RESUMO
Research has shown that some circular RNAs (circRNA) are abnormally expressed in the process of myocardial fibrosis, but the mechanism behind this was unknown. In the process of inducing cardiac fibroblast (CF) activation with TGF-ß1 or Ang II, the expression of circRNA circ_BMP2K and miR-455-3p were significantly inhibited, whereas the expression of SUMO1 was promoted. The results from our dual luciferase reporter gene assays, RIP assays, and pull-down assays show that miR-455-3p directly binds circ_BMP2K, thereby mutually promoting their expression levels. SUMO1 is a target gene of miR-455-3p, and circ_BMP2K enhances the inhibitory effects of miR-455-3p on the expression of SUMO1. Further study showed that both circ_BMP2K and miR-455-3p inhibited the expression of alpha-SMA as well as type I and type III collagen, whereas SUMO1 promoted their expression, and miR-455-3p inhibitors or overexpression of SUMO1 reversed the effects of circ_BMP2K and miR-455-3p. Circ_BMP2K and miR-455-3p inhibits cell proliferation and migration and promotes the apoptosis of CFs, but SUMO1 has the opposite effects; miR-455-3p inhibitors or overexpression of SUMO1 reverses the effects of circ_BMP2K and miR-455-3p. Thus, circ_BMP2K promotes the expression of miR-455-3p, down-regulates the expression of SUMO1, and finally, inhibits the activation, growth, and migration of CFs. These results could provide important therapeutic targets and a theoretical basis for regulating the process of myocardial fibrosis.
Assuntos
Fibroblastos/metabolismo , MicroRNAs/metabolismo , RNA Circular/metabolismo , Proteína SUMO-1/metabolismo , Movimento Celular , Proliferação de Células , Células Cultivadas , Fibroblastos/patologia , Humanos , MicroRNAs/genética , RNA Circular/genética , Proteína SUMO-1/genéticaRESUMO
BACKGROUND/AIMS: To detect the changes of high density lipoprotein (HDL) and its subtypes in serum of patients with coronary heart disease (CHD). METHODS: 337 hospitalized patients were selected from our hospital during August, 2014 - January, 2015, and divided into CHD group (n = 190) and control group (n = 127). Lipoprint lipoprotein analyzer was used to classify low density lipoprotein (LDL) particle size and its sub-components, as well as HDL particle size and its sub-components. The changes of the subtypes in patients with CHD were statistically analyzed. The possible mechanism was explored. RESULTS: (1) Compared with the control group, the concentration of HDL in CHD patients reduced, HDLL significantly decreased (P < 0.001), while HDLS increased (P < 0.001); (2) In the patients with HDL less than 1.04 mmol/L among CHD, all HDL subtypes reduced, but HDLL had the most significant decreased; (3) HDL and all HDL subtypes were positively correlated with apolipoprotein A-I (apoA-I), of which, HDLL had the biggest correlation with apoA-I (P < 0.001); (4) HDL subtypes had good correlation with HDL, of which, HDLM had a maximum correlation with HDL (P < 0.001). CONCLUSION: HDL maturation disorders existed in the serum of CHD patients, HDLL may be protected factor for CHD, whose decrease was closely related wit the risk increase of CHD. The cardiovascular protection function of HDLL may be related with apoA-I content.
