RESUMO
Diabetic kidney disease is associated with the dysbiosis of the gut microbiota and its metabolites. db/db mice were fed chow diet with or without 0.4% resveratrol for 12 weeks, after which the gut microbiota, faecal short-chain fatty acids (SCFAs), and renal fibrosis were analysed. Resveratrol ameliorated the progression of diabetic kidney disease and alleviated tubulointerstitial fibrosis. Further studies showed that gut microbiota dysbiosis was modulated by resveratrol, characterised by the expansion of SCFAs-producing bacteria Faecalibaculum and Lactobacillus, which increased the concentrations of SCFAs (especially acetic acid) in the faeces. Moreover, microbiota transplantation experiments found that alteration of the gut microbiota contributed to the prevention of diabetic kidney disease. Acetate treatment ameliorated proteinuria, glomerulosclerosis and tubulointerstitial fibrosis in db/db mice. Overall, resveratrol improved the progression of diabetic kidney disease by suppressing tubulointerstitial fibrosis, which may be involved, at least in part, in the regulation of the gut microbiota-SCFAs axis.
Assuntos
Nefropatias Diabéticas , Ácidos Graxos Voláteis , Microbioma Gastrointestinal , Resveratrol , Animais , Microbioma Gastrointestinal/efeitos dos fármacos , Ácidos Graxos Voláteis/metabolismo , Nefropatias Diabéticas/tratamento farmacológico , Resveratrol/farmacologia , Camundongos , Masculino , Fibrose , Fezes/microbiologia , Disbiose , Rim/efeitos dos fármacos , Camundongos Endogâmicos C57BLRESUMO
BACKGROUND: Adverse environmental exposure during the prenatal period can lead to diseases in the offspring, including hypertension. Whether or not the hypertensive phenotype can be transgenerationally transmitted is not known. METHODS: Pregnant Sprague Dawley rats were intraperitoneally injected with lipopolysaccharide (LPS) on gestation days 6, 8, 10, and 12 to generate the prenatal LPS exposure model. Blood pressure was monitored by both telemetry and tail-cuff method. RNA sequencing was performed to analyze transcriptome alteration in the kidney of the third generation. Tempol and spironolactone were used to test the potential preventative and therapeutic effect of targeting reactive oxygen species and mineralocorticoid receptor signaling, respectively. Molecular biological experiments were performed to illustrate the mechanism of epigenetic and transcription regulation. RESULTS: Prenatal LPS exposure can impair the ability to excrete a salt load and induce hypertension from the first to the third generations, with the fourth and fifth generations, inducing salt-sensitive hypertension. Compared with control pups, the transcriptome in the kidney of the hypertensive third-generation prenatal LPS-exposed offspring have upregulation of the Ras-related C3 botulinum toxin substrate 1 (Rac1) gene and activation of mineralocorticoid receptor signaling. Furthermore, we found that LPS exposure during pregnancy triggered oxidative stress that upregulated KDM3B (histone lysine demethylase 3B) in the oocytes of first-generation female rats, leading to an inheritable low level of H3K9me2 (histone H3 lysine 9 dimethylation), resulting in the transgenerational upregulation of Rac1. Based on these findings, we treated the LPS-exposed pregnant rats with the reactive oxygen species scavenger, tempol, which successfully prevented hypertension in the first-generation offspring and the transgenerational inheritance of hypertension. CONCLUSIONS: These findings show that adverse prenatal exposure induces transgenerational hypertension through an epigenetic-regulated mechanism and identify potentially preventive and therapeutic strategies for hypertension.
Assuntos
Hipertensão , Efeitos Tardios da Exposição Pré-Natal , Animais , Óxidos N-Cíclicos , Feminino , Histona Desmetilases , Histonas , Hipertensão/induzido quimicamente , Hipertensão/genética , Histona Desmetilases com o Domínio Jumonji , Lipopolissacarídeos/toxicidade , Lisina , Gravidez , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Efeitos Tardios da Exposição Pré-Natal/etiologia , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio , Receptores de Mineralocorticoides/genética , Marcadores de Spin , Espironolactona , Proteínas rac1 de Ligação ao GTP/genéticaRESUMO
Exercise has been recommended as a nonpharmaceutical therapy to treat insulin resistance (IR). Previous studies showed that dopamine D1-like receptor agonists, such as fenoldopam, could improve peripheral insulin sensitivity, while antipsychotics, which are dopamine receptor antagonists, increased susceptibility to Type 2 diabetes mellitus (T2DM). Meanwhile, exercise has been proved to stimulate dopamine receptors. However, whether the dopamine D1 receptor (D1R) is involved in exercise-mediated amelioration of IR remains unclear. We found that the D1-like receptor antagonist, SCH23390, reduced the effect of exercise on lowering blood glucose and insulin in insulin-resistant mice and inhibited the contraction-induced glucose uptake in C2C12 myotubes. Similarly, the opposite was true for the D1-like receptor agonist, fenoldopam. Furthermore, the expression of D1R was decreased in skeletal muscles from streptozotocin (STZ)- and high-fat intake-induced T2DM mice, accompanied by increased D1R phosphorylation, which was reversed by exercise. A screening study showed that G protein-coupled receptor kinase 4 (GRK4) may be the candidate kinase for the regulation of D1R function, because, in addition to the increased GRK4 expression in skeletal muscles of T2DM mice, GRK4 transgenic T2DM mice exhibited lower insulin sensitivity, accompanied by higher D1R phosphorylation than control mice, whereas the AAV9-shGRK4 mice were much more sensitive to insulin than AAV9-null mice. Mechanistically, the up-regulation of GRK4 expression caused by increased reactive oxygen species (ROS) in IR was ascribed to the enhanced expression of c-Myc, a transcriptional factor of GRK4. Taken together, the present study shows that exercise, via regulation of ROS/c-Myc/GRK4 pathway, ameliorates D1R dysfunction and improves insulin sensitivity.
