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1.
Endocrine ; 2024 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-39198343

RESUMO

PURPOSE: Diabetes mellitus and metabolic homeostasis disorders may benefit from white adipose tissue (WAT) browning, which is associated with mitochondrial fission. Resveratrol, a dietary polyphenol, exhibits beneficial effects against abnormalities related to metabolic diseases. However, it remains unknown whether resveratrol contributes to WAT browning by regulating mitochondrial fission. METHODS: We administered resveratrol (0.4% mixed with control) to db/db mice for 12 weeks, measuring body weight, oral glucose tolerance, insulin tolerance, and histological changes. The uncoupling protein 1 (UCP1) and dynamin-related protein 1 (DRP1) expressions in the epididymal WAT were assessed via immunoblotting. RESULTS: We found that resveratrol improved systemic glucose homeostasis and insulin resistance in db/db mice, which was associated with increased UCP1 in epididymal WAT. Resveratrol-treated mice exhibited more fragmented mitochondria and increased phosphorylation of DRP1 in the epididymal WAT of the db/db mice. These results were further confirmed in vitro, where resveratrol induced extracellular signal-regulated kinase (ERK) signaling activation, leading to phosphorylation of DRP1 at the S616 site (p-DRP1S616) and mitochondrial fission, which was reversed by an ERK inhibitor in 3T3-L1 adipocytes. CONCLUSION: Resveratrol plays a role in regulating the phosphorylation of ERK and DRP1, resulting in the promotion of beige cells with epididymal WAT and the improvement of glucose homeostasis. Our present study provides novel insights into the potential mechanism of resveratrol-mediated effects on WAT browning, suggesting that it is, at least in part, mediated through ERK/DRP1-mediated mitochondrial fission.

2.
J Ethnopharmacol ; 305: 116040, 2023 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-36539071

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: JinHong Formula (JHF) was derived from the famous Rhubarb and Moutan Decoction which was prescribed for appendicitis. It was originally recorded in the classic of "Jingui Yaolve" written by Zhang Zhongjing. It is a kind of traditional Chinese medicine, widely used in the treatment of inflammation. However, the clinical effect of JHF for sepsis and its comprehensive mechanism in sepsis remained largely unknown. RESEARCH PURPOSE: The aim of our study was to evaluate the clinical effect of JHF in the treatment of sepsis, and to explore its mechanism from the perspective of network pharmacology. RESEARCH METHODS: The single-center randomized clinical trial was conducted to assess the effect of JHF in the treatment of sepsis. Additionally, we used the Chinese herbal medicine pharmacology database and analysis platform to identify the active components and therapeutic target of JHF. Numerous well-known disease target databases have been used to screen therapeutic target proteins for sepsis. Furthermore, we have established a Protein-Protein Interaction (PPI) network and carried out Gene Onotology/Kyoto Encyclopedia of Genes and Genomes (GO/KEGG) enrichment analysis. In order to conclude which active compounds from JHF may be responsible for signaling pathway, we performed network analysis. RESEARCH RESULTS: The study included 114 patients. By comparing participants with and without JHF, the results suggested that JHF significantly reduced all-cause mortality on 28 and 60 days after intervention, and improved Sequential Organ Failure Assessment (SOFA) on 7th day after intervention as well as. JHF had an effect of anti-inflammatories and antioxidants (SOD). By using network pharmacological analysis, we identified 72 active components and 426 target genes of JHF, and successfully constructed a "JHF-compound target-sepsis" network. 116 mentioned targets revealed by GO/KEGG enrichment analysis played a significant role in the inflammatory reaction and immunoregulation via interleukin-17 (IL-17) and tumor necrosis factor (TNF) signaling pathway. Moreover, the analysis of "pathway target-active component" revealed that Sennidin A, Rheidin A, Rheidin B, Rheidin C, (E)-4-Phenyl-3-Buten-2-One, Osmanthuside H, Esculetin, and Caffeicacid were responsible for IL-17, TNF signaling pathways. CONCLUSION: JHF contains potential active substance of anti-inflammatory and antioxidant. These active compounds may come into play through IL-17 and TNF signaling pathways. For sepsis, JHF may be a promising and effective treatment strategy.