Assuntos
Doença da Artéria Coronariana/patologia , Idoso , Apolipoproteína A-I/sangue , Índice de Massa Corporal , Estudos de Casos e Controles , Doença da Artéria Coronariana/sangue , Feminino , Humanos , Lipoproteínas HDL/sangue , Lipoproteínas LDL/sangue , Masculino , Pessoa de Meia-Idade , Tamanho da Partícula , Triglicerídeos/sangueRESUMO
Angiotensin (Ang)-(1-7), a metabolite of AngI and AngII, is a counter-regulatory mediator of AngII. In the present study, we investigated the effects of Ang-(1-7) on AngII-induced apoptosis in human umbilical vein endothelial cells (HUVEC). To this end, HUVEC were pretreated with 10(-9), 10(-8), 10(-7) or 10(-6) mol/L Ang-(1-7) at for 30 min before being stimulated with 10(-6) mol/L Ang-II for another 24 h. Acridine orange/ethidium bromide and propidium iodide staining were used to analyse the effects of Ang-(1-7) on AngII-induced apoptosis. Alone, 10(-6) mol/L Ang-(1-7) had no effect on the apoptosis of HUVEC following exposure of cells for 30 min, whereas AngII (10(-6) mol/L, 24 h) significantly enhanced the number of apoptotic cells (P < 0.01). The AngII-induced apoptosis of HUVEC was suppressed by 10(-9)-10(-6) mol/L Ang-(1-7). The anti-apoptotic effects of Ang-(1-7) were almost completely abolished by A-779 (10(-6) mol/L, 30 min), a specific Mas receptor antagonist. In addition, Ang-(1-7) inhibited AngII-induced accumulation of cleaved caspase 3 and enhanced the expression of the anti-apoptotic factor Bcl-2 at both the mRNA and protein levels. Angiotensin II upregulated the expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1), which is involved in endothelial apoptosis, at both the mRNA and protein levels. This effect was blocked by Ang-(1-7) in a concentration-dependent manner, although A-779 almost completely reversed Ang-(1-7)-mediated inhibition of AngII-induced upregulation of LOX-1. Silencing of LOX-1 using short interference RNA enhanced the protective effects of Ang-(1-7) against AngII-induced apoptosis in HUVEC. Together, the results suggest that Ang-(1-7) ameliorates AngII-induced apoptosis of HUVEC at least in part by suppressing LOX-1 expression.
Assuntos
Angiotensina II/farmacologia , Angiotensina I/farmacologia , Apoptose/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Receptores Depuradores Classe E/metabolismo , Western Blotting , Caspase 3/metabolismo , Técnicas de Cultura de Células , Relação Dose-Resposta a Droga , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Citometria de Fluxo , Células Endoteliais da Veia Umbilical Humana , Humanos , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Receptores Depuradores Classe E/genética , Regulação para CimaRESUMO
The aim of the present study was to investigate the effects of adiponectin (APN) on the expression of T-cadherin in cultured Sprague-Dawley (SD) rat cardiomyocytes injured by hypoxia/reoxygenation (H/R). Primary myocardial cells from neonatal rats were obtained by enzymatic digestion. The cells were divided into control group, H/R group and H/R+APN (3, 10, 20 and 30 µg/mL) groups. The H/R group was incubated in anoxic environment (anoxic solution saturated with high concentration N2) for 3 h, and then in the reoxygenation environment (the reoxygenation solution saturated with pure oxygen) for 1 h. The H/R+APN group was pretreated with different concentrations of APN for 24 h prior to the initiation of H/R. The content of lactate dehydrogenase (LDH) was measured by chemistry chromatometry. Cellular apoptosis was analyzed by flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The expression of T-cadherin was detected by RT-PCR and Western blotting. The results showed that, compared with control group, the apoptotic rate and release of LDH were significantly increased in the H/R group, whereas the expressions of T-cad mRNA and protein were decreased. Pretreating with APN significantly and dose-dependently decreased apoptotic rate and LDH release, and up-regulated T-cad mRNA and protein level in rat neonatal cardiomyocytes under H/R conditions. These results suggest that APN may protect cardiomyocytes against H/R-induced injury by up-regulating H/R-decreased T-cad expression.