Assuntos
Diabetes Mellitus Tipo 2 , Resistência à Insulina , Animais , Camundongos , Fenoldopam , Insulina , Músculo Esquelético , Espécies Reativas de Oxigênio , Receptores de Dopamina D1/genéticaRESUMO
Activation of the angiotensin II type 2 receptor (AT2R) induces diuresis and natriuresis. Increased expression or/and activity of G-protein-coupled receptor kinase 4 (GRK4) or genetic variants (e.g., GRK4γ142V) cause sodium retention and hypertension. Whether GRK4 plays a role in the regulation of AT2R in the kidney remains unknown. In the present study, we found that spontaneously hypertensive rats (SHRs) had increased AT2R phosphorylation and impaired AT2R-mediated diuretic and natriuretic effects, as compared with normotensive Wistar-Kyoto (WKY) rats. The regulation by GRK4 of renal AT2R phosphorylation and function was studied in human (h) GRK4γ transgenic mice. hGRK4γ142V transgenic mice had increased renal AT2R phosphorylation and impaired AT2R-mediated natriuresis, relative to hGRK4γ wild-type (WT) littermates. These were confirmed in vitro; AT2R phosphorylation was increased and AT2R-mediated inhibition of Na+-K+-ATPase activity was decreased in hGRK4γ142V, relative to hGRK4γ WT-transfected renal proximal tubule (RPT) cells. There was a direct physical interaction between renal GRK4 and AT2R that was increased in SHRs, relative to WKY rats. Ultrasound-targeted microbubble destruction of renal GRK4 decreased the renal AT2R phosphorylation and restored the impaired AT2R-mediated diuresis and natriuresis in SHRs. In vitro studies showed that GRK4 siRNA reduced AT2R phosphorylation and reversed the impaired AT2R-mediated inhibition of Na+-K+-ATPase activity in SHR RPT cells. Our present study shows that GRK4, at least in part, impairs renal AT2R-mediated diuresis and natriuresis by increasing its phosphorylation; inhibition of GRK4 expression and/or activity may be a potential strategy to improve the renal function of AT2R.
Assuntos
Quinase 4 de Receptor Acoplado a Proteína G , Hipertensão , Adenosina Trifosfatases/metabolismo , Angiotensina II/metabolismo , Angiotensina II/farmacologia , Animais , Quinase 4 de Receptor Acoplado a Proteína G/genética , Quinase 4 de Receptor Acoplado a Proteína G/metabolismo , Camundongos , Fosforilação , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor Tipo 1 de Angiotensina/metabolismo , Receptor Tipo 2 de Angiotensina/genética , Receptor Tipo 2 de Angiotensina/metabolismoRESUMO
The endothelin receptor type B (ETBR) regulates water and electrolyte balance and blood pressure, in part, by inhibiting renal sodium transport. Our preliminary study found that the ETBR-mediated diuresis and natriuresis are impaired in hypertension with unknown mechanism. Persistently increased activity of G protein-coupled receptor kinase 4 (GRK4), caused by increased expression or genetic variants (eg, GRKγ142V), impairs the ability of the kidney to excrete a sodium load, in part, by impairing renal dopamine D1 receptor function through persistent phosphorylation. Our present study found that although renal ETBR expression was not different between Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHRs), renal ETBR phosphorylation was higher in SHRs. The role of hyper-phosphorylation in impaired ETBR-function was supported by results in human (h) GRK4γ transgenic mice. Stimulation of ETBR by BQ3020-induced natriuresis in human (h) GRK4γ wild-type (WT) mice. However, in hGRK4γ 142V transgenic mice, the renal ETBR was hyperphosphorylated and ETBR-mediated natriuresis and diuresis were not evident. There were co-localization and co-immunoprecipitation of ETBR and GRK4 in renal proximal tubule (RPT) cells from both WKY and SHRs but was greater in the latter than the former group. SiRNA-mediated downregulation of GRK4 expression, recovered the impaired inhibitory effect of ETBR on Na+ -K+ -ATPase activity in RPT cells from SHR. In vivo downregulation of renal GRK4 expression, via ultrasound-targeted microbubble destruction, decreased ETBR phosphorylation and restored ETBR-mediated natriuresis and diuresis in SHRs. This study provides a mechanism by which GRK4, via regulation of renal ETBR function, participates in the pathogenesis of hypertension.