Assuntos
Medicamentos de Ervas Chinesas , Sepse , Humanos , Medicamentos de Ervas Chinesas/farmacologia , Medicamentos de Ervas Chinesas/uso terapêutico , Interleucina-17 , Farmacologia em Rede , Sepse/tratamento farmacológico , Medicina Tradicional Chinesa , Antioxidantes , Inflamação , Simulação de Acoplamento Molecular
3.
Clin Sci (Lond) ; 136(12): 989-1003, 2022 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-35695067

RESUMO

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/metabolismo
4.
J Clin Invest ; 131(24)2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34907911

RESUMO

Circular RNAs (circRNAs) have been recently recognized as playing a role in the pathogenesis of vascular remodeling-related diseases by modulating the functions of miRNAs. However, the interplay between circRNAs and proteins during vascular remodeling remains poorly understood. Here, we investigated a previously identified circRNA, circEsyt2, whose expression is known to be upregulated during vascular remodeling. Loss- and gain-of­function mutation analyses in vascular smooth muscle cells (VSMCs) revealed that circEsyt2 enhanced cell proliferation and migration and inhibited apoptosis and differentiation. Furthermore, the silencing of circEsyt2 in vivo reduced neointima formation, while circEsyt2 overexpression enhanced neointimal hyperplasia in the injured carotid artery, confirming its role in vascular remodeling. Using unbiased protein-RNA screening and molecular validation, circEsyt2 was found to directly interact with polyC-binding protein 1 (PCBP1), an RNA splicing factor, and regulate PCBP1 intracellular localization. Additionally, circEsyt2 silencing substantially enhanced p53ß splicing via the PCBP1-U2AF65 interaction, leading to the altered expression of p53 target genes (cyclin D1, p21, PUMA, and NOXA) and the decreased proliferation of VSMCs. Thus, we identified a potentially novel circRNA that regulated vascular remodeling, via altered RNA splicing, in atherosclerotic mouse models.


Assuntos
Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Splicing de RNA , RNA Circular/metabolismo , Proteína Supressora de Tumor p53/biossíntese , Remodelação Vascular , Animais , Aterosclerose/genética , Aterosclerose/metabolismo , Proliferação de Células , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Hiperplasia/genética , Hiperplasia/metabolismo , Camundongos , Camundongos Knockout para ApoE , RNA Circular/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Fator de Processamento U2AF/genética , Fator de Processamento U2AF/metabolismo , Proteína Supressora de Tumor p53/genética
5.
Front Cell Dev Biol ; 9: 673993, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34790655

RESUMO

Infection is thought to be involved in the pathogenesis of atherosclerosis. Studies have shown the association between helicobacter pylori (H. pylori) and coronary artery disease. It is interesting to find H. pylori DNA and cytotoxin-associated gene A (CagA) protein in atherosclerotic plaque. Outer membrane vesicles (OMVs), secreted by H. pylori, exert effects in the distant organ or tissue. However, whether or not OMVs from H. pylori are involved in the pathogenesis of atherosclerosis remains unknown. Our present study found that treatment with OMVs from CagA-positive H. pylori accelerated atherosclerosis plaque formation in ApoE-/- mice. H. pylori-derived OMVs inhibited proliferation and promoted apoptosis of human umbilical vein endothelial cells (HUVECs), which was also reflected in in vivo studies. These effects were normalized to some degree after treatment with lipopolysaccharide (LPS)-depleted CagA-positive OMVs or CagA-negative OMVs. Treatment with H. pylori-derived OMVs increased reactive oxygen species (ROS) levels and enhanced the activation of nuclear factor-κB (NF-κB) in HUVECs, which were reversed to some degree in the presence of a superoxide dismutase mimetic TEMPOL and a NF-κB inhibitor BAY11-7082. Expressions of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α), two inflammatory factors, were augmented after treatment with OMVs from H. pylori. These suggest that H. pylori-derived OMVs accelerate atherosclerosis plaque formation via endothelium injury. CagA and LPS from H. pylori-OMVs, at least in part, participate in these processes, which may be involved with the activation of ROS/NF-κB signaling pathway. These may provide a novel strategy to reduce the incidence and development of atherosclerosis.