Assuntos
Adiponectina/farmacologia , Caderinas/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Apoptose , Hipóxia Celular , L-Lactato Desidrogenase/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Oxigênio/efeitos adversos , Ratos , Ratos Sprague-Dawley , Regulação para CimaRESUMO
OBJECTIVE: To investigate the effects of p53 upregulated modulator of apoptosis (PUMA) on the apoptosis of H9C2 cardiomyocytes induced by high glucose and its mechanisms. METHODS: H9C2 cardiomyocytes were treated with 5.5mmol/L (control group) or 35 mmol/L glucose (HG group) for 6 h, 12 h, 24 h or 48 h respectively to induce apoptosis, each group sets 5 multiple wells. Apoptosis was tested by TUNEL assay. PUMA mRNA was measured by RT-PCR and protein expression was measured by Western blot assay. The mitochondrial membrane potential was detected by JC-1 method. The expressions of cleaved caspase-3 and cytochrome C (Cyt C) protein in mitochondria and cytoplasm were determined by Western blot assay. H9C2 cardiomyocytes were randomly divided into four groups, control group (5.5 mmol/L), HG (35 mmol/L) group, HG+si-scramble group(si-scramble treatment for 24 h, then 35 mmol/L high glucose treatment for 24 h) and HG-si-PUMA group (si-PUMA treatment for 24 h, then 35mmol/L high glucose treatment for 24 h). Si-PUMA was transfected into cardiomyocytes and the effects of PUMA on high glucose-induced apoptosis were studied. RESULTS: Compared with the control group, high glucose increased cardiomyocyte apoptosis and enhanced PUMA mRNA and protein expressions significantly (Pï¼0.05 or Pï¼0.01). Cell injury and increased PUMA expression were time-dependent and there was no significant difference between the high glucose 24 h group and the high glucose 48h group. The following experiment used high glucose 24 h as the stimulation time. The cardiomyocytes transfected with si-PUMA to inhibit PUMA expression had decreased apoptotic rate and cleaved caspase-3, increased mitochondria membrane potential and decreased Cyt C release (Pï¼0.05 or Pï¼0.01). There were no significant differences between the HG+si-scramble group and the high glucose group (Pï¼0.05). CONCLUSION: PUMA mediates high glucose-induced cardiomyocyte apoptosis suggesting PUMA may be an important target gene of diabetic cardiomyopathy.
Assuntos
Apoptose , Miócitos Cardíacos , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Caspase 3/metabolismo , Glucose/efeitos adversos , Glucose/metabolismo , Miócitos Cardíacos/metabolismo , RNA Mensageiro/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , RatosRESUMO
The present study was to investigate the effect of glucagon like peptide-1 (GLP-1) on high glucose-induced oxidative stress of cardiomyocytes and the possible role of the PI3K-Akt signal path in this process in the neonatal SD rats. With enzymatic digestion and immunofluorescence identification, cardiomyocytes after 72-96 h of primary culture were used in experiment. The cells were divided into 5 groups: normal control group, high glucose group, high glucose + GLP-1 group, high glucose + GLP-1 + LY294002 group and high osmolarity control group. The content of MDA was detected by TBA colouration method. The content of SOD was detected by xanthine oxidase method. The change of NADPH P47phox subunit mRNA quantity was detected by PCR gel electrophoresis. The level of ROS was detected by flow cytometry, and was also observed by fluorescence microscope. The DNA ladder was examined by agarose gel electrophoresis, and the cell apoptosis was determined by Annexin-V-FITC/PI flow cytometry, and the phosphorylation of Akt was determined by Western blotting. Compared with those in the normal control group, in the high glucose group, the cells grew poorly, and the beating rate was significantly lower (P < 0.05); The apoptotic rate was significantly increased (P < 0.05); The MDA content was increased (P < 0.05); It showed the typical DNA ladder, which is the characteristic of apoptosis; The SOD activity was decreased (P < 0.05); The level of intracellular ROS increased (P < 0.05); And the expression of NADPH P47phox subunit mRNA was increased; However the phosphorylation level of Akt was decreased. Pretreatment with GLP-1 improved the above-mentioned parameters and decreased the expression of NADPH P47phox subunit mRNA (P < 0.05). However, compared with the high glucose + GLP-1 group, LY294002, an inhibitor of PI3K-Akt signal path, attenuated the protective effect of GLP-1 in the high glucose + GLP-1 + LY294002 group. It is suggested that GLP-1 plays a protective role in the high glucose-induced injury and apoptosis of cardiomyocytes, and the PI3K-Akt signal path is involved in this process.
