RESUMO
Torilis japonica (TJ) fruit, is a herb that is traditionally used for erectile dysfunction (ED). Given the shared mechanisms of ED and hypertension through vascular smooth muscle, we hypothesized that TJ would be effective in vasodilation and blood pressure reduction. This study confirmed the authenticity of TJ samples via DNA barcoding and quantified the main active compound, torilin, using HPLC. TJ was extracted with distilled water (TJW) and 50% ethanol (TJE), yielding torilin contents of 0.35 ± 0.01% and 2.84 ± 0.02%, respectively. Ex vivo tests on thoracic aortic rings from Sprague-Dawley rats showed that TJE (3-300 µg/mL) induced endothelium-independent, concentration-dependent vasodilation, unlike TJW. Torilin caused concentration-dependent relaxation with an EC50 of 210 ± 1.07 µM. TJE's effects were blocked by a voltage-dependent K+ channel blocker and alleviated contractions induced by CaCl2 and angiotensin II. TJE inhibited vascular contraction induced by phenylephrine or KCl via extracellular CaCl2 and enhanced inhibition with nifedipine, indicating involvement of voltage-dependent and receptor-operated Ca2+ channels. Oral administration of TJE (1000 mg/kg) significantly reduced blood pressure in spontaneously hypertensive rats. These findings suggest TJ extract's potential for hypertension treatment through vasorelaxant mechanisms, though further research is needed to confirm its efficacy and safety.
Assuntos
Pressão Sanguínea , Endotélio Vascular , Frutas , Extratos Vegetais , Ratos Sprague-Dawley , Vasodilatação , Animais , Ratos , Vasodilatação/efeitos dos fármacos , Extratos Vegetais/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Masculino , Frutas/química , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Anti-Hipertensivos/farmacologia , Vasodilatadores/farmacologia , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Ratos Endogâmicos SHR , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Hipertensão/fisiopatologiaRESUMO
BACKGROUND: Cellular communication among different types of vascular cells is indispensable for maintaining vascular homeostasis and preventing atherosclerosis. However, the biological mechanism involved in cellular communication among these cells and whether this biological mechanism can be used to treat atherosclerosis remain unknown. We hypothesized that endothelial autophagy mediates the cellular communication in vascular tissue through exosome-mediated delivery of atherosclerosis-related genes. METHODS: Rapamycin and adeno-associated virus carrying Atg7 short hairpin RNA under the Tie (TEK receptor tyrosine kinase) promoter were used to activate and inhibit vascular endothelial autophagy in high-fat diet-fed ApoE-/- mice, respectively. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue were used to explore the effects of endothelial autophagy on endothelial function and atherosclerosis and its molecular mechanisms. Quantitative polymerase chain reaction and miRNA sequencing were performed to determine changes in miRNA expression in exosomes. Immunofluorescence and exosome coculture experiments were conducted to examine the role of endothelial autophagy in regulating the communication between endothelial cells and smooth muscle cells (SMCs) via exosomal miRNA. RESULTS: Endothelial autophagy was inhibited in thoracic aortas of high-fat diet-fed ApoE-/- mice. Furthermore, rapamycin alleviated high-fat diet-induced atherosclerotic burden and endothelial dysfunction, while endothelial-specific Atg7 depletion aggravated the atherosclerotic burden. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue analysis revealed that miR-204-5p was significantly increased in endothelial cells after high-fat diet exposure, which directly targeted Bcl2 to regulate endothelial cell apoptosis. Importantly, endothelial autophagy activation decreased excess miR-204-5p by loading miR-204-5p into multivesicular bodies and secreting it through exosomes. Moreover, exosomal miR-204-5p can effectively transport to SMCs, alleviating SMC calcification by regulating target proteins such as RUNX2 (runt-related transcription factor 2). CONCLUSIONS: Our study revealed the exosomal pathway by which endothelial autophagy protects atherosclerosis: endothelial autophagy activation transfers miR-204-5p from endothelial cells to SMCs via exosomes, both preventing endothelial apoptosis and alleviating SMC calcification. REGISTRATION: URL: https://www.chictr.org.cn/; Unique identifier: ChiCTR2200064155.