6.
Sci Rep ; 11(1): 15836, 2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34349170

RESUMO

Acute myocardial infarction (MI) is one of the leading causes of death in humans. Our previous studies showed that gastrin alleviated acute myocardial ischaemia-reperfusion injury. We hypothesize that gastrin might protect against heart injury after MI by promoting angiogenesis. An MI model was simulated by ligating the anterior descending coronary artery in adult male C57BL/6J mice. Gastrin was administered twice daily by intraperitoneal injection for 2 weeks after MI. We found that gastrin reduced mortality, improved myocardial function with reduced infarct size and promoted angiogenesis. Gastrin increased HIF-1α and VEGF expression. Downregulation of HIF-1α expression by siRNA reduced the proliferation, migration and tube formation of human umbilical vein endothelial cells. These results indicate that gastrin restores cardiac function after MI by promoting angiogenesis via the HIF-1α/VEGF pathway.


Assuntos
Gastrinas/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Infarto do Miocárdio/tratamento farmacológico , Neovascularização Patológica/prevenção & controle , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Proliferação de Células , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/patologia , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/genética
7.
Cell Prolif ; 53(11): e12910, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33047378

RESUMO

OBJECTIVES: The mechanisms responsible for the postnatal loss of mammalian cardiac regenerative capacity are not fully elucidated. The aim of the present study is to investigate the role of progesterone in cardiac regeneration and explore underlying mechanism. MATERIALS AND METHODS: Effect of progesterone on cardiomyocyte proliferation was analysed by immunofluorescent staining. RNA sequencing was performed to screen key target genes of progesterone, and yes-associated protein (YAP) was knocked down to demonstrate its role in pro-proliferative effect of progesterone. Effect of progesterone on activity of YAP promoter was measured by luciferase assay and interaction between progesterone receptor and YAP promoter by electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP). Adult mice were subjected to myocardial infarction, and then, effects of progesterone on adult cardiac regeneration were analysed. RESULTS: Progesterone supplementation enhanced cardiomyocyte proliferation in a progesterone receptor-dependent manner. Progesterone up-regulated YAP expression and knockdown of YAP by small interfering RNA reduced progesterone-mediated cardiomyocyte proliferative effect. Progesterone receptor interacted with the YAP promoter, determined by ChIP and EMSA; progesterone increased luciferase activity of YAP promoter and up-regulated YAP target genes. Progesterone administration also promoted adult cardiomyocyte proliferation and improved cardiac function in myocardial infarction. CONCLUSION: Our data uncover a role of circulating progesterone withdrawal as a novel mechanism for the postnatal loss of mammalian cardiac regenerative potential. Progesterone promotes both neonatal and adult cardiomyocyte proliferation by up-regulating YAP expression.


Assuntos
Proteínas Reguladoras de Apoptose/genética , Cardiotônicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Progesterona/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Cardiotônicos/uso terapêutico , Proteínas de Ciclo Celular/genética , Células Cultivadas , Masculino , Camundongos Endogâmicos C57BL , Infarto do Miocárdio/tratamento farmacológico , Infarto do Miocárdio/genética , Miócitos Cardíacos/citologia , Miócitos Cardíacos/metabolismo , Progesterona/uso terapêutico , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacos , Proteínas de Sinalização YAP
8.
Circ Heart Fail ; 13(1): e006525, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31957467