Assuntos
Apoptose/efeitos dos fármacos , Peptídeo 1 Semelhante ao Glucagon/farmacologia , Glucose/farmacologia , Miócitos Cardíacos/citologia , Estresse Oxidativo/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Células Cultivadas , Feminino , Masculino , Miócitos Cardíacos/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Substâncias Protetoras/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Sprague-Dawley , Transdução de SinaisRESUMO
The aim of the present study is to investigate the effects of adiponectin (APN) on hypoxia/reoxygenation (H/R) injury in cultured cardiomyocytes. Primary cardiomyocytes were obtained from neonatal rats by enzymatic digestion method and identified by immunofluorescent technique. Primary cells cultured for 72 h were used in experiment and divided into 5 groups randomly: Control group, H/R group, H/R+APN group, H/R+APN+adenine 9-beta-D-arabinfuranoside (AraA, AMPK inhibitor) group, and H/R+AraA group. The cardiocyte morphology and beating rate were observed under inverted microscope. The DNA ladder was examined by agarose gel electrophoresis, and the cell apoptosis was determined by flow cytometry. Moreover, the malondialchehyche (MDA) content in myocardial cells and the superoxide dismutase (SOD) activity in the supernatant were measured using kits, the fluorescence intensity of intracellular Ca2+ was observed by laser scanning confocal microscope, and the phosphorylation of AMPK was determined by Western blotting. Compared with control group, H/R group showed increased apoptotic rate, oxidative stress level, intracellular Ca2+ concentration and phosphorylation level of AMPK (P<0.05), while significant ameliorations in the above indices were seen in H/R+APN group. On the contrast, AraA attenuated the protective effect of APN and decreased the phosphorylation of AMPK. These results suggest that adiponectin can protect cardiomyocytes from H/R-induced oxidative stress and apoptosis through AMPK pathway.
Assuntos
Adiponectina/farmacologia , Apoptose/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/citologia , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Animais Recém-Nascidos , Cálcio/metabolismo , Cardiotônicos/farmacologia , Hipóxia Celular , Células Cultivadas , Feminino , Masculino , Estresse Oxidativo , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
OBJECTIVE: To investigate the effects of RNA interference (RNAi) targeting angiotensin II Type 1 receptor (ATlR) and angiotensin-converting enzyme (ACE) on blood pressure and myocardial remodeling in spontaneous hypertensive rats (SHRs). METHODS: Saline (control), adenovirus (Ad5) and recombinant adenoviral vectors (Ad5-ACE-shRNA, Ad5-AT1R-shRNA and Ad5-ACE-AT1R-shRNA expressing ACE, AT1R, ACE and AT1R gene-specific shRNA, respectively) were randomly administered by caudal intravasation to SHRs (n = 12 each group) at day 1 and 17. Normotensive Wistar-Kyoto rats (WKY) served as normal controls. Systolic blood pressure (SBP) of the caudal artery was measured daily. Expression of ACE and AT1R at mRNA levels in ventricle and aorta were evaluated by fluorescence quantitative PCR. Angiotension II serum concentration was measured by ELISA at day 3 (n = 6 each group). The ratio of left ventricular to body weight (LVW/BW) and myocardial collagen content were measured, myocardial ultrastructure observed under transmission electron microscope at the study end. RESULTS: The caudal artery pressure of saline and Ad5 group was equally increased by about 26 mm Hg(1 mm Hg = 0.133 kPa) compared to baseline (both P < 0.05). Ad5-ACE-shRNA, Ad5-AT1R-shRNA and Ad5-ACE-AT1R-shRNA injection significantly reduced SBP (-24 mm Hg, -22 mm Hg and -26 mm Hg respectively, all P < 0.05 vs. baseline) and the antihypertensive effect could last at least 15 days post each injection. SBP was not affected by saline and Ad5 injections. ACE and AT1 mRNA expressions at ventricle and aorta were significantly decreased in Ad5-ACE-shRNA, Ad5-ACE-AT1R-shRNA and Ad5-AT1R-shRNA, Ad5-ACE-AT1R-shRNA treated SHRs compared to those in saline and Ad5 groups (all P < 0.05) and was comparable to that in WKY group (P > 0.05). The LVW/BW ratio [(2.22 +/- 0.18) microg/mg, (2.23 +/- 0.19) microg/mg, (2.17 +/- 0.16) microg/mg] and myocardial collagen content [(1.291 +/- 0.019) microg/mg, (1.298 +/- 0.019) microg/mg, (1.276 +/- 0.019) microg/mg] in Ad5-ACE-shRNA, Ad5-AT1R-shRNA and Ad5-ACE-AT1R-shRNA treated SHRs were also significantly lower than those in saline treated [(3.23 +/- 0.13) microg/mg and(1.683 +/- 0.013) microg/mg, both P < 0.05] and Ad5 treated SHRs [(3.25 +/- 0.12) microg/mg and(1.693 +/- 0.013) microg/mg, both P < 0.05], but still higher than those of WKY group [(2.06 +/- 0.12) microg/mg and (1.258 +/- 0.019) microg/mg, both P < 0.05]. Myocardial ultrastructure was also significantly improved in all SHRs underwent RNAi treatments compared to saline and Ad5 treated SHRs. CONCLUSION: RNAi targeting ACE and AT1R gene significantly inhibited myocardial and aortic ACE and AT1R mRNA expressions and resulted in prolonged antihypertensive effects and myocardial ultrastructure improvements in SHRsl. The RNAi technology may be a potential new strategy of gene therapy for hypertension.