Assuntos
Aterosclerose , Autofagia , Comunicação Celular , Modelos Animais de Doenças , Exossomos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , MicroRNAs , Miócitos de Músculo Liso , MicroRNAs/metabolismo , MicroRNAs/genética , Exossomos/metabolismo , Exossomos/genética , Animais , Aterosclerose/patologia , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/prevenção & controle , Humanos , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Masculino , Camundongos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Proteína 7 Relacionada à Autofagia/metabolismo , Proteína 7 Relacionada à Autofagia/genética , Células Cultivadas , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Placa Aterosclerótica , Doenças da Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/prevenção & controle , Doenças da Aorta/metabolismo , Técnicas de Cocultura , Transdução de Sinais , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Dieta HiperlipídicaAssuntos
Aorta Torácica , Aneurisma da Aorta Torácica , Síndrome de Marfan , Músculo Liso Vascular , Miócitos de Músculo Liso , Proteômica , Síndrome de Marfan/metabolismo , Síndrome de Marfan/complicações , Síndrome de Marfan/diagnóstico , Síndrome de Marfan/genética , Aneurisma da Aorta Torácica/metabolismo , Aneurisma da Aorta Torácica/patologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Humanos , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Músculo Liso Vascular/efeitos dos fármacos , Células Cultivadas , AnimaisRESUMO
BACKGROUND: In early atherosclerosis, circulating LDLs (low-density lipoproteins) traverse individual endothelial cells by an active process termed transcytosis. The CANTOS trial (Canakinumab Antiinflammatory Thrombosis Outcome Study) treated advanced atherosclerosis using a blocking antibody for IL-1ß (interleukin-1ß); this significantly reduced cardiovascular events. However, whether IL-1ß regulates early disease, particularly LDL transcytosis, remains unknown. METHODS: We used total internal reflection fluorescence microscopy to quantify transcytosis by human coronary artery endothelial cells exposed to IL-1ß. To investigate transcytosis in vivo, we injected wild-type and knockout mice with IL-1ß and LDL to visualize acute LDL deposition in the aortic arch. RESULTS: Exposure to picomolar concentrations of IL-1ß induced transcytosis of LDL but not of albumin by human coronary artery endothelial cells. Surprisingly, expression of the 2 known receptors for LDL transcytosis, ALK-1 (activin receptor-like kinase-1) and SR-BI (scavenger receptor BI), was unchanged or decreased. Instead, IL-1ß increased the expression of the LDLR (LDL receptor); this was unexpected because LDLR is not required for LDL transcytosis. Overexpression of LDLR had no effect on basal LDL transcytosis. However, knockdown of LDLR abrogated the effect of IL-1ß on transcytosis rates while the depletion of Cav-1 (caveolin-1) did not. Since LDLR was necessary but overexpression had no effect, we reasoned that another player must be involved. Using public RNA sequencing data to curate a list of Rab (Ras-associated binding) GTPases affected by IL-1ß, we identified Rab27a. Overexpression of Rab27a alone had no effect on basal transcytosis, but its knockdown prevented induction by IL-1ß. This was phenocopied by depletion of the Rab27a effector JFC1 (synaptotagmin-like protein 1). In vivo, IL-1ß increased LDL transcytosis in the aortic arch of wild-type but not Ldlr-/- or Rab27a-deficient mice. The JFC1 inhibitor nexinhib20 also blocked IL-1ß-induced LDL accumulation in the aorta. CONCLUSIONS: IL-1ß induces LDL transcytosis by a distinct pathway requiring LDLR and Rab27a; this route differs from basal transcytosis. We speculate that induction of transcytosis by IL-1ß may contribute to the acceleration of early disease.
Assuntos
Vasos Coronários , Células Endoteliais , Interleucina-1beta , Lipoproteínas LDL , Camundongos Knockout , Receptores de LDL , Transdução de Sinais , Transcitose , Proteínas rab de Ligação ao GTP , Interleucina-1beta/metabolismo , Animais , Humanos , Receptores de LDL/genética , Receptores de LDL/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Proteínas rab de Ligação ao GTP/genética , Células Endoteliais/metabolismo , Células Endoteliais/efeitos dos fármacos , Lipoproteínas LDL/metabolismo , Vasos Coronários/metabolismo , Vasos Coronários/efeitos dos fármacos , Células Cultivadas , Camundongos Endogâmicos C57BL , Caveolina 1/metabolismo , Caveolina 1/genética , Doenças da Aorta/metabolismo , Doenças da Aorta/genética , Doenças da Aorta/patologia , Modelos Animais de Doenças , Aorta Torácica/metabolismo , Aorta Torácica/efeitos dos fármacos , Aterosclerose/metabolismo , Aterosclerose/genética , Aterosclerose/patologia , Masculino , CamundongosRESUMO
BACKGROUND: The prevention of obesity represents a major health and socio-economic challenge. Nutraceuticals are regularly highlighted for their beneficial effects in preventing the metabolic disturbances associated with obesity. However, few studies have described the combined action of nutraceutical mixtures combining polyphenols with alkaloids. OBJECTIVE: The aim of this study was to evaluate the effects of long-term dietary supplementation with a mixture of Berberine, Citrus and Apple extracts (BCA) in the primary prevention of obesity and its metabolic and vascular complications in the obese Zucker rat, a spontaneous model of genetic obesity and insulin resistance. METHODS: Sixteen 8-week-old obese Zucker male rats were randomly divided into two groups: all rats received oral gavage daily either with water, untreated obese (U-ObZ) or BCA (BCA-ObZ) mixture for thirteen weeks. Morphological and metabolic parameters were measured along the study. Cumulative concentration-response curves to insulin, acetylcholine and phenylephrine were determined on isolated thoracic aorta. Colon permeability measurements were performed using the Ussing chamber technique. Fecal samples collected at the beginning and the end of the protocol were used as a template for amplification of the V3-V4 region of the 16S rDNA genes. RESULTS: BCA supplementation reduced weight gain (p<0.05) and food intake (p<0.05) in the BCA-ObZ group rats compared to the U-ObZ group rats. It also improved glucose tolerance (p<0.001) and decreased fasting insulin and Homeostasis model assessment index (p<0.05). Through ex vivo experiments, the BCA mixture enhanced significantly aortic insulin relaxation (p<0.01), reduced α1-adrenoceptor-mediated vasoconstriction (p<0.01), and decreased distal colon permeability. Moreover, short-chain fatty acid producers such as Bacteroides, Blautia, and Akkermansia were found to be increased by the BCA mixture supplementation. CONCLUSION: The results showed that a 13-week-supplementation with BCA mixture prevented weight gain and improved glucose metabolism in obese Zucker rats. We also demonstrated that BCA supplementation improved vascular function, colonic barrier permeability and gut microbiota profile.