RESUMO

BACKGROUND: Long noncoding RNA (lncRNA) can regulate various physiological and pathological processes through multiple molecular mechanisms in cis and in trans. However, the role of lncRNAs in cardiac hypertrophy is yet to be fully elucidated. METHODS: A mouse lncRNA microarray was used to identify differentially expressed lncRNAs in the mouse hearts following transverse aortic constriction-induced pressure overload comparing to the sham-operated samples. The direct impact of one lncRNA, Ahit, on cardiomyocyte hypertrophy was characterized in neonatal rat cardiomyocytes in response to phenylephrine by targeted knockdown and overexpression. The in vivo function of Ahit was analyzed in mouse hearts by using cardiac-specific adeno-associated virus, serotype 9-short hairpin RNA to knockdown Ahit in combination with transverse aortic constriction. Using catRAPID program, an interaction between Ahit and SUZ12 (suppressor of zeste 12 protein homolog) was predicted and validated by RNA immunoprecipitation and immunoblotting following RNA pull-down. Chromatin immunoprecipitation was performed to determine SUZ12 or H3K27me3 occupancy on the MEF2A (myocyte enhancer factor 2A) promoter. Finally, the expression of human Ahit (leukemia-associated noncoding IGF1R activator RNA 1 [LUNAR1]) in the serum samples from patients of hypertrophic cardiomyopathy was tested by quantitative real-time polymerase chain reaction. RESULTS: A previously unannotated lncRNA, antihypertrophic interrelated transcript (Ahit), was identified to be upregulated in the mouse hearts after transverse aortic constriction. Inhibition of Ahit induced cardiac hypertrophy, both in vitro and in vivo, associated with increased expression of MEF2A, a critical transcriptional factor involved in cardiac hypertrophy. In contrast, overexpression of Ahit significantly attenuated stress-induced cardiac hypertrophy in vitro. Furthermore, Ahit was significantly upregulated in serum samples of patients diagnosed with hypertensive heart disease versus nonhypertrophic hearts (1.46±0.17 fold, P=0.0325). Mechanistically, Ahit directly bound and recruited SUZ12, a core PRC2 (polycomb repressive complex 2) protein, to the promoter of MEF2A, triggering its trimethylation on H3 lysine 27 (H3K27me3) residues and mediating the downregulation of MEF2A, thereby preventing cardiac hypertrophy. CONCLUSIONS: Ahit is a lncRNA with a significant role in cardiac hypertrophy regulation through epigenomic modulation. Ahit is a potential therapeutic target of cardiac hypertrophy.


Assuntos
Cardiomegalia/metabolismo , Insuficiência Cardíaca/metabolismo , Complexo Repressor Polycomb 2/metabolismo , RNA Longo não Codificante/metabolismo , Animais , Cardiomegalia/patologia , Regulação para Baixo , Regulação da Expressão Gênica/genética , Insuficiência Cardíaca/genética , Humanos , Fatores de Transcrição MEF2/metabolismo , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/metabolismo , Proteínas de Neoplasias , RNA Longo não Codificante/genética , Ratos Sprague-Dawley , Fatores de Transcrição
9.
Am J Hypertens ; 33(1): 31-42, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31328772

RESUMO

BACKGROUND: Essential hypertension is associated with increased plasma concentrations of extracellular vesicles (EVs). We aimed to determine the role of monocyte miR-27a in EVs on arterial Mas receptor expression, and its involvement in the pathogenesis of hypertension. METHODS: THP-1 cells were transfected with miR-27a mimic and miR-27a inhibitor, and EVs were collected. Mas receptor expression and endothelial nitric oxide synthase (eNOS) phosphorylation were determined by immunoblotting. Sprague-Dawley (SD) rats received EVs via tail-vein injection. Blood pressure (BP) was measured with the tail-cuff method. The vasodilatory response of mesenteric arteries was measured using a small vessel myograph. RESULTS: EVs from THP-1 cells increased rat BP by impairing Ang-(1-7)-mediated vasodilation in mesenteric arteries, which was further exaggerated by EVs from lipopolysaccharides-treated THP-1 cells. As the receptor and key signaling of Ang-(1-7), next experiments found that Mas receptor expression and eNOS phosphorylation were decreased in mesenteric arteries from EVs-treated SD rats. Screening studies found miR-27a in EVs may be involved in this process. Through transfection with miR-27a inhibitor or miR-27a mimic, we found that miR-27a downregulates Mas receptor expression in endothelial cells. Injection of EVs from miR-27a-transfected HEK-293 cells decreased Mas receptor and eNOS phosphorylation in mesenteric arteries, impaired Ang-(1-7)-mediated vasodilation and increased BP. Earlier effects were reversed using cells with downregulation of miR-27 in EVs. CONCLUSIONS: Monocyte miR-27a in EVs decreases Mas receptor expression and eNOS phosphorylation in endothelium, impairs Ang-(1-7)-mediated vasodilation, and causes hypertension. Understanding the contributions of EVs in the pathogenesis of hypertension may facilitate their use as a diagnostic biomarker.