Assuntos
Pressão Sanguínea , Hipertensão/fisiopatologia , Peptidil Dipeptidase A/genética , Interferência de RNA , Receptor Tipo 1 de Angiotensina/genética , Remodelação Ventricular , Animais , Inativação Gênica , Frequência Cardíaca , Hipertensão/genética , Masculino , Peptidil Dipeptidase A/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor Tipo 1 de Angiotensina/metabolismoRESUMO
OBJECTIVE: To investigate the effects of RNA interference (RNAi) targeting angiotensin-converting enzyme (ACE) on the blood pressure and myocardial remodeling in spontaneously hypertensive rats (SHRs). METHODS: Saline (control), adenovirus (Ad5) and recombinant adenoviral vectors (Ad5-ACE-shRNA expressing ACE gene-specific shRN) were randomly administered by caudal intravasation to SHRs (n = 12 each group) at day 1 and day 16. Normotensive Wistar-Kyoto rats (WKY) served as normal controls. Systolic blood pressure (SBP) of the caudal artery was measured daily. Expressions of ACE at mRNA and protein levels in myocardium and aorta were evaluated by RT-PCR and Western blot respectively, ACE serum concentration was measured by ELISA at day 3 (n = 6 each group). The ratio of left ventricular to body weight (LVW/BW), myocardial collagen content were measured and myocardial ultrastructure observed under transmission electron microscope at the study end. RESULTS: Ad5-ACE-shRNA injection significantly reduced SBP (-22 mm Hg, 1 mm Hg = 0.133 kPa) and the antihypertensive effect could last at least 14 days post each injection. SBP was not affected by saline and Ad5 injections. ACE expressions at mRNA and protein levels at myocardium and aorta as well as serum ACE were significantly decreased in Ad5-ACE-shRNA treated SHRs compared to that in saline and Ad5 groups (all P < 0.05) and was comparable to that in WKY group (P > 0.05). The LVW/BW ratio (2.24 +/- 0.19) and myocardial collagen content [(1.283 +/- 0.019) microg/mg] in Ad5-ACE-shRNA treated SHRs were also significantly lower than those in saline treated [3.21 +/- 0.13 and (1.686 +/- 0.013) microg/mg, both P < 0.05] and Ad5 treated SHRs [3.13 +/- 0.12, (1.682 +/- 0.009) microg/mg, both P < 0.05] but still higher than those of WKY group [2.06 +/- 0.11, (1.257 +/- 0.019) microg/mg, both P < 0.05]. Myocardial ultrastructure was also significantly improved in Ad5-ACE-shRNA treated SHRs compared to saline and Ad5 treated SHRs. CONCLUSION: RNAi targeting ACE gene significantly inhibited the expressions of ACE at mRNA and protein levels and resulted in prolonged antihypertensive effects and myocardial ultrastructure improvements in this SHR model.