Assuntos
Berberina , Citrus , Suplementos Nutricionais , Malus , Obesidade , Extratos Vegetais , Ratos Zucker , Animais , Berberina/farmacologia , Berberina/uso terapêutico , Masculino , Obesidade/metabolismo , Obesidade/prevenção & controle , Obesidade/tratamento farmacológico , Ratos , Citrus/química , Extratos Vegetais/farmacologia , Malus/química , Resistência à Insulina , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Insulina/sangue , Insulina/metabolismoRESUMO
Perivascular adipose tissue (PVAT) is an adipose layer, surrounding blood vessels, with a local modulatory role. Interleukin-10 (IL-10) has been shown to modulate vascular tissue. This study aimed to characterize the endogenous role of IL-10 in vascular remodeling, and PVAT phenotyping. Thoracic aortic segments from control (C57BL/6J) and IL-10 knockout (IL-10-/-) male mice were used. Analyzes of aorta/PVAT morphometry, and elastin, collagen and reticulin deposition were performed. Tissue uncoupling protein 1 (UCP1) was accessed by Western blotting. Endogenous absence of IL-10 reduced total PVAT area (p = 0.0310), and wall/lumen ratio (p = 0.0024), whereas increased vascular area and thickness (p < 0.0001). Total collagen deposition was augmented in IL-10-/-, but under polarized light, the reduction of collagen-I (p = 0.0075) and the increase of collagen-III (p = 0.0055) was found, simultaneously with reduced elastic fibers deposition (p = 0.0282) and increased deposition of reticular fibers (p < 0.0001). Adipocyte area was augmented in the IL-10 absence (p = 0.0225), and UCP1 expression was reduced (p = 0.0420). Moreover, relative frequency of white adipose cells and connective tissue was augmented in IL-10-/- (p < 0.0001), added to a reduction in brown adipose cells (p < 0.0001). Altogether, these data characterize aorta PVAT from IL-10-/- as a white-like adipocyte phenotype. Endogenous IL-10 prevents vascular remodeling and favors a brown-like adipocyte phenotype, suggesting a modulatory role for IL-10 in PVAT plasticity.
Assuntos
Tecido Adiposo Branco , Interleucina-10 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Remodelação Vascular , Animais , Interleucina-10/metabolismo , Interleucina-10/genética , Remodelação Vascular/genética , Masculino , Camundongos , Tecido Adiposo Branco/metabolismo , Proteína Desacopladora 1/metabolismo , Proteína Desacopladora 1/genética , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Colágeno/metabolismo , Tecido Adiposo/metabolismoRESUMO
Perivascular adipose tissue (PVAT) is known for being anti-contractile in healthy tissues. We discovered a new function of PVAT, the ability to stress relax and maintain a tone in response to a stretch. This is of note because stress relaxation has been attributed to smooth muscle, of which PVAT has none that is organized in a functional layer. We test the hypothesis the interactions of integrins with collagen play a role in stress relaxation. Our model is the thoracic aorta of the male Dahl SS rat. The PVAT and aorta were physically separated for most assays. Results from single nuclei RNA sequencing (snRNAseq) experiments, histochemistry and isometric contractility were also used. Masson Trichrome staining made evident the expression of collagen in PVAT. From snRNA seq experiments of the PVAT, mRNA for multiple collagen and integrin isoforms were detected: the α1 and ß1 integrin were most highly expressed. Pharmacological inhibition of integrin/collagen interaction was effected by the specific α1ß1 distintegrin obtustatin or general integrin inhibitor RGD peptide. RGD peptide but not obtustatin increased the stress relaxation. Cell-cell communication inference identified integrins αv and α5, two major RGD motif containing isoforms, as potential signaling partners of collagens. Collectively, these findings validate that stress relaxation can occur in a non-smooth muscle tissue, doing so in part through integrin-collagen interactions that may not include α1ß1 heterodimers. The importance of this lies in considering PVAT as a vascular layer that possesses mechanical functions.
Assuntos
Tecido Adiposo , Aorta Torácica , Colágeno , Integrinas , Ratos Endogâmicos Dahl , Animais , Masculino , Tecido Adiposo/metabolismo , Integrinas/metabolismo , Aorta Torácica/metabolismo , Colágeno/metabolismo , RatosRESUMO
Aortic stiffening is an inevitable manifestation of chronological aging, yet the mechano-molecular programs that orchestrate region- and layer-specific adaptations along the length and through the wall of the aorta are incompletely defined. Here, we show that the decline in passive cyclic distensibility is more pronounced in the ascending thoracic aorta (ATA) compared to distal segments of the aorta and that collagen content increases in both the medial and adventitial compartments of the ATA during aging. The single-cell RNA sequencing of aged ATA tissues reveals altered cellular senescence, remodeling, and inflammatory responses accompanied by enrichment of T-lymphocytes and rarefaction of vascular smooth muscle cells, compared to young samples. T lymphocyte clusters accumulate in the adventitia, while the activation of mechanosensitive Piezo-1 enhances vasoconstriction and contributes to the overall functional decline of ATA tissues. These results portray the immuno-mechanical aging of the ATA as a process that culminates in a stiffer conduit permissive to the accrual of multi-gerogenic signals priming to disease development.
