector specifically targeting VDR gene in rat was constructed by RNA interference. Rat ASMCs were divided into blank group, empty vector group and interference group. ASM cell line model stably silencing the VDR gene RNA expressing was selected by puromycin. Then MTT colorimetric assay and cell cycle analysis by flow cytometry were used to examine cell proliferation. The activation of nuclear factor-κB was determined by immunofluorescence double label method. Moreover, NF-κB-dependent transcription activity was tested through luciferase reporter gene assay. Western blotting was used for IκBα and phospho-IκBα protein levels and actinomycin D treatment was used to determine IκBα mRNA stability. All statistical analyses were performed using SPSS version 23.0 software. Differences between groups were analyzed using one-way ANOVA analysis. Multiple comparisons among groups were made by Student-Newman-Keuls test. Results: (1) As compared with those in the blank group and the empty vector group, the cell proliferation index (PI) and the percent of ASMCs at G2/M phase in the interference group were markedly increased (P<0.05), but their percent at G0/1 phase was decreased (P<0.05).(2) In the interference group, the nuclear translocation of NF-κB p65 in ASMCs was obviously induced. And its level of receptor gene NF-κB p65 (1.37±0.28) was significantly higher than that in the blank group (1.00±0.19,P=0.031) and in the empty vector group (0.96±0.18,P=0.027).(3) In the interference group, the IκBα protein level in ASMCs (0.13±0.04) was obviously less than that in the blank group (0.29±0.05, P=0.023) and in the empty vector group (0.32±0.07, P=0.014). Oppositely, the p-IκBα/IκBα level in the interference group (0.86±0.04) was much more than that in the blank control group (0.41±0.07, P=0.026) and in the empty vector group (0.37±0.05, P=0.017). (4) In the interference group, IκBα mRNA showed a shorter half-life, (171.31±9.67) min, compared to that in the blank group [(224.69±7.95) min,P=0.032] and in the empty vector group [(230.41±6.37) min,P=0.035]. Conclusion: VDR gene silencing could promote ASMC proliferation and the underlying mechanism may involve the activation of NF-κB signaling pathway.


Assuntos
NF-kappa B , Transdução de Sinais , Animais , Proliferação de Células , Miócitos de Músculo Liso/metabolismo , Inibidor de NF-kappaB alfa/genética , NF-kappa B/genética , NF-kappa B/metabolismo , Interferência de RNA , Ratos , Receptores de Calcitriol
4.
Aging (Albany NY) ; 13(12): 16485-16499, 2021 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-34153003

RESUMO

This study aimed to construct an atlas of the cell landscape and comprehensively characterize the cellular repertoire of the pulmonary endarterectomized tissues of patients with chronic thromboembolic pulmonary hypertension (CTEPH). Five pulmonary endarterectomized tissues were collected. 10× Genomics single-cell RNA sequencing was performed, followed by the identification of cluster marker genes and cell types. Gene Ontology (GO) enrichment analysis was conducted. Seventeen cell clusters were characterized, corresponding to 10,518 marker genes, and then classified into eight cell types, including fibroblast/smooth muscle cell, endothelial cell, T cell/NK cell, macrophage, mast cell, cysteine rich secretory protein LCCL domain containing 2 (CRISPLD2)+ cell, cancer stem cell, and undefined. The specific marker genes of fibroblast/smooth muscle cell, endothelial cell, T cell/NK cell, macrophage, mast cell, and cancer stem cell were significantly enriched for multiple functions associated with muscle cell migration, endothelial cell migration, T cell activation, neutrophil activation, erythrocyte homeostasis, and tissue remodeling, respectively. No functions were significantly enriched for the marker gene of CRISPLD2+ cell. Our study, for the first time, provides an atlas of the cell landscape of the pulmonary endarterectomized tissues of CTEPH patients at single-cell resolution, which may serve as a valuable resource for further elucidation of disease pathophysiology.


Assuntos
Endarterectomia , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/cirurgia , Análise de Sequência de RNA , Análise de Célula Única , Tromboembolia/genética , Agregação Celular , Doença Crônica , Feminino , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Humanos , Hipertensão Pulmonar/imunologia , Células Matadoras Naturais/imunologia , Pulmão/patologia , Macrófagos/imunologia , Masculino , Mastócitos/imunologia , Pessoa de Meia-Idade , Miócitos de Músculo Liso/metabolismo , Linfócitos T/imunologia
5.
Int J Mol Sci ; 22(10)2021 May 12.
Artigo em Inglês | MEDLINE | ID: mdl-34066088