Assuntos
Pressão Sanguínea , Hipertensão/fisiopatologia , Peptidil Dipeptidase A/metabolismo , Interferência de RNA , Remodelação Ventricular , Animais , Modelos Animais de Doenças , Frequência Cardíaca , Hipertensão/genética , Masculino , Peptidil Dipeptidase A/genética , RNA Mensageiro/genética , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKYRESUMO
Induction of oxidative stress has a causal role in atherosclerosis. The aim of the present study was to examine the role of lectinlike oxidized lowdensity lipoprotein receptor1 (LOX1) in oxidized lowdensity lipoprotein (OxLDL)induced oxidative stress in atherosclerosis. Small interfering RNA (siRNA) technology was employed to decrease the expression of LOX1 in mouse RAW264.7 macrophages and the effects of LOX1 silencing on OxLDLinduced reactive oxygen species (ROS) generation and NADPH oxidase (NOX) expression were investigated. The in vivo effects of reducing LOX1 were also examined in a mouse model (ApoE/) of highfat dietinduced atherosclerosis. Compared with the control cells, OxLDL exposure led to a significant (P<0.05) increase in the intracellular levels of malondialdehyde and ROS and a significant decrease in the activity of superoxide dismutase. Delivery of LOX1targeting siRNA significantly (P<0.05) reversed the alterations in oxidative stress parameters induced by OxLDL. LOX1 silencing downregulated the expression of NOX2, Rac1, p47phox and p22phox and impaired the activation of mitogenactivated protein kinases in OxLDLtreated cells. Adenoviral delivery of LOX1 siRNA caused a significant increase in the size of the fibrous cap and a decrease in the macrophage content in lesions, compared with the control mice. Western blot analysis demonstrated that the protein expression levels of NOX1, Rac1, p47phox and p22phox in aortic lesions were significantly lower in the LOX1 siRNA group than in the control group. LOX1 is implicated in OxLDLinduced oxidative stress of macrophages in atherosclerosis, which in part, involves the regulation of NADPH oxidases.
Assuntos
Aterosclerose/metabolismo , Aterosclerose/patologia , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , Estresse Oxidativo , Receptores Depuradores Classe E/metabolismo , Animais , Apolipoproteínas E/deficiência , Apolipoproteínas E/genética , Aterosclerose/genética , Linhagem Celular , Grupo dos Citocromos b/genética , Grupo dos Citocromos b/metabolismo , Modelos Animais de Doenças , Regulação da Expressão Gênica , Inativação Gênica , Lipídeos/sangue , Lipoproteínas LDL/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , NADPH Oxidase 2 , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Placa Aterosclerótica/genética , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologia , RNA Interferente Pequeno/genética , Receptores Depuradores Classe E/genética , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/metabolismoRESUMO
Angiotensin II (Ang II) and Ang-(1-7) are key effector peptides of the renin-angiotensin system. The present study aimed to investigate the effects of Ang-(1-7) on Ang II-stimulated cholesterol efflux and the associated molecular mechanisms. Differentiated THP-1 macrophages were treated with Ang II (1 µM) and/or Ang-(1-7) (10 and 100 nM) for 24 h and the cholesterol efflux and gene expression levels were assessed. Pharmacological inhibition of peroxisome proliferator-activated receptor (PPAR)γ and mitogen-activated protein kinases (MAPKs) were performed to identify the signaling pathways involved. The results demonstrated that Ang II significantly inhibited the cholesterol efflux from cholesterol-loaded THP-1 macrophages. Treatment with Ang-(1-7) led to a dose-dependent restoration of cholesterol efflux in the Ang II-treated cells. The co-treatment with Ang-(1-7) and Ang II significantly increased the expression levels of adenosine triphosphate (ATP)-binding cassette (ABC)A1 and ABCG1 compared with treatment with Ang II alone. This was coupled with increased expression levels of PPARγ and liver X receptor (LXR)α. The pharmacological inhibition of PPARγ significantly (P<0.05) eliminated the Ang-(1-7)-mediated induction of ABCA1 and ABCG1 mRNA expression. Treatment with Ang-(1-7) caused the inactivation of c-Jun N-terminal kinases (JNK) and p38 MAPK signaling in the Ang II-treated THP-1 macrophages. In addition, the inhibition of JNK or p38 MAPK signaling using specific pharmacological inhibitors mimicked the Ang-(1-7)-induced expression of PPARγ and LXRα. In conclusion, the data demonstrated that treatment with Ang-(1-7) promoted cholesterol efflux in Ang II-treated THP-1 macrophages, partly through inactivation of p38 and JNK signaling and by inducing the expression of PPARγ and LXRα. Ang (1-7) may, therefore, have therapeutic benefits for the treatment of atherosclerosis.