Assuntos
Envelhecimento , Aorta Torácica , Transcriptoma , Envelhecimento/fisiologia , Envelhecimento/genética , Transcriptoma/genética , Aorta Torácica/metabolismo , Humanos , Mecanotransdução Celular , Masculino , AnimaisRESUMO
BACKGROUND: Vasculopathy is the most common complication of diabetes. Endothelial cells located in the innermost layer of blood vessels are constantly affected by blood flow or vascular components; thus, their mechanosensitivity plays an important role in mediating vascular regulation. Endothelial damage, one of the main causes of hyperglycemic vascular complications, has been extensively studied. However, the role of mechanosensitive signaling in hyperglycemic endothelial damage remains unclear. METHODS: Vascular endothelial-specific Piezo1 knockout mice were generated to investigate the effects of Piezo1 on Streptozotocin-induced hyperglycemia and vascular endothelial injury. In vitro activation or knockdown of Piezo1 was performed to evaluate the effects on the proliferation, migration, and tubular function of human umbilical vein endothelial cells in high glucose. Reactive oxygen species production, mitochondrial membrane potential alternations, and oxidative stress-related products were used to assess the extent of oxidative stress damage caused by Piezo1 activation. RESULTS: Our study found that in VECreERT2;Piezo1flox/flox mice with Piezo1 conditional knockout in vascular endothelial cells, Piezo1 deficiency alleviated streptozotocin-induced hyperglycemia with reduced apoptosis and abscission of thoracic aortic endothelial cells, and decreased the inflammatory response of aortic tissue caused by high glucose. Moreover, the knockout of Piezo1 showed a thinner thoracic aortic wall, reduced tunica media damage, and increased endothelial nitric oxide synthase expression in transgenic mice, indicating the relief of endothelial damage caused by hyperglycemia. We also showed that Piezo1 activation aggravated oxidative stress injury and resulted in severe dysfunction through the Ca2+-induced CaMKII-Nrf2 axis in human umbilical vein endothelial cells. In Piezo1 conditional knockout mice, Piezo1 deficiency partially restored superoxide dismutase activity and reduced malondialdehyde content in the thoracic aorta. Mechanistically, Piezo1 deficiency decreased CaMKII phosphorylation and restored the expression of Nrf2 and its downstream molecules HO-1 and NQO1. CONCLUSION: In summary, our study revealed that Piezo1 is involved in high glucose-induced oxidative stress injury and aggravated endothelial dysfunction, which have great significance for alleviating endothelial damage caused by hyperglycemia.
Assuntos
Glicemia , Diabetes Mellitus Experimental , Células Endoteliais da Veia Umbilical Humana , Canais Iônicos , Camundongos Knockout , Óxido Nítrico Sintase Tipo III , Estresse Oxidativo , Animais , Humanos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Diabetes Mellitus Experimental/metabolismo , Canais Iônicos/metabolismo , Canais Iônicos/genética , Glicemia/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Mecanotransdução Celular , Fator 2 Relacionado a NF-E2/metabolismo , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/deficiência , Células Cultivadas , Proliferação de Células , Apoptose , Masculino , Angiopatias Diabéticas/metabolismo , Angiopatias Diabéticas/fisiopatologia , Angiopatias Diabéticas/patologia , Angiopatias Diabéticas/genética , Angiopatias Diabéticas/etiologia , Movimento Celular , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismo , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Camundongos , Estreptozocina , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Endotélio Vascular/patologia , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/genéticaRESUMO
Endothelial cells (ECs) in the descending aorta are exposed to high laminar shear stress, and this supports an antiinflammatory phenotype. High laminar shear stress also induces flow-aligned cell elongation and front-rear polarity, but whether these are required for the antiinflammatory phenotype is unclear. Here, we showed that caveolin-1-rich microdomains polarize to the downstream end of ECs that are exposed to continuous high laminar flow. These microdomains were characterized by high membrane rigidity, filamentous actin (F-actin), and raft-associated lipids. Transient receptor potential vanilloid (TRPV4) ion channels were ubiquitously expressed on the plasma membrane but mediated localized Ca2+ entry only at these microdomains where they physically interacted with clustered caveolin-1. These focal Ca2+ bursts activated endothelial nitric oxide synthase within the confines of these domains. Importantly, we found that signaling at these domains required both cell body elongation and sustained flow. Finally, TRPV4 signaling at these domains was necessary and sufficient to suppress inflammatory gene expression and exogenous activation of TRPV4 channels ameliorated the inflammatory response to stimuli both in vitro and in vivo. Our work revealed a polarized mechanosensitive signaling hub in arterial ECs that dampened inflammatory gene expression and promoted cell resilience.