RESUMO

Familial hypercholesterolemia (FH) is increasingly associated with inflammation, a phenotype that persists despite treatment with lipid lowering therapies. The alternative C3 complement system (C3), as a key inflammatory mediator, seems to be involved in the atherosclerotic process; however, the relationship between C3 and lipids during plaque progression remains unknown. The aim of the study was to investigate by a systems biology approach the role of C3 in relation to lipoprotein levels during atherosclerosis (AT) progression and to gain a better understanding on the effects of C3 products on the phenotype and function of human lipid-loaded vascular smooth muscle cells (VSMCs). By mass spectrometry and differential proteomics, we found the extracellular matrix (ECM) of human aortas to be enriched in active components of the C3 complement system, with a significantly different proteomic signature in AT segments. Thus, C3 products were more abundant in AT-ECM than in macroscopically normal segments. Furthermore, circulating C3 levels were significantly elevated in FH patients with subclinical coronary AT, evidenced by computed tomographic angiography. However, no correlation was identified between circulating C3 levels and the increase in plaque burden, indicating a local regulation of the C3 in AT arteries. In cell culture studies of human VSMCs, we evidenced the expression of C3, C3aR (anaphylatoxin receptor) and the integrin αMß2 receptor for C3b/iC3b (RT-PCR and Western blot). C3mRNA was up-regulated in lipid-loaded human VSMCs, and C3 protein significantly increased in cell culture supernatants, indicating that the C3 products in the AT-ECM have a local vessel-wall niche. Interestingly, C3a and iC3b (C3 active fragments) have functional effects on VSMCs, significantly reversing the inhibition of VSMC migration induced by aggregated LDL and stimulating cell spreading, organization of F-actin stress fibers and attachment during the adhesion of lipid-loaded human VSMCs. This study, by using a systems biology approach, identified molecular processes involving the C3 complement system in vascular remodeling and in the progression of advanced human atherosclerotic lesions.


Assuntos
Aterosclerose/patologia , Complemento C3/metabolismo , Hiperlipoproteinemia Tipo II/patologia , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/patologia , Proteoma/metabolismo , Adulto , Aterosclerose/imunologia , Aterosclerose/metabolismo , Estudos de Casos e Controles , Adesão Celular , Células Cultivadas , Feminino , Humanos , Hiperlipoproteinemia Tipo II/imunologia , Hiperlipoproteinemia Tipo II/metabolismo , Masculino , Pessoa de Meia-Idade , Músculo Liso Vascular/imunologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/imunologia , Miócitos de Músculo Liso/metabolismo , Proteoma/análise , Remodelação Vascular , Cicatrização , Adulto Jovem
6.
Int J Mol Sci ; 22(11)2021 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-34064112

RESUMO

Increasing adipose tissue mass in obesity directly correlates with elevated circulating leptin levels. Leptin is an adipokine known to play a role in numerous biological processes including regulation of energy homeostasis, inflammation, vascular function and angiogenesis. While physiological concentrations of leptin may exhibit multiple beneficial effects, chronically elevated pathophysiological levels or hyperleptinemia, characteristic of obesity and diabetes, is a major risk factor for development of atherosclerosis. Hyperleptinemia results in a state of selective leptin resistance such that while beneficial metabolic effects of leptin are dampened, deleterious vascular effects of leptin are conserved attributing to vascular dysfunction. Leptin exerts potent proatherogenic effects on multiple vascular cell types including macrophages, endothelial cells and smooth muscle cells; these effects are mediated via an interaction of leptin with the long form of leptin receptor, abundantly expressed in atherosclerotic plaques. This review provides a summary of recent in vivo and in vitro studies that highlight a role of leptin in the pathogenesis of atherosclerotic complications associated with obesity and diabetes.


Assuntos
Aterosclerose/metabolismo , Células Endoteliais/metabolismo , Leptina/metabolismo , Macrófagos/metabolismo , Miócitos de Músculo Liso/metabolismo , Animais , Diabetes Mellitus/metabolismo , Humanos , Obesidade/metabolismo
7.
Int J Mol Sci ; 22(11)2021 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-34063769

RESUMO

Maturation of the cardiovascular system is associated with crucial structural and functional remodeling. Thickening of the arterial wall, maturation of the sympathetic innervation, and switching of the mechanisms of arterial contraction from calcium-independent to calcium-dependent occur during postnatal development. All these processes promote an almost doubling of blood pressure from the moment of birth to reaching adulthood. This review focuses on the developmental alterations of potassium channels functioning as key smooth muscle membrane potential determinants and, consequently, vascular tone regulators. We present evidence that the pattern of potassium channel contribution to vascular control changes from Kir2, Kv1, Kv7 and TASK-1 channels to BKCa channels with maturation. The differences in the contribution of potassium channels to vasomotor tone at different stages of postnatal life should be considered in treatment strategies of cardiovascular diseases associated with potassium channel malfunction.