Assuntos
Quinase 4 de Receptor Acoplado a Proteína G/metabolismo , Hipertensão/metabolismo , Rim/metabolismo , Receptor de Endotelina B/metabolismo , Animais , Células Cultivadas , Feminino , Quinase 4 de Receptor Acoplado a Proteína G/genética , Hipertensão/genética , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos Transgênicos , Fosforilação , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor de Endotelina B/genética , Sódio/metabolismo , Especificidade da EspécieRESUMO
Background: Genetic variants of coding genes related to blood pressure regulation participate in the pathogenesis of hypertension and determines the response to specific antihypertensive drugs. G protein-coupled receptor kinase 4 (GRK4) and its variants are of great importance in pathogenesis of hypertension. However, little is known about role of GRK4 variants in determine circadian rhythm of blood pressure and response to candesartan in hypertension. The aim of this study was to analyze the correlation of GRK4 variants and circadian rhythm of blood pressure, and to explore their effect on antihypertensive efficiency of candestartan.Methods: In this study, a total of 1239 cases were eligible, completed ambulatory blood pressure monitoring (ABPm) observation and exon sequencing of G protein-coupled receptor kinase 4 (GRK4). ABPm was obtained before and after 4-week treatment of candesartan. Diurnal variation of systolic blood pressure and antihypertensive effect of candesartan were then assessed.Results: Compared to GRK4 wild type (GRK4-WT), patients with GRK4 variants were more likely to be non-dippers (odds ratio (OR) 6.672, 95% confidence interval (CI) 5.124-8.688, P < .001), with GRK4 A142V (OR 5.888, 95% CI 4.332-8.003, P < .001), A486V (OR 7.102, 95% CI 5.334-9.455, P < .001) and GRK4 R65L (OR 3.273, 95% CI 2.271-4.718, P < .001), respectively. Correlation analysis revealed that non-dippers rhythm of blood pressure were associated with GRK4 variants (r = .420, P < .001), with GRK4 A142V (r = .416, P < .001), A486V (r = .465, P < .001) and GRK4 R65L (r = .266, P < .001), respectively. When given 4-week candesartan, patients with GRK4 variants showed better antihypertensive effect as to drop in blood pressure (24 h mSBP, 21.21 ± 4.99 vs 12.34 ± 4.78 mmHg, P < .001) and morning peak (MP-SBP, 16.54 ± 4.37 vs 11.52 ± 4.14 mmHg, P < .001), as well as greater increase in trough to peak ratio (SBP-T/P, .71 ± .07 vs .58 ± .07, P < .001) and smoothness index (SBP-SI, 1.44 ± .16 vs 1.17 ± .11, P < .001) than those with GRK4 WT.Conclusion: This study indicates that hypertensive patients with GRK4 variants are more likely to be non-dippers. What's more, patients with GRK4 variants possess a significantly better antihypertensive response to candesartan than those with GRK4 WT.
Assuntos
Benzimidazóis/uso terapêutico , Compostos de Bifenilo/uso terapêutico , Monitorização Ambulatorial da Pressão Arterial , Ritmo Circadiano , Quinase 4 de Receptor Acoplado a Proteína G/genética , Hipertensão , Tetrazóis/uso terapêutico , Anti-Hipertensivos/farmacologia , Anti-Hipertensivos/uso terapêutico , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/genética , Ritmo Circadiano/genética , Variação Genética , Humanos , Hipertensão/tratamento farmacológico , Hipertensão/genéticaRESUMO
Mesenchymal stem cells (MSCs) exert therapeutic effect on treating acute myocardial infarction. Recent evidence showed that paracrine function rather than direct differentiation predominately contributes to the beneficial effects of MSCs, but how the paracrine factors function are not fully elucidated. In the present study, we tested if extracellular vesicles (EVs) secreted by MSC promotes angiogenesis in infracted heart via microRNAs. Immunostaining of CD31 and matrigel plug assay were performed to detect angiogenesis in a mouse myocardial infarction (MI) model. The cardiac function and structure was examined with echocardiographic analysis. Capillary-like tube formation, migration and proliferation of human umbilical vein endothelial cells (HUVECs) were determined. As a result, MSC-EVs significantly improved angiogenesis and cardiac function in post-MI heart. MSC-EVs increased the proliferation, migration and tube formation capacity of HUVECs. MicroRNA (miR)-210 was found to be enriched in MSC-EVs. The EVs collected from MSCs with miR-210 silence largely lost the pro-angiogenic effect both in-vitro and in-vivo. The miR-210 target gene Efna3, which plays a role in angiogenesis, was down-regulated by MSC-EVs treatment in HUVECs. In conclusion, MSC-EVs are sufficient to improve angiogenesis and exert therapeutic effect on MI, its pro- angiogenesis effect might be associated with a miR-210-Efna3 dependent mechanism. This article is part of a Special Issue entitled: Genetic and epigenetic control of heart failure - edited by Jun Ren & Megan Yingmei Zhang.
Assuntos
Micropartículas Derivadas de Células/transplante , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Infarto do Miocárdio/terapia , Neovascularização Fisiológica , Animais , Micropartículas Derivadas de Células/metabolismo , Micropartículas Derivadas de Células/patologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Células-Tronco Mesenquimais/patologia , Camundongos , MicroRNAs/genética , Infarto do Miocárdio/genética , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologiaRESUMO
UNLABELLED: Nox4-based NADPH oxidase is a major reactive oxygen species-generating enzyme in the vasculature, but its role in atherosclerosis remains controversial. OBJECTIVE: Our goal was to investigate the role of smooth muscle Nox4 in atherosclerosis. APPROACH AND RESULTS: Atherosclerosis-prone conditions (disturbed blood flow and Western diet) increased Nox4 mRNA level in smooth muscle of arteries. To address whether upregulated smooth muscle Nox4 under atherosclerosis-prone conditions was directly involved in the development of atherosclerosis, mice carrying a human Nox4 P437H dominant negative mutation (Nox4DN), specifically in smooth muscle, were generated on a FVB/N ApoE deficient genetic background to counter the effect of increased smooth muscle Nox4. Nox4DN significantly decreased aortic stiffness and atherosclerotic lesions, with no effect on blood pressure. Gene analysis indicated that soluble epoxide hydrolase 2 (sEH) was significantly downregulated in Nox4DN smooth muscle cells (SMC), at both mRNA and protein levels. Downregulation of sEH by siRNA decreased SMC proliferation and migration, and suppressed inflammation and macrophage adhesion to SMC. CONCLUSIONS: Downregulation of smooth muscle Nox4 inhibits atherosclerosis by suppressing sEH, which, at least in part, accounts for inhibition of SMC proliferation, migration and inflammation.