Assuntos
Cálcio , Células Endoteliais , Inflamação , Mecanotransdução Celular , Canais de Cátion TRPV , Canais de Cátion TRPV/metabolismo , Canais de Cátion TRPV/genética , Animais , Inflamação/metabolismo , Inflamação/patologia , Inflamação/genética , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Cálcio/metabolismo , Camundongos , Humanos , Microdomínios da Membrana/metabolismo , Caveolina 1/metabolismo , Caveolina 1/genética , Sinalização do Cálcio , Estresse Mecânico , Aorta Torácica/metabolismo , Aorta Torácica/patologiaRESUMO
Apigenin and baicalein are structurally related flavonoids that have been reported to have multiple pharmacological activities. The aim of this study was to investigate the protective effects and potential mechanisms of apigenin and baicalein in D-galactose-induced aging rats. First, apigenin and baicalein showed remarkable antioxidant activity and anti-glycation activity in vitro. Secondly, the protective effects of apigenin and baicalein on aging rats were investigated. We found that apigenin and baicalein supplementation significantly ameliorated aging-related changes such as declines in the spatial learning and memory and histopathological damage of the hippocampus and thoracic aorta. In addition, our data showed that apigenin and baicalein alleviated oxidative stress as illustrated by decreasing MDA level, increasing SOD activity and GSH level. Further data showed that they significantly reduced the accumulation of advanced glycation end products (AGEs), inhibited the expression of RAGE, down-regulated phosphorylated nuclear factor (p-NF-κB (p65)). Our results suggested that the protective effects of apigenin and baicalein on aging rats were at least partially related to the inhibition of AGEs/RAGE/NF-κB pathway and the improvement of oxidative damage. Overall, apigenin and baicalein showed almost equal anti-aging efficacy. Our results provided an experimental basis for the application of apigenin and baicalein to delay the aging process.
Assuntos
Envelhecimento , Aorta Torácica , Apigenina , Flavanonas , Galactose , Produtos Finais de Glicação Avançada , NF-kappa B , Estresse Oxidativo , Ratos Sprague-Dawley , Receptor para Produtos Finais de Glicação Avançada , Transdução de Sinais , Animais , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Flavanonas/farmacologia , Flavanonas/uso terapêutico , Apigenina/farmacologia , Apigenina/uso terapêutico , Envelhecimento/efeitos dos fármacos , Envelhecimento/metabolismo , Masculino , NF-kappa B/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipocampo/patologia , Disfunção Cognitiva/tratamento farmacológico , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/induzido quimicamente , Antioxidantes/farmacologiaRESUMO
ABSTRACT: Vascular calcification (VC), a major complication in chronic kidney disease (CKD), is predominantly driven by osteoblastic differentiation. Recent studies have highlighted the crucial role of microRNAs in CKD's pathogenesis. Here, our research focused on the effects of miR-204-5p and its molecular mechanisms within VC. We initially found a notable decrease in miR-204-5p levels in human aortic vascular smooth muscle cells stimulated with inorganic phosphate, using this as a VC model in vitro. Following the overexpression of miR-204-5p, a decrease in VC was observed, as indicated by alizarin red S staining and measurements of calcium content. This decrease was accompanied by lower levels of the osteogenic marker, runt-related transcription factor 2, and higher levels of α-smooth muscle actin, a marker of contractility. Further investigation showed that calcium/calmodulin-dependent protein kinase 1 (CAMK1), which is a predicted target of miR-204-5p, promotes VC. Conversely, overexpressing miR-204-5p reduced VC by suppressing CAMK1 activity. Overexpressing miR-204-5p also effectively mitigated aortic calcification in an in vivo rat model. In summary, our research indicated that targeting the miR-204-5p/CAMK1 pathway could be a viable strategy for mitigating VC in CKD patients.
Assuntos
Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina , MicroRNAs , Insuficiência Renal Crônica , Calcificação Vascular , Masculino , Animais , Ratos , Ratos Sprague-Dawley , Aorta Torácica/metabolismo , Aorta Torácica/patologia , MicroRNAs/farmacologia , Proteína Quinase Tipo 1 Dependente de Cálcio-Calmodulina/metabolismo , Miócitos de Músculo Liso/metabolismo , Diferenciação Celular , Calcificação Vascular/tratamento farmacológico , Calcificação Vascular/metabolismo , Calcificação Vascular/patologia , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/metabolismo , Fosfatos/metabolismoRESUMO
BACKGROUND: ß-aminopropionitrile (BAPN) is a pharmacological inhibitor of LOX (lysyl oxidase) and LOXLs (LOX-like proteins). Administration of BAPN promotes aortopathies, although there is a paucity of data on experimental conditions to generate pathology. The objective of this study was to define experimental parameters and determine whether equivalent or variable aortopathies were generated throughout the aortic tree during BAPN administration in mice. METHODS: BAPN was administered in drinking water for a period ranging from 1 to 12 weeks. The impacts of BAPN were first assessed with regard to BAPN dose, and mouse strain, age, and sex. BAPN-induced aortic pathological characterization was conducted using histology and immunostaining. To investigate the mechanistic basis of regional heterogeneity, the ascending and descending thoracic aortas were harvested after 1 week of BAPN administration before the appearance of overt pathology. RESULTS: BAPN-induced aortic rupture predominantly occurred or originated in the descending thoracic aorta in young C57BL/6J or N mice. No apparent differences were found between male and female mice. For mice surviving 12 weeks of BAPN administration, profound dilatation was consistently observed in the ascending region, while there were more heterogeneous changes in the descending thoracic region. Pathological features were distinct between the ascending and descending thoracic regions. Aortic pathology in the ascending region was characterized by luminal dilatation and elastic fiber disruption throughout the media. The descending thoracic region frequently had dissections with false lumen formation, collagen deposition, and remodeling of the wall surrounding the false lumen. Cells surrounding the false lumen were predominantly positive for α-SMA (α-smooth muscle actin). One week of BAPN administration compromised contractile properties in both regions equivalently, and RNA sequencing did not show obvious differences between the 2 aortic regions in smooth muscle cell markers, cell proliferation markers, and extracellular components. CONCLUSIONS: BAPN-induced pathologies show distinct, heterogeneous features within and between ascending and descending aortic regions in mice.