Assuntos
Artérias/metabolismo , Miócitos de Músculo Liso/metabolismo , Canais de Potássio/metabolismo , Animais , Pressão Sanguínea/fisiologia , Cálcio/metabolismo , Potenciais da Membrana/fisiologia , Músculo Liso Vascular/metabolismo , Cuidado Pós-Natal/métodos
8.
Int J Mol Sci ; 22(8)2021 Apr 16.
Artigo em Inglês | MEDLINE | ID: mdl-33923431

RESUMO

Human atherosclerotic plaque contains smooth muscle cells (SMCs) negative for the contractile phenotype (α-smooth muscle actin) but positive for proprotein convertase subtilisin/kexin type 9 (PCSK9). Thus, we generated rat SMCs which overexpressed human PCSK9 (SMCsPCSK9) with the aim of investigating the role of PCSK9 in the phenotype of SMCs. PCSK9 overexpression in SMCsPCSK9 led to a significant downregulation of the low-density lipoprotein receptor (Ldlr) as well as transgelin (Sm22α), a marker of the contractile phenotype. The cell proliferation rate of SMCsPCSK9 was significantly faster than that of the control SMCs (SMCspuro). Interestingly, overexpression of PCSK9 did not impact the migratory capacity of SMCs in response to 10% FCS, as determined by Boyden's chamber assay. Expression and activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase (Hmgcr) was significantly increased in the presence of PCSK9, both in SMCPCSK9 and after treatment with recombinant PCSK9. The transcriptional activity of sterol regulatory element-binding protein (SREBP) was also increased in the presence of PSCK9, suggesting a direct role of PCSK9 in the control of SRE-responsive genes, like HMGCR. We also observed that cholesterol biosynthesis is elevated in SMCPCSK9, potentially explaining the increased proliferation observed in these cells. Finally, concentration-dependent experiments with simvastatin demonstrated that SMCsPCSK9 were partially resistant to the antiproliferative and antimigratory effect of this drug. Taken together, these data further support a direct role of PCSK9 in proliferation, migration, and phenotypic changes in SMCs-pivotal features of atherosclerotic plaque development. We also provide new evidence on the role of PCSK9 in the pharmacological response to statins-gold standard lipid-lowering drugs with pleiotropic action.


Assuntos
Anticolesterolemiantes/farmacologia , Proliferação de Células , Miócitos de Músculo Liso/metabolismo , Pró-Proteína Convertase 9/metabolismo , Sinvastatina/farmacologia , Animais , Movimento Celular , Células Cultivadas , Colesterol/metabolismo , Feminino , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/fisiologia , Pró-Proteína Convertase 9/genética , Ratos , Receptores de LDL
9.
Int J Mol Sci ; 22(7)2021 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-33808324

RESUMO

Calcium (Ca2+) is an important mediator of multicellular homeostasis and is involved in several diseases. The interplay among the kidney, bone, intestine, and parathyroid gland in Ca2+ homeostasis is strictly modulated by numerous hormones and signaling pathways. The calcium-sensing receptor (CaSR) is a G protein-coupled receptor, that is expressed in calcitropic tissues such as the parathyroid gland and the kidney, plays a pivotal role in Ca2+ regulation. CaSR is important for renal Ca2+, as a mutation in this receptor leads to hypercalciuria and calcium nephrolithiasis. In addition, CaSR is also widely expressed in the vascular system, including vascular endothelial cells (VECs) and vascular smooth muscle cells (VSMCs) and participates in the process of vascular calcification. Aberrant Ca2+ sensing by the kidney and VSMCs, owing to altered CaSR expression or function, is associated with the formation of nephrolithiasis and vascular calcification. Based on emerging epidemiological evidence, patients with nephrolithiasis have a higher risk of vascular calcification, but the exact mechanism linking the two conditions is unclear. However, a dysregulation in Ca2+ homeostasis and dysfunction in CaSR might be the connection between the two. This review summarizes renal calcium handling and calcium signaling in the vascular system, with a special focus on the link between nephrolithiasis and vascular calcification.


Assuntos
Sinalização do Cálcio/fisiologia , Nefrolitíase/metabolismo , Calcificação Vascular/metabolismo , Animais , Cálcio/metabolismo , Células Endoteliais/metabolismo , Humanos , Hipercalciúria/genética , Hipercalciúria/metabolismo , Hipercalciúria/fisiopatologia , Rim/metabolismo , Cálculos Renais/metabolismo , Miócitos de Músculo Liso/metabolismo , Nefrolitíase/fisiopatologia , Receptores de Detecção de Cálcio/genética , Calcificação Vascular/genética , Calcificação Vascular/fisiopatologia
10.
Life Sci ; 277: 119492, 2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-33864819

RESUMO

AIMS: Non-invasive and simultaneous recording of gastrointestinal (GI) activity during stress induction is still an unexplored field. In our previous investigation, the stress-induced alteration of the gastrointestinal tract was explored in rats. Our aims were to expand our previous rat experiment and to induce stress response in rats (Study 1) and humans (Study 2) to detect the GI tract activity, heart rate and body temperature. MATERIALS AND METHODS: In the preclinical sample, acute stress was induced by immobilization in Sprague-Dawley rats (N = 10). Acute stress response was generated by the Trier Social Stress Test among healthy volunteers (N = 16). Detection of acute stress was measured by using smooth muscle electromyography, which recorded the myoelectric waves of the gastrointestinal tract (stomach, ileum and colon) simultaneously with heart rate and body temperature in rats and humans. KEY FINDINGS: The myoelectric waves of the stomach, the cecum and the ileum increased during immobilization in rats, rising in parallel with heart rate and the dermal temperature of the abdominal surface. The same alterations were found during the stress period among humans, except in the case of the colon, where no change was detected. SIGNIFICANCE: The crucial role of the GI tract in stress response was revealed by translating the outcome of basic research into human results. The similar GI alterations during stress in rats and humans underpin the robustness of our findings. In summary, our preliminary translational-based study can serve as an appropriate basis for further human studies.