Assuntos
Aterosclerose/genética , Inflamação/genética , Miócitos de Músculo Liso/metabolismo , NADPH Oxidases/genética , Animais , Aorta/metabolismo , Aorta/patologia , Apolipoproteínas E/genética , Aterosclerose/patologia , Pressão Sanguínea/genética , Movimento Celular/genética , Proliferação de Células/genética , Regulação da Expressão Gênica , Humanos , Inflamação/patologia , Camundongos , Camundongos Transgênicos , Miócitos de Músculo Liso/patologia , NADPH Oxidase 4 , NADPH Oxidases/biossíntese , RNA Interferente Pequeno/genética , Espécies Reativas de Oxigênio/metabolismoRESUMO
Tirofiban, a glycoprotein IIb/IIIa inhibitor, is an antiplatelet drug extensively used in patients with acute coronary syndrome (ACS) and exerts an therapeutic effect on no-reflow phenomenon during percutaneous coronary intervention (PCI). Previous studies elucidated the vasodilation caused by tirofiban in the peripheral artery. However, whether tirofiban exerts a vasodilator effect on the coronary artery is unclear. Our present study found that tirofiban induced endothelium-dependent vasodilation in a concentration- and time-dependent manner in the isolated rat coronary artery pre-constricted by 5-hydroxytryptamine (5-HT). Further study showed that incubation of human umbilical venous endothelial cells (HUVECs) with tirofiban increased NO production, which was ascribed to the increased eNOS phosphorylation. This was confirmed by the loss of the vasorelaxant effect of tirofiban in the presence of l-NAME (eNOS inhibitor) and L-NMMA (NOS inhibitor) but not SMT (iNOS inhibitor) on isolated rat coronary arteries. The vasorelaxation was also blocked by the PI3K inhibitors, wortmannin and LY294002, as well as the Akt inhibitor SH-5, indicating the role of PI3K and Akt in tirofiban-mediated vasodilation. Moreover, further study showed that soluble guanylyl cyclase (sGC) inhibitor ODQ, or blockers of potassium channel (big-conductance calcium-activated potassium channel) blocked tirofiban-induced vasodilation of the coronary artery. These findings suggest that tirofiban induces vasorelaxation via an endothelium-dependent NO-cGMP signaling through the activation of the Akt/eNOS/sGC pathway.
Assuntos
Vasos Coronários/fisiologia , GMP Cíclico/metabolismo , Endotélio Vascular/metabolismo , Óxido Nítrico/metabolismo , Tirosina/análogos & derivados , Vasodilatação/fisiologia , Animais , Circulação Coronária/efeitos dos fármacos , Circulação Coronária/fisiologia , Vasos Coronários/efeitos dos fármacos , Relação Dose-Resposta a Droga , Endotélio Vascular/efeitos dos fármacos , Técnicas In Vitro , Masculino , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Tirofibana , Tirosina/administração & dosagem , Vasodilatação/efeitos dos fármacos , Vasodilatadores/administração & dosagemRESUMO
Cardiac troponin I (cTnI), a biomarker for myocardial damage and risk stratification, may be involved in the pathogenesis of cardiovascular diseases, which was ascribed to the effect of cTnI auto-antibodies. Whether or not cTnI itself has a direct impact on acute myocardial injury is unknown. To exclude the influence of cTnI antibody on the cardiac infarct size, we studied the effect of cTnI shortly after myocardial ischaemia-reperfusion (I/R) injury when cTnI antibodies were not elevated. Pretreatment with cTnI augmented the myocardial infarct size caused by I/R, accompanied by an increase in inflammatory markers in the blood and myocardium. Additional experiments using human umbilical vein endothelial cells (HUVECs) showed that the detrimental effect of cTnI was related to cTnI-induced increase in vascular cell adhesion molecule-1 (VCAM-1) expression and VCAM-1 mediated adhesion of human monocytes (THP-1) to HUVECs, which could be neutralized by VCAM-1 antibody. Both toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) were involved in the signalling pathway, because blockade of either TLR4 or NF-κB inhibited the cTnI's effect on VCAM-1 expression and adhesion of monocytes to endothelial cells. Moreover, TLR4 inhibition reduced cTnI-augmented cardiac injury in rats with I/R injury. We conclude that cTnI exacerbates myocardial I/R injury by inducing the adhesion of monocytes to vascular endothelial cells via activation of the TLR4/NF-κB pathway. Inhibition of TLR4 may be an alternative strategy to reduce cTnI-induced myocardial I/R injury.