Assuntos
Aminopropionitrilo , Aorta Torácica , Ruptura Aórtica , Modelos Animais de Doenças , Camundongos Endogâmicos C57BL , Animais , Aminopropionitrilo/toxicidade , Aminopropionitrilo/farmacologia , Aorta Torácica/patologia , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Feminino , Masculino , Ruptura Aórtica/induzido quimicamente , Ruptura Aórtica/patologia , Ruptura Aórtica/metabolismo , Ruptura Aórtica/prevenção & controle , Camundongos , Remodelação Vascular/efeitos dos fármacos , Dilatação Patológica , Músculo Liso Vascular/patologia , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Fatores Etários , Fatores de Tempo , Fatores Sexuais , Proliferação de Células/efeitos dos fármacos , Proteína-Lisina 6-Oxidase/metabolismoRESUMO
Long-term ß-adrenoceptor (ß-AR) stimulation is a pathological mechanism associated with cardiovascular diseases resulting in endothelial and perivascular adipose tissue (PVAT) dysfunction. In this study, we aimed to identify whether ß-adrenergic signaling has a direct effect on PVAT. Thoracic aorta PVAT was obtained from male Wistar rats and cultured ex vivo with the ß-AR agonist isoproterenol (Iso; 1â µM) or vehicle for 24 hours. Conditioned culture medium (CCM) from Iso-treated PVAT induced a marked increase in aorta contractile response, induced oxidative stress, and reduced nitric oxide production in PVAT compared to vehicle. In addition, Iso-treated PVAT and PVAT-derived differentiated adipocytes exhibited higher corticosterone release and protein expression of 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1), an enzyme responsible for de novo synthesis of corticosterone. Macrophages exposed to Iso also exhibited increased corticosterone release in response to ß-AR stimulation. Incubation of Iso-treated PVAT and PVAT-derived differentiated adipocytes with ß3-AR antagonist restored aorta contractile function modulated by Iso-CCM and normalized 11ß-HSD1 protein expression. These results show that ß3-AR signaling leads to upregulation of 11ß-HSD1 in PVAT, thus increasing corticosterone release and contributing to impair the anticontractile function of this tissue.
Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1 , Corticosterona , Isoproterenol , Animais , Masculino , Ratos , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/genética , Adipócitos/metabolismo , Adipócitos/efeitos dos fármacos , Tecido Adiposo/metabolismo , Agonistas Adrenérgicos beta/farmacologia , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Corticosterona/metabolismo , Meios de Cultivo Condicionados/farmacologia , Isoproterenol/farmacologia , Óxido Nítrico/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos Wistar , Receptores Adrenérgicos beta/metabolismoRESUMO
BACKGROUND: The resiliency of embryonic development to genetic and environmental perturbations has been long appreciated; however, little is known about the mechanisms underlying the robustness of developmental processes. Aberrations resulting in neonatal lethality are exemplified by congenital heart disease arising from defective morphogenesis of pharyngeal arch arteries (PAAs) and their derivatives. METHODS: Mouse genetics, lineage tracing, confocal microscopy, and quantitative image analyses were used to investigate mechanisms of PAA formation and repair. RESULTS: The second heart field (SHF) gives rise to the PAA endothelium. Here, we show that the number of SHF-derived endothelial cells (ECs) is regulated by VEGFR2 (vascular endothelial growth factor receptor 2) and Tbx1. Remarkably, when the SHF-derived EC number is decreased, PAA development can be rescued by the compensatory endothelium. Blocking such compensatory response leads to embryonic demise. To determine the source of compensating ECs and mechanisms regulating their recruitment, we investigated 3-dimensional EC connectivity, EC fate, and gene expression. Our studies demonstrate that the expression of VEGFR2 by the SHF is required for the differentiation of SHF-derived cells into PAA ECs. The deletion of 1 VEGFR2 allele (VEGFR2SHF-HET) reduces SHF contribution to the PAA endothelium, while the deletion of both alleles (VEGFR2SHF-KO) abolishes it. The decrease in SHF-derived ECs in VEGFR2SHF-HET and VEGFR2SHF-KO embryos is complemented by the recruitment of ECs from the nearby veins. Compensatory ECs contribute to PAA derivatives, giving rise to the endothelium of the aortic arch and the ductus in VEGFR2SHF-KO mutants. Blocking the compensatory response in VEGFR2SHF-KO mutants results in embryonic lethality shortly after mid-gestation. The compensatory ECs are absent in Tbx1+/- embryos, a model for 22q11 deletion syndrome, leading to unpredictable arch artery morphogenesis and congenital heart disease. Tbx1 regulates the recruitment of the compensatory endothelium in an SHF-non-cell-autonomous manner. CONCLUSIONS: Our studies uncover a novel buffering mechanism underlying the resiliency of PAA development and remodeling.