Assuntos
Trato Gastrointestinal/fisiologia , Miócitos de Músculo Liso/metabolismo , Estresse Fisiológico/fisiologia , Adulto , Animais , Ceco/fisiologia , Colo/fisiologia , Eletromiografia/métodos , Feminino , Motilidade Gastrointestinal/fisiologia , Voluntários Saudáveis , Humanos , Íleo/fisiologia , Masculino , Músculo Liso/fisiologia , Ratos , Ratos Sprague-Dawley , Estômago/fisiologia
11.
Biomed Res Int ; 2021: 5551504, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33928148

RESUMO

Background: Previous studies have demonstrated the ubiquitin-proteasome inhibitor bortezomib (BTZ) can effectively alleviate hypoxia-induced pulmonary hypertension (HPH) by suppressing the intracellular calcium homeostasis in pulmonary arterial smooth muscle cells (PASMCs). Further evaluation showed that the antiproliferation roles of BTZ are mainly mediated by inhibition of the intracellular calcium homeostasis. Caveolin-1 belongs to one of the key regulators of the intracellular calcium homeostasis in PASMCs, which can regulate the store-operated calcium entry (SOCE). However, the effects of BTZ on Caveolin-1 remain unclear. Methods: Primarily cultured human PASMCs were used as the cell model. CCK-8 assay was performed to assess the PASMCs proliferation. Western blotting and real-time qPCR were used to detect the mRNA and protein expressions. Fura-2-based fluorescence imaging experiments were used to determine the intracellular calcium concentration ([Ca2+]i). The protein synthesis inhibitor cycloheximide (CHX) was utilized to determine the protein degradation process. Results: Firstly, in cultured human PASMCs, treatment of BTZ for 24 or 60 hours significantly downregulates Caveolin-1 at both mRNA and protein levels. Secondly, in the presence CHX, BTZ treatment also leads to downregulated protein expression and fastened protein degradation of Caveolin-1, indicating that BTZ can promote the Caveolin-1 protein degradation, other than the BTZ on Caveolin-1 mRNA transcription. Then, BTZ significantly attenuates the hypoxia-elevated baseline [Ca2+]i, SOCE, and cell proliferation. Conclusion: We firstly observed that the ubiquitin-proteasome inhibitor BTZ can inhibit the Caveolin-1 expression at both mRNA transcription and protein degradation processes, providing new mechanistic basis of BTZ on PASMC proliferation.


Assuntos
Bortezomib/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Cálcio/metabolismo , Caveolina 1/metabolismo , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Artéria Pulmonar/patologia , Caveolina 1/genética , Hipóxia Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Cloroquina/farmacologia , Homeostase/efeitos dos fármacos , Humanos , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Proteólise/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
12.
Nutr Metab Cardiovasc Dis ; 31(5): 1635-1644, 2021 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-33812737

RESUMO

BACKGROUND AND AIM: Our previous study found carotid baroreceptor stimulation (CBS) reduces body weight and white adipose tissue (WAT) weight, restores abnormal secretion of adipocytokines and inflammation factors, decreases systolic blood pressure (SBP) by inhibiting activation of sympathetic nervous system (SNS) and renin-angiotensin system (RAS) in obese rats. In this study, we explore effects of CBS on aortic remodeling in obese rats. METHODS AND RESULTS: Rats were fed high-fat diet (HFD) for 16 weeks to induce obesity and underwent either CBS device implantation and stimulation or sham operation at 8 weeks. BP and body weight were measured weekly. RAS activity of WAT, histological, biochemical and functional profiles of aortas were detected after 16 weeks. CBS effectively decreased BP in obese rats, downregulated mRNA expression of angiotensinogen (AGT) and renin in WAT, concentrations of AGT, renin, angiotensin II (Ang II), protein levels of Ang II receptor 1 (AT1R) and Ang II receptor 2 (AT2R) in WAT were declined. CBS inhibited reactive oxygen species (ROS) generation, inflammatory response and endoplasmic reticulum (ER) stress in aortas of obese rats, restrained vascular wall thickening and vascular smooth muscle cells (VSMCs) phenotypic switching, increased nitric oxide (NO) synthesis, promoted endothelium-dependent vasodilatation by decreasing protein expression of AT1R and leptin receptor (LepR), increasing protein expression of adiponectin receptor 1 (AdipoR1) in aortic VSMCs. CONCLUSION: CBS reduced BP and reversed aortic remodeling in obese rats, the underlying mechanism might be related to the suppressed SNS activity, restored adipocytokine secretion and restrained RAS activity of WAT.