RESUMO
BACKGROUND: Long noncoding RNAs (lncRNAs) have recently been implicated in many biological processes and diseases. Atherosclerosis is a major risk factor for cardiovascular disease. However, the functional role of lncRNAs in atherosclerosis is largely unknown. METHODS AND RESULTS: We identified lincRNA-p21 as a key regulator of cell proliferation and apoptosis during atherosclerosis. The expression of lincRNA-p21 was dramatically downregulated in atherosclerotic plaques of ApoE(-/-) mice, an animal model for atherosclerosis. Through loss- and gain-of-function approaches, we showed that lincRNA-p21 represses cell proliferation and induces apoptosis in vascular smooth muscle cells and mouse mononuclear macrophage cells in vitro. Moreover, we found that inhibition of lincRNA-p21 results in neointimal hyperplasia in vivo in a carotid artery injury model. Genome-wide analysis revealed that lincRNA-p21 inhibition dysregulated many p53 targets. Furthermore, lincRNA-p21, a transcriptional target of p53, feeds back to enhance p53 transcriptional activity, at least in part, via binding to mouse double minute 2 (MDM2), an E3 ubiquitin-protein ligase. The association of lincRNA-p21 and MDM2 releases MDM2 repression of p53, enabling p53 to interact with p300 and to bind to the promoters/enhancers of its target genes. Finally, we show that lincRNA-p21 expression is decreased in patients with coronary artery disease. CONCLUSIONS: Our studies identify lincRNA-p21 as a novel regulator of cell proliferation and apoptosis and suggest that this lncRNA could serve as a therapeutic target to treat atherosclerosis and related cardiovascular disorders.
Assuntos
Apoptose/genética , Macrófagos/citologia , Músculo Liso Vascular/citologia , Neointima/genética , RNA Longo não Codificante/genética , Proteína Supressora de Tumor p53/metabolismo , Animais , Aterosclerose/genética , Aterosclerose/metabolismo , Aterosclerose/patologia , Doenças das Artérias Carótidas/genética , Doenças das Artérias Carótidas/metabolismo , Doenças das Artérias Carótidas/patologia , Linhagem Celular , Proliferação de Células , Modelos Animais de Doenças , Humanos , Macrófagos/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/fisiologia , Neointima/patologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/genéticaRESUMO
BACKGROUND: Both angiotensin II type 1 receptor (AT1R) and nuclear factor-kappa B (NF-κB) play significant roles in the pathogenesis of hypertension and type 2 diabetes. However, the role of NF-κB in perpetuating renal AT1 receptors dysfunction remains unclear. The aim of the present study to determine whether blockade of NF-κB, could reverse the exaggerated renal AT1R function, reduce inflammatory state and oxidative stress, lower blood pressure in Zucker diabetic fatty (ZDF) rats. METHODS: Pyrrolidine dithiocarbamate (PDTC), a NF-κB inhibitor (150 mg/kg in drinking water)or vehicle was administered orally to 12-weeks-old ZDF rats and their respective control lean Zucker (LZ) rats for 4 weeks. Blood pressure was measured weekly by tail-cuff method. AT1R functions were determined by measuring diuretic and natriuretic responses to AT1R antagonist (candesartan; 10 µg/kg/min iv). The mRNA and protein levels of NF-κB, oxidative stress maker and AT1R were determined using quantitative real-time PCR and Western blotting, respectively. The NF-κB-DNA binding activity in renal cortex was measured by Electrophoretic mobility shift assay (EMSA). RESULTS: As compared with LZ rats, ZDF rats had higher blood pressure, impaired natriuresis and diuresis, accompanied with higher levels of oxidative stress and inflammation. Furthermore, AT1R expression was higher in renal cortex from ZDF rats; candesartan induced natriresis and diuresis, which was augmented in ZDF rats. Treatment with PDTC lowered blood pressure and improved diuretic and natriuretic effects in ZDF rats; meanwhile, the increased oxidative stress and inflammation were reduced; the increased AT1R expression and augmented candesartan-mediated natriuresis and diuresis were recoverd in ZDF rats. Our further study investigated the mechanisms of PDTC on AT1R receptor expression. It resulted that PDTC inhibited NF-κB translocation from cytosol to nucleus, inhibited binding of NF-κB with AT1R promoter, therefore, reduced AT1R expression and function. CONCLUSIONS: Our present study indicates blockade of NF-κB, via inhibition of binding of NF-κB with AT1R promoter, reduces renal AT1R expression and function, improves oxidative stress and inflammatory/anti-inflammatory balance, therefore, lowers blood pressure and recovers renal function in ZDF rats.
Assuntos
Antioxidantes/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Rim/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Estresse Oxidativo/efeitos dos fármacos , Pirrolidinas/farmacologia , Receptor Tipo 1 de Angiotensina/efeitos dos fármacos , Tiocarbamatos/farmacologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Benzimidazóis/farmacologia , Compostos de Bifenilo , Diabetes Mellitus Tipo 2 , Diurese/efeitos dos fármacos , Rim/metabolismo , Masculino , NF-kappa B/genética , NF-kappa B/metabolismo , Natriurese/efeitos dos fármacos , Regiões Promotoras Genéticas , Ratos , Ratos Zucker , Reação em Cadeia da Polimerase em Tempo Real , Receptor Tipo 1 de Angiotensina/metabolismo , Tetrazóis/farmacologiaRESUMO
We set out to investigate whether and how SRY (sex-determining region, Y) DNAs in plasma EVs (extracellular vesicles) is involved in the pathogenesis of atherosclerosis. PCR and gene sequencing found the SRY gene fragment in plasma EVs from male, but not female, patients; EVs from male patients with CAD (coronary artery disease) had a higher SRY GCN (gene copy number) than healthy subjects. Additional studies found that leucocytes, the major source of plasma EVs, had higher SRY GCN and mRNA and protein expression in male CAD patients than controls. After incubation with EVs from SRY-transfected HEK (human embryonic kidney)-293 cells, monocytes (THP-1) and HUVECs (human umbilical vein endothelial cells), which do not endogenously express SRY protein, were found to express newly synthesized SRY protein. This resulted in an increase in the adherence factors CD11-a in THP-1 cells and ICAM-1 (intercellular adhesion molecule 1) in HUVECs. EMSA showed that SRY protein increased the promoter activity of CD11-a in THP-1 cells and ICAM-1 in HUVECs. There was an increase in THP-1 cells adherent to HUVECs after incubation with SRY-EVs. SRY DNAs transferred from EVs have pathophysiological significance in vivo; injection of SRY EVs into ApoE-/- (apolipoprotein-knockout) mice accelerated atherosclerosis. The SRY gene in plasma EVs transferred to vascular endothelial cells may play an important role in the pathogenesis of atherosclerosis; this mechanism provides a new approach to the understanding of inheritable CAD in men.