Assuntos
Pressão Sanguínea , Vesículas Extracelulares/metabolismo , Hipertensão/enzimologia , Artérias Mesentéricas/enzimologia , MicroRNAs/metabolismo , Monócitos/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animais , Modelos Animais de Doenças , Vesículas Extracelulares/genética , Vesículas Extracelulares/transplante , Células HEK293 , Humanos , Hipertensão/genética , Hipertensão/fisiopatologia , Masculino , Artérias Mesentéricas/fisiopatologia , MicroRNAs/genética , Monócitos/transplante , Fosforilação , Proto-Oncogene Mas , Ratos Sprague-Dawley , Transdução de Sinais , Células THP-1 , Vasodilatação
10.
Stem Cell Reports ; 13(3): 559-571, 2019 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-31402338

RESUMO

Development of spermatogonia and spermatocytes are the critical steps of spermatogenesis, impacting on male fertility. Investigation of the related regulators benefits the understanding of male reproduction. The proteasome system has been reported to regulate spermatogenesis, but the mechanisms and key contributing factors in vivo are poorly explored. Here we found that ablation of REGγ, a proteasome activator, resulted in male subfertility. Analysis of the mouse testes after birth showed there was a decreased number of PLZF+ spermatogonia and spermatocytes. Molecular analysis found that REGγ loss significantly increased the abundance of p53 protein in the testis, and directly repressed PLZF transcription in cell lines. Of note, allelic p53 haplodeficiency partially rescued the defects in spermatogenesis observed in REGγ-deficient mice. In summary, our results identify REGγ-p53-PLZF to be a critical pathway that regulates spermatogenesis and establishes a new molecular link between the proteasome system and male reproduction.


Assuntos
Autoantígenos/metabolismo , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Espermatogênese , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose , Autoantígenos/genética , Masculino , Meiose , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oligonucleotídeos Antissenso , Regiões Promotoras Genéticas , Proteína com Dedos de Zinco da Leucemia Promielocítica/antagonistas & inibidores , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Complexo de Endopeptidases do Proteassoma/deficiência , Complexo de Endopeptidases do Proteassoma/genética , Ligação Proteica , Transdução de Sinais , Motilidade dos Espermatozoides , Espermatócitos/citologia , Espermatócitos/metabolismo , Espermatogônias/citologia , Espermatogônias/metabolismo , Testículo/metabolismo , Proteína Supressora de Tumor p53/genética
11.
Sci Rep ; 9(1): 7693, 2019 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-31118464

RESUMO

Genome-wide association studies (GWAS) have identified many genetic variants in genes related to lipid metabolism. However, how these variations affect lipid levels remains elusive. Long non-coding RNAs (lncRNAs) have been implicated in a variety of biological processes. We hypothesize lncRNAs are likely to be located within disease or trait-associated DNA regions to regulate lipid metabolism. The aim of this study was to investigate whether and how lncRNAs in lipid- associated DNA regions regulate cholesterol homeostasis in hepatocytes. In this study, we identified a novel long non-coding RNA in Lipid Associated Single nucleotide polymorphism gEne Region (LASER) by bioinformatic analysis. We report that LASER is highly expressed in both hepatocytes and peripheral mononuclear cells (PBMCs). Clinical studies showed that LASER expression is positively related with that of cholesterol containing apolipoprotein levels. In particular, we found that LASER is positively correlated with plasma PCSK9 levels in statin free patients. siRNAs mediated knock down of LASER dramatically reduces intracellular cholesterol levels and affects the expression of genes involved in cholesterol metabolism. Transcriptome analyses show that knockdown of LASER affects the expression of genes involved in metabolism pathways. We found that HNF-1α and PCSK9 were reduced after LASER knock-down. Interestingly, the reduction of PCSK9 can be blocked by the treatment of berberine, a natural cholesterol-lowering compound which functions as a HNF-1α antagonist. Mechanistically, we found that LASER binds to LSD1 (lysine-specific demethylase 1), a member of CoREST/REST complex, in nucleus. LASER knock-down enhance LSD1 targeting to genomic loci, resulting in decreased histone H3 lysine 4 mono-methylation at the promoter regions of HNF-1α gene. Conversely, LSD1 knock-down abolished the effect of LASER on HNF-1α and PCSK9 expressions. Finally, we found that statin treatment increased LASER expression, accompanied with increased PCSK9 expression, suggesting a feedback regulation of cholesterol on LASER expression. This observation may partly explain the statin escape during anti-cholesterol treatment. These findings identified a novel lncRNA in cholesterol homeostasis. Therapeutic targeting LASER might be an effective approach to augment the effect of statins on cholesterol levels in clinics.