Assuntos
Aorta Torácica , Células Endoteliais , Cardiopatias Congênitas , Proteínas com Domínio T , Receptor 2 de Fatores de Crescimento do Endotélio Vascular , Animais , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Camundongos , Aorta Torácica/embriologia , Aorta Torácica/metabolismo , Cardiopatias Congênitas/genética , Cardiopatias Congênitas/metabolismo , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/embriologia , Proteínas com Domínio T/metabolismo , Proteínas com Domínio T/genética , Células Endoteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Diferenciação Celular , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: The pineal product melatonin (MEL) modulates blood vessels through G protein-coupled receptors (GPCRs) called melatonin type 1 receptor (MT1R) and melatonin type 2 receptor (MT2R), in that order. The renin-angiotensin system (RAS), which breaks down angiotensin II (Ang II) to create Ang 1-7, is thought to be mostly controlled by angiotensin-converting enzyme-2 (ACE2). AIM: The current work examines the involvement of ACE2 inhibitor, MEL, and ramelteon (RAM) in the vascular response to Ang II activities in the endothelial denuded (E-) and intact (E+) rat isolated thoracic aortic rings. METHOD: The isometric tension was measured to evaluate the vascular Ang II contractility using dose response curve (DRC). RESULTS: MEL and RAM caused a rightward shift of Ang II in endothelium E + and endothelium E- aorta. CONCLUSION: According to the current study, the distribution of MEL receptors and the endothelium's condition are related to the vasomodulatory effect of MEL and ACE2 on Ang II attenuation. These physiological interactions can control vascular tone and increase Ang II reactivity denude endothelial layaer.
Assuntos
Angiotensina II , Enzima de Conversão de Angiotensina 2 , Melatonina , Animais , Melatonina/farmacologia , Angiotensina II/metabolismo , Angiotensina II/farmacologia , Ratos , Enzima de Conversão de Angiotensina 2/metabolismo , Sistema Renina-Angiotensina/efeitos dos fármacos , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/metabolismo , Masculino , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Peptidil Dipeptidase A/metabolismo , Aorta/efeitos dos fármacos , Aorta/metabolismo , Receptor MT2 de Melatonina/metabolismo , Receptor MT2 de Melatonina/antagonistas & inibidores , Inibidores da Enzima Conversora de Angiotensina/farmacologiaRESUMO
Artificial light at night (ALAN) disrupts 24-h variability of blood pressure, but the molecular mechanisms underlying these effects are unknown. Therefore, we analysed the daily variability of pulse pressure, the maximum value of acceleration rate of aortic pressure (dP/dt(max)) measured by telemetry and protein expression in the thoracic aorta of normotensive male rats exposed to ALAN (1-2 lx) for 3 weeks. Daily, 24-h variability of pulse pressure and dP/dt(max) was observed during a regular light/dark regimen with higher values during the dark compared to the light phase of the day. ALAN suppressed 24-h variability and enhanced ultradian (<12-h) periods of pulse pressure and dP/dt(max) in duration-dependent manners. From beat-to-beat blood pressure variability, ALAN decreased low-frequency bands (a sympathetic marker) and had minimal effects on high-frequency bands. At the molecular level, ALAN decreased angiotensin II receptor type 1 expression and reduced 24-h variability. ALAN caused the appearance of 12-h oscillations in transforming growth factor ß1 and fibulin 4. Expression of sarco/endoplasmic reticulum Ca2+-ATPase type 2 was increased in the middle of the light and dark phase of the day, and ALAN did not affect its daily and 12-h variability. In conclusion, ALAN suppressed 24-h variability of pulse pressure and dP/dt(max), decreased the power of low-frequency bands and differentially affected the expression of specific proteins in the rat thoracic aorta. Suppressed 24-h oscillations by ALAN underline the pulsatility of individual endocrine axes with different periods, disrupting the cardiovascular control of central blood pressure.