Assuntos
Aterosclerose/genética , Aterosclerose/metabolismo , Células Endoteliais/metabolismo , Leucócitos/metabolismo , Proteína da Região Y Determinante do Sexo/metabolismo , Animais , Adesão Celular , Células Cultivadas , Células Endoteliais/citologia , Endotélio Vascular/metabolismo , Espaço Extracelular/metabolismo , Humanos , Leucócitos/citologia , Masculino , Camundongos , Proteína da Região Y Determinante do Sexo/genética , Molécula 1 de Adesão de Célula Vascular/biossínteseRESUMO
Angiotensin type 1a receptor (AT1aR) is thought to play an important role in the development of hypertension. However, it is unknown how the AT1aR expression in vascular tissue is changed during the development of hypertension or if the degree of methylation in the AT1aR promoter correlates with the expression of AT1aR. To address these questions, we measured AT1aR mRNA, protein expression, and methylation status of the AT1aR promoter in the aorta and mesenteric artery of male spontaneously hypertensive rats (SHRs) and age-matched Wistar-Kyoto (WKY) rats acting as controls at pre-hypertensive (4 weeks), evolving (10 weeks), and established (20 weeks) stages of hypertension. The expression of the AT1aR mRNA and protein was not different between the SHRs and WKY rats at 4 weeks. However, they were significantly greater in SHRs than in WKY rats at 20 weeks. Bisulfite sequencing revealed that the AT1aR promoter from the aorta and mesenteric artery of the SHRs was progressively hypo-methylated with age as compared with their WKY rat counterparts. These results suggest that the heightened AT1aR expression in SHRs is related to the AT1aR promoter hypo-methylation, which might be a consequence of the increased blood pressure and may be important in the maintenance of high blood pressure.
Assuntos
Hipertensão/genética , Receptor Tipo 1 de Angiotensina/genética , Animais , Aorta/metabolismo , Ilhas de CpG , Metilação de DNA , Epigênese Genética , Expressão Gênica , Hipertensão/metabolismo , Masculino , Artérias Mesentéricas/metabolismo , Regiões Promotoras Genéticas , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor Tipo 1 de Angiotensina/metabolismoRESUMO
RATIONALE: Transplantation of stem cells into damaged hearts has had modest success as a treatment for ischemic heart disease. One of the limitations is the poor stem cell survival in the diseased microenvironment. Prolyl hydroxylase domain protein 2 (PHD2) is a cellular oxygen sensor that regulates 2 key transcription factors involved in cell survival and inflammation: hypoxia-inducible factor and nuclear factor-κB. OBJECTIVE: We studied whether and how PHD2 silencing in human adipose-derived stem cells (ADSCs) enhances their cardioprotective effects after transplantation into infarcted hearts. METHODS AND RESULTS: ADSCs were transduced with lentiviral short hairpin RNA against prolyl hydroxylase domain protein 2 (shPHD2) to silence PHD2. ADSCs, with or without shPHD2, were transplanted after myocardial infarction in mice. ADSCs reduced cardiomyocyte apoptosis, fibrosis, and infarct size and improved cardiac function. shPHD2-ADSCs exerted significantly more protection. PHD2 silencing induced greater ADSC survival, which was abolished by short hairpin RNA against hypoxia-inducible factor-1α. Conditioned medium from shPHD2-ADSCs decreased cardiomyocyte apoptosis. Insulin-like growth factor-1 (IGF-1) levels were significantly higher in the conditioned medium of shPHD2-ADSCs versus ADSCs, and depletion of IGF-1 attenuated the cardioprotective effects of shPHD2-ADSC-conditioned medium. Nuclear factor-κB activation was induced by shPHD2 to induce IGF-1 secretion via binding to IGF-1 gene promoter. CONCLUSIONS: PHD2 silencing promotes ADSCs survival in infarcted hearts and enhances their paracrine function to protect cardiomyocytes. The prosurvival effect of shPHD2 on ADSCs is hypoxia-inducible factor-1α dependent, and the enhanced paracrine function of shPHD2-ADSCs is associated with nuclear factor-κB-mediated IGF-1 upregulation. PHD2 silencing in stem cells may be a novel strategy for enhancing the effectiveness of stem cell therapy after myocardial infarction.