Assuntos
Aterosclerose/genética , Colesterol/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/biossíntese , Receptores Nucleares Órfãos/biossíntese , Proteínas Quinases p38 Ativadas por Mitógeno/biossíntese , Transportador 1 de Cassete de Ligação de ATP/biossíntese , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/biossíntese , Angiotensina I/administração & dosagem , Angiotensina II/administração & dosagem , Aterosclerose/tratamento farmacológico , Aterosclerose/patologia , Regulação Enzimológica da Expressão Gênica , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Receptores X do Fígado , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/patologia , Receptores Nucleares Órfãos/genética , PPAR gama/antagonistas & inibidores , Fragmentos de Peptídeos/administração & dosagem , RNA Mensageiro/biossíntese , Sistema Renina-Angiotensina , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
OBJECTIVE: An insertion/deletion (I/D) variant in the angiotensin-converting enzyme (ACE) gene was associated with ACE inhibitor (ACEI)-related cough in previous studies. However, the results were inconsistent. Our objective was to assess the relationship between the ACE I/D polymorphism and ACEI-related cough by meta-analysis and to summarize all studies that are related to ACE I/D polymorphism and ACEI-cough and make a summary conclusion to provide reference for the researchers who attempt to conduct such a study. METHODS: Databases including PubMed, EMbase, Cochrane Library, and China National Knowledge Infrastructure, were searched for genetic association studies. Data were extracted by two independent authors and pooled odds ratio (OR) with 95% confidence interval (CI) was calculated. Metaregression and subgroup analyses were performed to identify the source of heterogeneity. RESULTS: Eleven trials, including 906 cases (ACEI-related cough) and 1,175 controls, were reviewed in the present meta-analysis. The random effects pooled OR was 1.16 (95%CI: 0.78-1.74, p=0.46) in the dominant model and 1.61 (95%CI: 1.18-2.20, p=0.003) in the recessive model. Heterogeneity was found among and within studies. Metaregression indicated that the effect size was positively associated with age and negatively associated with follow-up duration of ACEI treatment. Subgroup analysis revealed a significant association between ACE I/D polymorphism and ACEI-related cough in studies with mean age >60 y, but not in studies with mean age ≤ 60 y. No heterogeneity was found within each mean age subgroup. We also found no association between ACE I/D polymorphism and ACEI-related cough in studies with follow-up>2 mo or in studies in Caucasians. No heterogeneity was detected in these two subgroups. CONCLUSIONS: Synthesis of the available evidence supports ACE I/D polymorphism as an age-dependent predictor for risk of ACEI-related cough.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/efeitos adversos , Tosse/induzido quimicamente , Tosse/genética , Deleção de Genes , Mutagênese Insercional , Peptidil Dipeptidase A/genética , Alelos , Estudos de Casos e Controles , Frequência do Gene , Genótipo , Humanos , Polimorfismo GenéticoRESUMO
AIM: To investigate the effects of H2-Bl gene on biological behaviour of mouse vascular smooth muscle cells (VSMCs) including apoptosis, proliferation and anti-cytolysis to peripheral blood mononuclear cells (PBMCs), then speculate the possible mechanism of immunological rejection after heart transplantation in maternal-fetal immune tolerance. METHODS: The mouse VSMC were transfected with 0.5 mg/L of pEGFP-N1 plasmid vector, 0.5 mg/L of pEGFP-N1-H2B1 plasmid vector and 1.0 mg/L of pEGFP-N1-H2B1 plasmid vector, respectively. The efficiency of transfection was detected, and the H2-Bl mRNA level, the apoptosis, the proliferation and cytotoxicity of VSMCs were also investigated at 24 h, 48 h and 72 h, respectively. RESULTS: Expression of the H2-Bl gene was determined by real time quantitative PCR. Expressions of the H2-Bl in experimental groups were higher than that of the control group (P<0.001). VSMC apoptosis was observed after 24 h in the 0.5 mg/L and 1.0 mg/L H2-Bl experimental groups compared with control group (P<0.05, P<0.001, respectively). VSMC proliferation was also inhibited in the 0.5 mg/L and 1.0 mg/L H2-Bl gene groups at 24 h and 72 h (P<0.05, P<0.01, respectively). The cytotoxicity of PBMC in the 0.5 mg/L and 1.0 mg/L H2-Bl gene groups at 24 h was lower than the control group (P<0.005, P<0.001, respectively). CONCLUSION: Transfection of pEGFP-N1-H2B1 plasmid to mouse VSMC causes the over expression of H2-Bl mRNA, induces the apoptosis, inhibits the proliferation and attenuates the cytotoxicity of PBMC, leading to immune tolerance.