Assuntos
Aorta Torácica , Pressão Sanguínea , Ritmo Circadiano , Animais , Masculino , Aorta Torácica/metabolismo , Pressão Sanguínea/fisiologia , Ratos , Ritmo Circadiano/fisiologia , Fator de Crescimento Transformador beta1/metabolismo , Receptor Tipo 1 de Angiotensina/metabolismo , Luz , Proteínas da Matriz Extracelular/metabolismo , Ratos Sprague-Dawley , Proteínas de Ligação ao Cálcio/metabolismoRESUMO
Perivascular adipose tissue (PVAT) is increasingly recognized for its function in mechanotransduction. However, major gaps remain in our understanding of the cells present in PVAT, as well as how different cells contribute to mechanotransduction. We hypothesized that snRNA-seq would reveal the expression of mechanotransducers, and test one (PIEZO1) to illustrate the expression and functional agreement between single-nuclei RNA sequencing (snRNA-seq) and physiological measurements. To contrast two brown tissues, subscapular brown adipose tissue (BAT) was also examined. We used snRNA-seq of the thoracic aorta PVAT (taPVAT) and BAT from male Dahl salt-sensitive (Dahl SS) rats to investigate cell-specific expression mechanotransducers. Localization and function of the mechanostransducer PIEZO1 were further examined using immunohistochemistry (IHC) and RNAscope, as well as pharmacological antagonism. Approximately 30,000 nuclei from taPVAT and BAT each were characterized by snRNA-seq, identifying eight major cell types expected and one unexpected (nuclei with oligodendrocyte marker genes). Cell-specific differential gene expression analysis between taPVAT and BAT identified up to 511 genes (adipocytes) with many (≥20%) being unique to individual cell types. Piezo1 was the most highly, widely expressed mechanotransducer. The presence of PIEZO1 in the PVAT but not the adventitia was confirmed by RNAscope and IHC in male and female rats. Importantly, antagonism of PIEZO1 by GsMTX4 impaired the PVAT's ability to hold tension. Collectively, the cell compositions of taPVAT and BAT are highly similar, and PIEZO1 is likely a mechanotransducer in taPVAT.NEW & NOTEWORTHY This study describes the atlas of cells in the thoracic aorta perivascular adipose tissue (taPVAT) of the Dahl-SS rat, an important hypertension model. We show that mechanotransducers are widely expressed in these cells. Moreover, PIEZO1 expression is shown to be restricted to the taPVAT and is functionally implicated in stress relaxation. These data will serve as the foundation for future studies investigating the role of taPVAT in this model of hypertensive disease.
Assuntos
Tecido Adiposo Marrom , Aorta Torácica , Canais Iônicos , Mecanotransdução Celular , Proteínas de Membrana , Ratos Endogâmicos Dahl , Animais , Aorta Torácica/metabolismo , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Masculino , Canais Iônicos/metabolismo , Canais Iônicos/genética , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo/metabolismo , Ratos , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Hipertensão/genética , Hipertensão/patologia , RNA-SeqRESUMO
AIM: The present study evaluated whether topiramate (TPM) treatment during the peripubertal period affects vascular parameters of male rats and whether oxidative stress plays a role in these changes. MAIN METHODS: Rats were treated with TPM (41 mg/kg/day, gavage) or vehicle (CTR group) from the postnatal day (PND) 28 to 50. At PND 51 and 120 the rats were evaluated for: thoracic aorta reactivity to phenylephrine, in the presence (Endo+) or absence of endothelium (Endo-), to acetylcholine and to sodium nitroprusside (SNP), aortic thickness and endothelial nitric oxide synthase (eNOS) expression. In serum were analyzed: the antioxidant capacity by ferric reducing antioxidant power assay; endogenous antioxidant reduced glutathione, and superoxide anion. Results were expressed as mean ± s.e.m., differences when p < 0.05. STATISTICS: Two-way ANOVA (and Tukey's) or Student t-test. KEY FINDINGS: At PND 51, the contraction induced by phenylephrine in Endo+ ring was higher in TPM when compared to CTR. At PND 120, the aortic sensitivity to acetylcholine in TPM rats was reduced in comparison with CTR. The aortic eNOs expression and the aortic thickness were similar between the groups. At PND 51 and 120, TPM group presented a decrease in antioxidants when compared to CTR groups and at PND 120, in TPM group the superoxide anion was increased. SIGNIFICANCE: Taken together, the treatment of rats with TPM during peripubertal period promoted permanent impairment of endothelial function probably mediated by oxidative stress.
Assuntos
Acetilcolina , Antioxidantes , Ratos , Animais , Masculino , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Topiramato/farmacologia , Acetilcolina/metabolismo , Superóxidos/metabolismo , Endotélio Vascular/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Estresse Oxidativo , Aorta Torácica/metabolismo , Fenilefrina/farmacologia , Óxido Nítrico/metabolismoRESUMO
The telocyte (TC) is a new interstitial cell type described in a wide variety of organs and loose connective tissues around small vessels, but its presence in large arteries remains unexplored. TCs have small cell bodies and remarkably thin, long, moniliform processes called telopods (Tps). Using transmission electron microscopy and immunofluorescence, we identified TCs in normal human thoracic aortas and in those with aneurysm or acute dissection (TAAD). In normal aortas the TCs were distributed throughout the connective tissue of the adventitial layer, in its innermost portion and at the zone of transition with the medial layer, with their long axes oriented parallel to the external elastic lamellae, forming a three-dimensional network, without prevalence in the media layer. In contrast, TAAD TCs were present in the medial layer and in regions of neovascularization. The most important feature of the adventitia of diseased aortas was the presence of numerous contacts between TCs and stem cells, including vascular progenitor cells. Although the biologically functional correlations need to be elucidated, the morphological observations presented here provide strong evidence of the involvement of TCs in maintaining vascular homeostasis in pathological situations of tissue injury.