Assuntos
Tecido Adiposo/metabolismo , Inativação Gênica/fisiologia , Infarto do Miocárdio/genética , Infarto do Miocárdio/cirurgia , Comunicação Parácrina/genética , Pró-Colágeno-Prolina Dioxigenase/biossíntese , Transplante de Células-Tronco , Tecido Adiposo/citologia , Animais , Sobrevivência Celular/genética , Células Cultivadas , Humanos , Prolina Dioxigenases do Fator Induzível por Hipóxia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/enzimologia , Pró-Colágeno-Prolina Dioxigenase/genética , RNA Interferente Pequeno/biossíntese , RNA Interferente Pequeno/genética , Transplante de Células-Tronco/métodosRESUMO
OBJECTIVE: The abnormal migration of vascular smooth muscle cells (VSMCs) has been implicated to contribute to lesion formation in the adult vasculature. The renin-angiotensin-aldosterone system (RAAS) is intensively involved in the pathogenesis of a variety of cardiovascular diseases. There are increasing pieces of evidence for interactions between RAAS and dopamine receptors. We hypothesize that the D3 receptor has an inhibitory effect on angiotensin II (Ang II)/aldosterone-induced VSMC migration. METHOD: In this study, embryonic thoracic aortic smooth muscle cells were cultured. VSMC migration was determined by the Boyden chamber and wound healing assays. RESULTS: VSMC migration was increased by Ang II (10(-10)-10(-7) mol/L) in a concentration-dependent manner, but not by aldosterone (10(-10)-10(-7) mol/L), and a synergistic effect of Ang II (10(-10) mol/L)/aldosterone (10(-10)mol/L) was also observed in VSMC migration. The migratory effects of Ang II alone/with aldosterone were attenuated by the activation of D3 receptors (10(-10)-10(-7) mol/L), although a D3 receptor agonist, PD128907, by itself, had no effect on VSMC migration. The inhibitory effect of the D3 receptor on Ang II/ aldosterone-mediated VSMC migration was blocked by the blocker of PKA (14-22 amide, 10(-7) mol/L), indicating that PKA was involved in the signaling pathway. CONCLUSION: These results indicate that activation of vascular D3 receptors inhibits Ang II/aldosterone-induced VSMC migration through the PKA signal pathway, which may be important in the regulation of vascular remodeling.
Assuntos
Aldosterona/farmacologia , Angiotensina II/farmacologia , Movimento Celular/efeitos dos fármacos , Hipertensão/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/patologia , Receptores de Dopamina D3/metabolismo , Aorta Torácica/patologia , Células Cultivadas , Sinergismo Farmacológico , Humanos , Hipertensão/tratamento farmacológico , Hipertensão/patologia , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
The dopaminergic and sympathetic systems interact to regulate blood pressure. Our previous studies showed regulation of α1-adrenergic receptor function by D1-like dopamine receptors in vascular smooth muscle cells. Because renalase could regulate circulating epinephrine levels and dopamine production in renal proximal tubules (RPTs), we tested the hypothesis that D1-like receptors regulate renalase expression in kidney. The effect of D1-like receptor stimulation on renalase expression and function was measured in immortalized RPT cells from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHRs). We found that the D1-like receptor agonist fenoldopam (10(-7)-10(-5) mol/l) increased renalase protein expression and function in WKY RPT cells but decreased them in SHR cells. Fenoldopam also increased renalase mRNA levels in WKY but not in SHR cells. In contrast, fenoldopam increased the degradation of renalase protein in SHR cells but not in WKY cells. The regulation of renalase by the D1-like receptor was mainly via the D5 receptor because silencing of the D5 but not D1 receptor by antisense oligonucleotides blocked the stimulatory effect of the D1-like receptor on renalase expression in WKY cells. Moreover, inhibition of PKC, by the PKC inhibitor 19-31, blocked the stimulatory effect of fenoldopam on renalase expression while stimulation of PKC, by a PKC agonist (PMA), increased renalase expression, indicating that PKC is involved in the process. Our studies suggest that the D5 receptor positively regulates renalase expression in WKY but not SHR RPT cells; aberrant regulation of renalase by the D5 receptor may be involved in the pathogenesis of hypertension.
Assuntos
Túbulos Renais Proximais/efeitos dos fármacos , Monoaminoxidase/biossíntese , Receptores de Dopamina D5/fisiologia , Animais , Células Cultivadas , Fenoldopam/farmacologia , Túbulos Renais Proximais/metabolismo , Masculino , Proteína Quinase C/fisiologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptores de Dopamina D1/agonistasRESUMO
Vascular smooth muscle cells (VSMCs) proliferation and migration, which are central in the development of vascular diseases, are regulated by numerous hormones and humoral factors. Activation of the insulin receptor stimulates VSMCs proliferation while dopamine receptors, via D1 and D3 receptors, inhibit the stimulatory effects of norepinephrine on VSMCs proliferation. We hypothesize that activation of the D4 dopamine receptor may also inhibit the proliferation and migration of VSMCs, therefore, inhibit atherosclerosis. Our current study found that insulin increased the proliferation and migration of A10 cells, an effect that was reduced in the presence of a D4 receptor agonist, PD168077. The negative effect of the D4 receptor on insulin's action may be via decreasing insulin receptor expression, because activation of the D4 receptor inhibited insulin receptor protein and mRNA expressions, indicating that the regulation occured at the transcriptional or post-transcriptional levels. To determine whether or not the inhibition of D4 receptor on insulin-mediated proliferation and migration of VSMCs has physiological significance, hyper-insulinemic Sprague-Dawley rats with balloon-injured carotid artery were treated with a D4 agonist, PD168077, (6 mg/kg/d) for 14 days. We found that PD168077 significantly inhibited neointimal formation by inhibition of VSMC proliferation. This study suggests that activation of the D4 receptor suppresses the proliferation and migration of VSMCs, therefore, inhibit atherosclerosis. The D4 receptor may be a potential therapeutic target to reduce the effects of insulin on artery remodeling.