Assuntos
Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Transfecção , Animais , Apoptose/genética , Proliferação de Células , Células Cultivadas , Citotoxicidade Imunológica/genética , Citotoxicidade Imunológica/imunologia , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Genes MHC da Classe II , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcrição GênicaRESUMO
Adiponectin (APN) is a potent cardioprotective molecule. The present study aims to investigate the underlying mechanism(s) for its cardioprotective effect. We isolated primary cardiomyocytes from neonatal rats and established an in vitro model of hypoxia/reoxygenation (H/R). The cardiomyocytes were randomly divided into 6 groups: saline group (control), dithiothreitol group (5 mM dithiothreitol for 2 hours), H/R group, H/R + APN group (incubation with 30 µg/mL APN, followed by H/R), H/R + APN + SB203580 (SB) group (treatment with 30 µg/mL APN and 5 µM SB, followed by H/R), and H/R + SB group (exposure to 5 µM SB and then H/R). Cell death was detected by measuring lactate dehydrogenase release. The expression levels of hypoxia-inducible factor 1α and endoplasmic reticulum stress-related genes including GRP78, caspase-12, C/EBP homologous protein, and p38 mitogen-activated protein kinase were examined. Cardiomyocytes exposed to H/R showed a significant increase in lactate dehydrogenase leakage and hypoxia-inducible factor 1α protein levels compared with the control cells (P < 0.05). The H/R-provoked cell death was profoundly attenuated by the pretreatment with APN alone, SB alone, or both, which was coupled with decreased expression of GRP78, caspase-12, C/EBP homologous protein, and p38 mitogen-activated protein kinase. These results provide new insights into the mechanism of APN-mediated cardioprotection, which may be partially due to inhibition of endoplasmic reticulum stress response.
Assuntos
Adiponectina/farmacologia , Hipóxia/tratamento farmacológico , Miócitos Cardíacos/efeitos dos fármacos , Oxigênio/química , Animais , Animais Recém-Nascidos , Ditiotreitol/farmacologia , Retículo Endoplasmático/metabolismo , Chaperona BiP do Retículo Endoplasmático , Inibidores Enzimáticos/farmacologia , Coração/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Imidazóis/farmacologia , L-Lactato Desidrogenase/metabolismo , Miócitos Cardíacos/metabolismo , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND AND AIMS: Amlodipine, a long-acting dihydropyridine calcium channel blocker, is able to improve angiotensin II-mediated vascular endothelial dysfunction. However, the underlying mechanism remains not fully understood. In the present study we attempted to determine whether the protective effect of amlodipine against Ang II-induced endothelial impairment was mediated through blockage of endothelial cell apoptosis. METHODS: We pretreated human umbilical venous endothelial cells with increasing doses of amlodipine (10(-8)-10(-6) M) followed by the addition of Ang II. Cell apoptosis was assessed by acridine orange/ethidium bromide staining and by annexin-V/propidium iodide double-labeled cytometry. The involvement of the apoptosis regulators, Bcl-2, Bax, and lectin-like oxidized low-density lipoprotein receptor-1, was determined. RESULTS: Pretreatment with amlodipine resulted in a dose-dependent suppression of Ang II-induced HUVEC apoptosis. Moreover, the Bcl-2/Bax ratio was found to be increased in cells pretreated with amlodipine, indicating an enhanced anti-apoptosis potential. Additionally, the induction of LOX-1 by Ang II was remarkably counteracted by the pre-exposure to amlodipine. CONCLUSIONS: Our data demonstrate that amlodipine ameliorates Ang II-induced endothelial apoptosis, which is likely associated with the elevation of Bcl-2/Bax ratio and reduction of the LOX-1 expression.