Assuntos
Colesterol/metabolismo , Hepatócitos/metabolismo , Leucócitos Mononucleares/metabolismo , RNA Longo não Codificante/fisiologia , Anticolesterolemiantes/farmacologia , Anticolesterolemiantes/uso terapêutico , Apolipoproteínas/metabolismo , Atorvastatina/farmacologia , Atorvastatina/uso terapêutico , Benzoatos/farmacologia , Benzilaminas/farmacologia , Berberina/farmacologia , Cromossomos Humanos Par 11/genética , Doença das Coronárias/tratamento farmacológico , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Células Hep G2 , Fator 1-alfa Nuclear de Hepatócito/biossíntese , Fator 1-alfa Nuclear de Hepatócito/genética , Código das Histonas/efeitos dos fármacos , Histona Desmetilases/metabolismo , Homeostase/genética , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Hipercolesterolemia/tratamento farmacológico , Hipercolesterolemia/metabolismo , Pró-Proteína Convertase 9/biossíntese , Pró-Proteína Convertase 9/genética , Ligação Proteica , Mapeamento de Interação de Proteínas , Interferência de RNA , RNA Longo não Codificante/genética , RNA Interferente Pequeno/genética
12.
Hypertension ; 72(4): 962-970, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30354705

RESUMO

Epidemiological and experimental studies suggest that maternal diabetes mellitus programs hypertension that is associated with impaired sodium excretion in the adult offspring. However, the underlying mechanisms are not clear. Because dopamine receptor function is involved in the pathogenesis of hypertension, we hypothesized that impaired renal dopamine D1 receptor function is also involved in the hypertension in offspring of maternal diabetes mellitus. Maternal diabetes mellitus was induced by a single intraperitoneal injection of streptozotocin (35 mg/kg) to pregnant Sprague-Dawley rats at day 0 of gestation. Compared with the offspring of mothers injected with citrate buffer (control mother offspring), the diabetic mother offspring (DMO) had increased systolic blood pressure and impaired D1 receptor-mediated diuresis and natriuresis, accompanied by increased renal PKC (protein kinase C) expression and activity, GRK-2 (G protein-coupled receptor kinase-2) expression, D1 receptor phosphorylation, D1 receptor/Gαs uncoupling, and loss of D1 receptor-mediated inhibition of Na+-K+-ATPase activity in renal proximal tubule cells from DMO. Inhibition of PKC reduced the increased GRK-2 expression and normalized D1 receptor function in primary cultures of renal proximal tubule cells from DMO. In addition, DMO, relative to control mother offspring, in vivo, had increased oxidative stress, indicated by decreased renal glutathione and increased renal malondialdehyde and urine 8-isoprostane. Normalization of oxidative stress with tempol also normalized the renal D1 receptor phosphorylation, D1 receptor-mediated diuresis and natriuresis, and blood pressure in DMO. Our present study indicates that maternal diabetes mellitus-programed hypertension in the offspring is caused by impaired renal D1 receptor function because of oxidative stress that is mediated by increased PKC-GRK-2 activity.


Assuntos
Diabetes Mellitus , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Hipertensão , Estresse Oxidativo/efeitos dos fármacos , Complicações na Gravidez/metabolismo , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Proteína Quinase C , Receptores de Dopamina D1/metabolismo , Animais , Antioxidantes/farmacologia , Pressão Sanguínea/fisiologia , Óxidos N-Cíclicos/farmacologia , Diabetes Mellitus/etiologia , Diabetes Mellitus/metabolismo , Feminino , Hipertensão/diagnóstico , Hipertensão/metabolismo , Masculino , Fosforilação , Gravidez , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Ratos , Ratos Sprague-Dawley , Eliminação Renal , ATPase Trocadora de Sódio-Potássio/metabolismo , Marcadores de Spin
13.
J Am Heart Assoc ; 7(14)2018 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-30005556