Assuntos
Movimento Celular/fisiologia , Proliferação de Células , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Receptor de Insulina/antagonistas & inibidores , Receptores de Dopamina D4/fisiologia , Animais , Benzamidas/farmacologia , Masculino , Músculo Liso Vascular/efeitos dos fármacos , Piperazinas/farmacologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Receptor de Insulina/biossíntese , Receptores de Dopamina D4/agonistasRESUMO
OBJECTIVE: To evaluate the efficacy and safety of Huashi Baidu Granules (HSBD) in treating patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant. METHODS: A single-center retrospective cohort study was conducted during COVID-19 Omicron epidemic in the Mobile Cabin Hospital of Shanghai New International Expo Center from April 1st to May 23rd, 2022. All COVID-19 patients with asymptomatic or mild infection were assigned to the treatment group (HSBD users) and the control group (non-HSBD users). After propensity score matching in a 1:1 ratio, 496 HSBD users of treatment group were matched by propensity score to 496 non-HSBD users. Patients in the treatment group were administrated HSBD (5 g/bag) orally for 1 bag twice a day for 7 consecutive days. Patients in the control group received standard care and routine treatment. The primary outcomes were the negative conversion time of nucleic acid and negative conversion rate at day 7. Secondary outcomes included the hospitalized days, the time of the first nucleic acid negative conversion, and new-onset symptoms in asymptomatic patients. Adverse events (AEs) that occurred during the study were recorded. Further subgroup analysis was conducted in vaccinated (378 HSBD users and 390 non-HSBD users) and unvaccinated patients (118 HSBD users and 106 non-HSBD users). RESULTS: The median negative conversion time of nucleic acid in the treatment group was significantly shortened than the control group [3 days (IQR: 2-5 days) vs. 5 days (IQR: 4-6 days); P<0.01]. The negative conversion rate of nucleic acid in the treatment group were significantly higher than those in the control group at day 7 (91.73% vs. 86.90%, P=0.014). Compared with the control group, the hospitalized days in the treatment group were significantly reduced [10 days (IQR: 8-11 days) vs. 11 days (IQR: 10.25-12 days); P<0.01]. The time of the first nucleic acid negative conversion had significant differences between the treatment and control groups [3 days (IQR: 2-4 days) vs. 5 days (IQR: 4-6 days); P<0.01]. The incidence of new-onset symptoms including cough, pharyngalgia, expectoration and fever in the treatment group were lower than the control group (P<0.05 or P<0.01). In the vaccinated patients, the median negative conversion time and hospitalized days were significantly shorter than the control group after HSDB treatment [3 days (IQR: 2-5 days) vs. 5 days (IQR: 4-6 days), P<0.01; 10 days (IQR: 8-11 days) vs. 11 days (IQR: 10-12 days), P<0.01]. In the unvaccinated patients, HSBD treatment efficiently shorten the median negative conversion time and hospitalized days [4 days (IQR: 2-6 days) vs. 5 days (IQR: 4-7 days), P<0.01; 10.5 days (IQR: 8.75-11 days) vs. 11.0 days (IQR: 10.75-13 days); P<0.01]. No serious AEs were reported during the study. CONCLUSION: HSBD treatment significantly shortened the negative conversion time of nuclear acid, the length of hospitalization, and the time of the first nucleic acid negative conversion in patients infected with SARS-COV-2 Omicron variant (Trial registry No. ChiCTR2200060472).
Assuntos
COVID-19 , Medicamentos de Ervas Chinesas , Ácidos Nucleicos , Humanos , SARS-CoV-2 , Estudos Retrospectivos , ChinaRESUMO
Importance: The global COVID-19 pandemic does not appear to end in the near future. Currently, limited data are available on the risk factors for delayed viral clearance in patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant infection. Objective: This study aimed to investigate the association of clinical characteristics and vaccination with prolonged viral clearance. Methods: This retrospective cohort included 16,985 patients who had contracted the SARS-CoV-2 Omicron variant between April 5 and May 30, 2022, in Shanghai, China, and had mild or no symptoms. The patients were admitted to the quarantine venue at the Shanghai New International Expo Center. Results: Of the 16,985 participants, the occurrence of viral clearance was ≤8 and > 8 days in 11,009 (64.8 %) and 5976 (35.2 %) participants, respectively. Risk factors related to patients who remained persistently polymerase chain reaction (PCR)-positive were sex (Male, odds ratio [OR] 1.221, p < 0.001), older age (35-49, OR 1.389, p < 0.001; 50-64, OR 1.659, p < 0.001; ≥65, OR 2.139, p < 0.001), presence of symptoms (OR 1.093, p = 0.030), number of vaccinations (two doses, OR 0.753, p < 0.001; three doses, OR 0.797, p < 0.001; four doses, OR 0.543, p < 0.001), and cycle threshold (Ct) value for ORF1ab gene at diagnosis (25-35, OR 0.235, p < 0.001; >35, OR 0.079, p < 0.001). The lower rates of increase in Ct values were observed in the later viral shedding group than in the early viral shedding group for ORF1ab (ß = -0.791, p < 0.001) and N genes (ß = -0.825, p < 0.001). Conclusion: Prolonged SARS-CoV-2 RNA detection and higher viral concentrations were associated with factors such as male sex, older age, symptomatic status, and fewer doses of vaccination in patients admitted to Shanghai Makeshift Hospital between April 5 and May 30, 2022.