Assuntos
Tecido Adiposo Branco/metabolismo , Terapia por Estimulação Elétrica , Músculo Liso Vascular/patologia , Obesidade/terapia , Pressorreceptores/fisiopatologia , Sistema Renina-Angiotensina , Remodelação Vascular , Adipocinas/metabolismo , Animais , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Pressão Arterial , Modelos Animais de Doenças , Terapia por Estimulação Elétrica/instrumentação , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Neuroestimuladores Implantáveis , Masculino , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiopatologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Obesidade/metabolismo , Obesidade/patologia , Obesidade/fisiopatologia , Ratos Sprague-Dawley , Receptor Tipo 1 de Angiotensina/metabolismo , Receptores de Adiponectina , Receptores para Leptina/metabolismo , Vasodilatação
13.
Int J Mol Sci ; 22(6)2021 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-33805714

RESUMO

Trifluoperazine (TFP), an antipsychotic drug approved by the Food and Drug Administration, has been show to exhibit anti-cancer effects. Pulmonary arterial hypertension (PAH) is a devastating disease characterized by a progressive obliteration of small pulmonary arteries (PAs) due to exaggerated proliferation and resistance to apoptosis of PA smooth muscle cells (PASMCs). However, the therapeutic potential of TFP for correcting the cancer-like phenotype of PAH-PASMCs and improving PAH in animal models remains unknown. PASMCs isolated from PAH patients were exposed to different concentrations of TFP before assessments of cell proliferation and apoptosis. The in vivo therapeutic potential of TFP was tested in two preclinical models with established PAH, namely the monocrotaline and sugen/hypoxia-induced rat models. Assessments of hemodynamics by right heart catheterization and histopathology were conducted. TFP showed strong anti-survival and anti-proliferative effects on cultured PAH-PASMCs. Exposure to TFP was associated with downregulation of AKT activity and nuclear translocation of forkhead box protein O3 (FOXO3). In both preclinical models, TFP significantly lowered the right ventricular systolic pressure and total pulmonary resistance and improved cardiac function. Consistently, TFP reduced the medial wall thickness of distal PAs. Overall, our data indicate that TFP could have beneficial effects in PAH and support the view that seeking new uses for old drugs may represent a fruitful approach.


Assuntos
Fármacos Cardiovasculares/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hipertensão Pulmonar/tratamento farmacológico , Hipóxia/prevenção & controle , Miócitos de Músculo Liso/efeitos dos fármacos , Trifluoperazina/farmacologia , Animais , Antipsicóticos/farmacologia , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Reposicionamento de Medicamentos , Feminino , Proteína Forkhead Box O3/genética , Proteína Forkhead Box O3/metabolismo , Hemodinâmica/efeitos dos fármacos , Humanos , Hipertensão Pulmonar/induzido quimicamente , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/fisiopatologia , Hipóxia/induzido quimicamente , Hipóxia/genética , Hipóxia/fisiopatologia , Indóis/administração & dosagem , Monocrotalina/administração & dosagem , Músculo Liso Vascular/citologia , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Artéria Pulmonar/citologia , Artéria Pulmonar/efeitos dos fármacos , Artéria Pulmonar/metabolismo , Pirróis/administração & dosagem , Ratos , Ratos Sprague-Dawley , Survivina/genética , Survivina/metabolismo
14.
Sichuan Da Xue Xue Bao Yi Xue Ban ; 52(2): 188-193, 2021 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-33829690

RESUMO

Objective: To analyze the correlation between connexin 43 (Cx43) and the expression of P16 and P21, aging-related proteins, and to investigate the possible role of Cx43 in the development of cell senescence with an aging model prepared by D-galactose (D-gal) intervention in the vascular smooth muscle cells (VSMCs) of guinea pig spiral modiolar artery (SMA). Methods: The VSMCs of guinea pig SMA were cultured with the adhesion method, and the markers of VSMCs were detected with immunofluorescence technique. The experiment has a control group, a D-gal group, and a group that received D-gal and gap junction agonist AAP10 intervention, hereafter referred to as the AAP10 group. Cell Counting Kit-8 (CCK-8) was used to check VSMC activity and to determine the concentration and duration of D-gal intervention. The mRNA expression of Cx43 in each group was checked with qRT-PCR. The expression of Cx43, P16 and P21 proteins in each group was examined with the Western blot. The expression and distribution of P16 and P21 proteins were examined with immunofluorescence assay. Results: Immunofluorescence results showed that the positive expression rate of cell actin (α-SM-actin) was over 90%. CCK-8 results showed that the optimal concentration of D-gal intervention was 30 mg/mL and the intervention duration was 48 h. qRT-PCR test showed that the mRNA expression of Cx43 in VSMCs in the D-gal group was significantly lower than that in the control group ( P<0.01), while it is higher in the AAP10 group than that of the D-gal group ( P<0.01); Western blot assay showed that the Cx43 expression level in VSMCs in the D-gal group was significantly lower than that in the control group ( P<0.01) and the expression of P16 and P21 was significantly higher than that in the control group ( P<0.01), the expression of Cx43 protein in AAP10 group was significantly up-regulated compared with that in the D-gal group ( P<0.01), while the expression of P16 and P21 was down-regulated significantly ( P<0.01); The results of immunofluorescence showed that P16 and P21 were mainly expressed in the cell nucleus. Semi-quantitative analysis of fluorescence intensity showed that the level of P16 and P21 protein in the D-gal group was significantly higher than that in the control group, and the fluorescence intensity of AAP10 group was significantly lower than that in the D-gal group ( P<0.01). Conclusion: Up-regulation of Cx43 expression can reverse the D-gal-induced abnormal expression of P16 and P21, two aging-related proteins, in SMA. It is suggested that Cx43 on SMA may be involved in D-gal-induced cell senescence, which provides a theoretical basis and possible intervention target for the delay of cell senescence.