RESUMO

BACKGROUND: Ischemia/reperfusion injury (IRI) is one of the most predominant complications of ischemic heart disease. Gastrin has emerged as a regulator of cardiovascular function, playing a key protective role in hypoxia. Serum gastrin levels are increased in patients with myocardial infarction, but the pathophysiogical significance of this finding is unknown. The purpose of this study was to determine whether and how gastrin protects cardiac myocytes from IRI. METHODS AND RESULTS: Adult male Sprague-Dawley rats were used in the experiments. The hearts in living rats or isolated Langendorff-perfused rat hearts were subjected to ischemia followed by reperfusion to induce myocardial IRI. Gastrin, alone or with an antagonist, was administered before the induction of myocardial IRI. We found that gastrin improved myocardial function and reduced the expression of myocardial injury markers, infarct size, and cardiomyocyte apoptosis induced by IRI. Gastrin increased the phosphorylation levels of ERK1/2 (extracellular signal-regulated kinase 1/2), AKT (protein kinase B), and STAT3 (signal transducer and activator of transcription 3), indicating its ability to activate the RISK (reperfusion injury salvage kinase) and SAFE (survivor activating factor enhancement) pathways. The presence of inhibitors of ERK1/2, AKT, or STAT3 abrogated the gastrin-mediated protection. The protective effect of gastrin was via CCK2R (cholecystokinin 2 receptor) because the CCK2R blocker CI988 prevented the gastrin-mediated protection of the heart with IRI. Moreover, we found a negative correlation between serum levels of cardiac troponin I and gastrin in patients with unstable angina pectoris undergoing percutaneous coronary intervention, suggesting a protective effect of gastrin in human cardiomyocytes. CONCLUSIONS: These results indicate that gastrin can reduce myocardial IRI by activation of the RISK and SAFE pathways.


Assuntos
Gastrinas/farmacologia , Coração/efeitos dos fármacos , Hormônios/farmacologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Idoso , Angina Instável/sangue , Angina Instável/cirurgia , Animais , Apoptose/efeitos dos fármacos , Feminino , Gastrinas/sangue , Humanos , Preparação de Coração Isolado , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Miocárdio/patologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Intervenção Coronária Percutânea , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Receptor de Colecistocinina B/antagonistas & inibidores , Receptor de Colecistocinina B/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/efeitos dos fármacos , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Troponina I/sangue
14.
J Am Soc Hypertens ; 12(2): 135-145, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29289466

RESUMO

Both renin-angiotensin systems and insulin participate in kidney-involved blood pressure regulation. Activation of angiotensin II type 2 receptor (AT2R) decreases sodium reabsorption in renal proximal tubule (RPT) cells, whereas insulin produces the opposite effect. We presume that AT2R has an inhibitory effect on insulin receptor expression in RPT cells, which may affect renal sodium transport and therefore be of physiological or pathological significance. Our present study found that activation of AT2R inhibited insulin receptor expression in a concentration and time-dependent manner in RPT cells from Wistar-Kyoto (WKY) rats. In the presence of a protein kinase C (PKC) inhibitor (PKC inhibitor peptide 19-31, 10-6 mol/L) or a phosphatidylinositol 3 kinase inhibitor (wortmannin, 10-6 mol/L), the inhibitory effect of AT2R on insulin receptor was blocked, indicating that both PKC and phosphatidylinositol 3 kinase were involved in the signaling pathway. There was a linkage between AT2R and insulin receptor which was determined by both laser confocal microscopy and coimmunoprecipitation. However, the effect of AT2R activation on insulin receptor expression was different in RPT cells from spontaneously hypertensive rats (SHRs). Being contrary to the effect in WKY RPT cells, AT2R stimulation increased insulin receptor in SHR RPT cells. Insulin (10-7 mol/L, 15 minutes) enhanced Na+-K+-ATPase activity in both WKY and SHR RPT cells. Pretreatment with CGP42112 decreased the stimulatory effect of insulin on Na+-K+-ATPase activity in WKY RPT cells, whereas pretreatment with CGP42112 increased it in SHR RPT cells. It is suggested that activation of AT2R inhibits insulin receptor expression and function in RPT cells. The lost inhibitory effect of AT2R on insulin receptor expression may contribute to the pathophysiology of hypertension.