Assuntos
Conexina 43 , Músculo Liso Vascular , Animais , Artérias , Senescência Celular , Conexina 43/genética , Conexina 43/metabolismo , Regulação para Baixo , Cobaias , Miócitos de Músculo Liso/metabolismo
15.
Int J Mol Sci ; 22(6)2021 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-33799449

RESUMO

(1) Background: Tissue non-specific alkaline phosphatase (TNAP) is suspected to induce atherosclerosis plaque calcification. TNAP, during physiological mineralization, hydrolyzes the mineralization inhibitor inorganic pyrophosphate (PPi). Since atherosclerosis plaques are characterized by the presence of necrotic cells that probably release supraphysiological concentrations of ATP, we explored whether this extracellular adenosine triphosphate (ATP) is hydrolyzed into the mineralization inhibitor PPi or the mineralization stimulator inorganic phosphate (Pi), and whether TNAP is involved. (2) Methods: Murine aortic smooth muscle cell line (MOVAS cells) were transdifferentiated into chondrocyte-like cells in calcifying medium, containing ascorbic acid and ß-glycerophosphate. ATP hydrolysis rates were determined in extracellular medium extracted from MOVAS cultures during their transdifferentiation, using 31P-NMR and IR spectroscopy. (3) Results: ATP and PPi hydrolysis by MOVAS cells increased during transdifferentiation. ATP hydrolysis was sequential, yielding adenosine diphosphate (ADP), adenosine monophosphate (AMP), and adenosine without any detectable PPi. The addition of levamisole partially inhibited ATP hydrolysis, indicating that TNAP and other types of ectonucleoside triphoshatediphosphohydrolases contributed to ATP hydrolysis. (4) Conclusions: Our findings suggest that high ATP levels released by cells in proximity to vascular smooth muscle cells (VSMCs) in atherosclerosis plaques generate Pi and not PPi, which may exacerbate plaque calcification.


Assuntos
Aterosclerose/genética , Transdiferenciação Celular/genética , Difosfatos/metabolismo , Calcificação Vascular/genética , Trifosfato de Adenosina , Fosfatase Alcalina/genética , Animais , Aorta/citologia , Aorta/metabolismo , Ácido Ascórbico/farmacologia , Aterosclerose/metabolismo , Aterosclerose/patologia , Condrócitos/metabolismo , Condrócitos/patologia , Glicerofosfatos/genética , Glicerofosfatos/metabolismo , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Fosfatos/metabolismo , Calcificação Vascular/metabolismo , Calcificação Vascular/patologia
16.
Int J Mol Sci ; 22(6)2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33803922

RESUMO

Pulmonary arterial hypertension (PAH) is a progressive disorder characterized by a sustained elevation of pulmonary artery (PA) pressure, right ventricular failure, and premature death. Enhanced proliferation and resistance to apoptosis (as seen in cancer cells) of PA smooth muscle cells (PASMCs) is a major pathological hallmark contributing to pulmonary vascular remodeling in PAH, for which current therapies have only limited effects. Emerging evidence points toward a critical role for Enhancer of Zeste Homolog 2 (EZH2) in cancer cell proliferation and survival. However, its role in PAH remains largely unknown. The aim of this study was to determine whether EZH2 represents a new factor critically involved in the abnormal phenotype of PAH-PASMCs. We found that EZH2 is overexpressed in human lung tissues and isolated PASMCs from PAH patients compared to controls as well as in two animal models mimicking the disease. Through loss- and gain-of-function approaches, we showed that EZH2 promotes PAH-PASMC proliferation and survival. By combining quantitative transcriptomic and proteomic approaches in PAH-PASMCs subjected or not to EZH2 knockdown, we found that inhibition of EZH2 downregulates many factors involved in cell-cycle progression, including E2F targets, and contributes to maintain energy production. Notably, we found that EZH2 promotes expression of several nuclear-encoded components of the mitochondrial translation machinery and tricarboxylic acid cycle genes. Overall, this study provides evidence that, by overexpressing EZH2, PAH-PASMCs remove the physiological breaks that normally restrain their proliferation and susceptibility to apoptosis and suggests that EZH2 or downstream factors may serve as therapeutic targets to combat pulmonary vascular remodeling.


Assuntos
Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteoma/genética , Hipertensão Arterial Pulmonar/genética , Transcriptoma/genética , Animais , Apoptose/genética , Proliferação de Células/genética , Ciclo do Ácido Cítrico/genética , Epigênese Genética/genética , Feminino , Ventrículos do Coração/metabolismo , Ventrículos do Coração/patologia , Humanos , Pulmão/metabolismo , Pulmão/patologia , Masculino , Pessoa de Meia-Idade , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Hipertensão Arterial Pulmonar/patologia , Artéria Pulmonar/crescimento & desenvolvimento , Artéria Pulmonar/patologia , Ratos
17.
Int J Mol Sci ; 22(5)2021 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-33806419

RESUMO

Arterial smooth muscle exhibits rhythmic oscillatory contractions called vasomotion and believed to be a protective mechanism against tissue hypoperfusion or hypoxia. Oscillations of vascular tone depend on voltage and follow oscillations of the membrane potential. Voltage-gated sodium channels (Nav), responsible for the initiation and propagation of action potentials in excitable cells, have also been evidenced both in animal and human vascular smooth muscle cells (SMCs). For example, they contribute to arterial contraction in rats, but their physiopathological relevance has not been established in human vessels. In the present study, we investigated the functional role of Nav in the human artery. Experiments were performed on human uterine arteries obtained after hysterectomy and on SMCs dissociated from these arteries. In SMCs, we recorded a tetrodotoxin (TTX)-sensitive and fast inactivating voltage-dependent INa current. Various Nav genes, encoding α-subunit isoforms sensitive (Nav 1.2; 1.3; 1.7) and resistant (Nav 1.5) to TTX, were detected both in arterial tissue and in SMCs. Nav channels immunostaining showed uniform distribution in SMCs and endothelial cells. On arterial tissue, we recorded variations of isometric tension, ex vivo, in response to various agonists and antagonists. In arterial rings placed under hypoxic conditions, the depolarizing agent KCl and veratridine, a specific Nav channels agonist, both induced a sustained contraction overlaid with rhythmic oscillations of tension. After suppression of sympathetic control either by blocking the release of catecholamine or by antagonizing the target adrenergic response, rhythmic activity persisted while the sustained contraction was abolished. This rhythmic activity of the arteries was suppressed by TTX but, in contrast, only attenuated by antagonists of calcium channels, Na+/Ca2+ exchanger, Na+/K+-ATPase and the cardiac Nav channel. These results highlight the role of Nav as a novel key element in the vasomotion of human arteries. Hypoxia promotes activation of Nav channels involved in the initiation of rhythmic oscillatory contractile activity.


Assuntos
Artérias/metabolismo , Hipóxia/metabolismo , Contração Muscular/fisiologia , Canais de Sódio Disparados por Voltagem/metabolismo , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Adulto , Animais , Artérias/efeitos dos fármacos , Canais de Cálcio/metabolismo , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Feminino , Humanos , Pessoa de Meia-Idade , Contração Muscular/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Trocador de Sódio e Cálcio/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Tetrodotoxina/farmacologia
18.
Oxid Med Cell Longev ; 2021: 6668971, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33824697

RESUMO

Background: Airway smooth muscle (ASM) remodeling is a hallmark in chronic obstructive pulmonary disease (COPD). NADPH oxidase 4- (NOX4-) mediated reactive oxygen species (ROS) production plays a crucial role in cell differentiation and extracellular matrix (ECM) synthesis in ASM remodeling. However, the precise mechanisms underpinning its pathogenic roles remain elusive. Methods: The expression of NOX4 and TGF-ß 1 in the airway of the lung was measured in COPD patients and the control group. Cigarette smoke- (CS-) induced emphysema mice were generated, and the alteration of α-SMA, NOX4, TGF-ß 1, and collagen I was accessed. The changes of the expression of ECM markers, NOX4, components of TGF-ß/Smad, and MAPK/Akt signaling in human bronchial smooth muscle cells (HBSMCs) were ascertained for delineating mechanisms of NOX4-mediated ROS production on cell differentiation and remodeling in human ASM cells. Results: An increased abundance of NOX4 and TGF-ß 1 proteins in the epithelial cells and ASM of lung was observed in COPD patients compared with the control group. Additionally, an increased abundance expression of NOX4 and α-SMA was observed in the lungs of the CS-induced emphysema mouse model. TGF-ß 1 displayed abilities to increase the oxidative burden and collagen I production, along with enhanced phosphorylation of ERK, p38MAPK, and p-Akt473 in HBSMCs. These effects of TGF-ß 1 could be inhibited by the ROS scavenger N-acetylcysteine (NAC), siRNA-mediated knockdown of Smad3 and NOX4, and pharmacological inhibitors SB203580 (p38MAPK inhibitor) and LY294002 (Akt inhibitor). Conclusions: NOX4-mediated ROS production alters TGF-ß 1-induced cell differentiation and collagen I protein synthesis in HBSMCs in part through the p38MAPK/Akt signaling pathway in a Smad-dependent manner.


Assuntos
Colágeno/metabolismo , Miócitos de Músculo Liso/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Idoso , Animais , Brônquios/citologia , Brônquios/metabolismo , Diferenciação Celular , Linhagem Celular , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Miócitos de Músculo Liso/citologia , NADPH Oxidase 4/genética , NADPH Oxidase 4/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
Int J Mol Sci ; 22(6)2021 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-33810164

RESUMO

The ductus arteriosus (DA) is a physiologic vessel crucial for fetal circulation. As a major regulating factor, the prostaglandin pathway has long been the target for DA patency maintenance or closure. However, the adverse effect of prostaglandins and their inhibitors has been a major unsolved clinical problem. Furthermore, a significant portion of patients with patent DA fail to respond to cyclooxygenase inhibitors that target the prostaglandin pathway. These unresponsive medical patients ultimately require surgical intervention and highlight the importance of exploring pathways independent from this well-recognized prostaglandin pathway. The clinical limitations of prostaglandin-targeting therapeutics prompted us to investigate molecules beyond the prostaglandin pathway. Thus, this article introduces molecules independent from the prostaglandin pathway based on their correlating mechanisms contributing to vascular remodeling. These molecules may serve as potential targets for future DA patency clinical management.


Assuntos
Canal Arterial/metabolismo , Canal Arterial/patologia , Remodelação Vascular , Animais , Biomarcadores , Movimento Celular , Proliferação de Células , Canal Arterial/embriologia , Permeabilidade do Canal Arterial/etiologia , Permeabilidade do Canal Arterial/metabolismo , Matriz Extracelular , Humanos , Miócitos de Músculo Liso/metabolismo , Prostaglandinas/metabolismo , Transdução de Sinais , Remodelação Vascular/genética
20.
J Pharmacol Exp Ther ; 377(2): 284-292, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33758056

RESUMO

Pulmonary hypertension (PH) is a disease of hyperplasia of pulmonary vascular cells. The pentose phosphate pathway (PPP)-a fundamental glucose metabolism pathway-is vital for cell growth. Because treatment of PH is inadequate, our goal was to determine whether inhibition of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP, prevents maladaptive gene expression that promotes smooth muscle cell (SMC) growth, reduces pulmonary artery remodeling, and normalizes hemodynamics in experimental models of PH. PH was induced in mice by exposure to 10% oxygen (Hx) or weekly injection of vascular endothelial growth factor receptor blocker [Sugen5416 (SU); 20 mg kg-1] during exposure to hypoxia (Hx + SU). A novel G6PD inhibitor (N-[(3ß,5α)-17-oxoandrostan-3-yl]sulfamide; 1.5 mg kg-1) was injected daily during exposure to Hx. We measured right ventricle (RV) pressure and left ventricle pressure-volume relationships and gene expression in lungs of normoxic, Hx, and Hx + SU and G6PD inhibitor-treated mice. RV systolic and end-diastolic pressures were higher in Hx and Hx + SU than normoxic control mice. Hx and Hx + SU decreased expression of epigenetic modifiers (writers and erasers), increased hypomethylation of the DNA, and induced aberrant gene expression in lungs. G6PD inhibition decreased maladaptive expression of genes and SMC growth, reduced pulmonary vascular remodeling, and decreased right ventricle pressures compared with untreated PH groups. Pharmacologic inhibition of G6PD activity, by normalizing activity of epigenetic modifiers and DNA methylation, efficaciously reduces RV pressure overload in Hx and Hx + SU mice and preclinical models of PH and appears to be a safe pharmacotherapeutic strategy. SIGNIFICANCE STATEMENT: The results of this study demonstrated that inhibition of a metabolic enzyme efficaciously reduces pulmonary hypertension. For the first time, this study shows that a novel inhibitor of glucose-6-phosphate dehydrogenase, the rate-limiting enzyme in the fundamental pentose phosphate pathway, modulates DNA methylation and alleviates pulmonary artery remodeling and dilates pulmonary artery to reduce pulmonary hypertension.


Assuntos
Cardiomegalia/tratamento farmacológico , Inibidores Enzimáticos/uso terapêutico , Glucosefosfato Desidrogenase/antagonistas & inibidores , Hipertensão Pulmonar/tratamento farmacológico , Animais , Pressão Sanguínea , Cardiomegalia/etiologia , Hipóxia Celular , Células Cultivadas , Metilação de DNA , Inibidores Enzimáticos/farmacologia , Feminino , Humanos , Hipertensão Pulmonar/etiologia , Indóis/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Pirróis/toxicidade , Função Ventricular Direita
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