Assuntos
Pressão Sanguínea/fisiologia , Túbulos Renais Proximais/metabolismo , Receptor Tipo 2 de Angiotensina/metabolismo , Receptor de Insulina/metabolismo , Sistema Renina-Angiotensina/fisiologia , Animais , Linhagem Celular , Regulação da Expressão Gênica/fisiologia , Hipertensão/patologia , Túbulos Renais Proximais/efeitos dos fármacos , Fosfatidilinositol 3-Quinase/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor de Insulina/fisiologia , Eliminação Renal , Sistema Renina-Angiotensina/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Sódio/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo
15.
Biochim Biophys Acta Mol Basis Dis ; 1863(8): 2085-2092, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28249798

RESUMO

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/patologia
16.
J Am Heart Assoc ; 5(10)2016 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-27792639

RESUMO

BACKGROUND: G protein-coupled receptor kinase type 4 (GRK4) plays a vital role in the long-term control of blood pressure (BP) and sodium excretion by regulating renal G protein-coupled receptor phosphorylation, including dopamine type 1 receptor (D1R). Ultrasound-targeted microbubble destruction (UTMD) is a promising method for gene delivery. Whether this method can deliver GRK4 small interfering RNA (siRNA) and lower BP is not known. METHODS AND RESULTS: BP, 24-hour sodium excretion, and urine volume were measured after UTMD-targeted GRK4 siRNA delivery to the kidney in spontaneously hypertensive rats. The expression levels of GRK4 and D1R were determined by immunoblotting. The phosphorylation of D1R was investigated using immunoprecipitation. The present study revealed that UTMD-mediated renal GRK4 siRNA delivery efficiently reduced GRK4 expression and lowered BP in spontaneously hypertensive rats, accompanied by increased sodium excretion. The increased sodium excretion might be accounted for by the UTMD regulation of D1R phosphorylation and function in spontaneously hypertensive rats. Further analysis showed that, although UTMD had no effect on D1R expression, it reduced D1R phosphorylation in spontaneously hypertensive rats kidneys and consequently increased D1R-mediated natriuresis and diuresis. CONCLUSIONS: Taken together, these study results indicate that UTMD-targeted GRK4 siRNA delivery to the kidney effectively reduces D1R phosphorylation by inhibiting renal GRK4 expression, improving D1R-mediated natriuresis and diuresis, and lowering BP, which may provide a promising novel strategy for gene therapy for hypertension.


Assuntos
Pressão Sanguínea/genética , Quinase 4 de Receptor Acoplado a Proteína G/genética , Técnicas de Silenciamento de Genes/métodos , Técnicas de Transferência de Genes , Receptores de Dopamina D1/metabolismo , Ondas Ultrassônicas , Animais , Regulação para Baixo , Quinase 4 de Receptor Acoplado a Proteína G/metabolismo , Rim , Masculino , Microbolhas , RNA Interferente Pequeno , Ratos , Ratos Endogâmicos SHR , Sódio/urina
17.
J Hypertens ; 34(8): 1599-606, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27254310

RESUMO

BACKGROUND: Dopamine receptors induce natriuresis in kidney. Previous studies have shown interactions between different subtypes of dopamine receptors in renal proximal tubule (RPT) cells. We hypothesize that D3 receptors have an interaction with D4 receptors in RPT cells from normotensive rats (Wistar-Kyoto, WKY) and spontaneously hypertensive rats (SHRs). METHODS: Immunoblotting and reverse transcriptase-polymerase chain reaction (RT-PCR) were used to examine the expression of D3 and D4 receptors. Na-K-ATPase activity was used to measure the function of receptors. The distribution and colocalization of D3 and D4 receptors were detected by confocal microscopy and co-immunoprecipitation. RESULTS: D3 receptor agonist PD128907 increased the mRNA and protein expression of D4 receptors in RPT cells from WKY rats, but decreased that from SHRs. In the presence of PLC blocker (U73122, 10-mol/l) or PKC inhibitor 19 -31 (10-mol/l), the up-regulation of D3 receptor on D4 receptor was lost in WKY cells. Moreover, stimulation with PD128907 for 30 minutes decreased D4 receptor degradation in WKY cells, not in SHR cells. D3 and D4 receptors colocalized and co-immunoprecipitated in RPT cells. PD128907 increased co-immunoprecipitation of D3 and D4 receptors in WKY RPT cells, but not in SHR RPT cells. Pre-treatment with D3 receptor agonist also increases D4 receptor mediated inhibitory effect on Na-K-ATPase activity in WKY cells, but not in SHR cells. CONCLUSION: Renal D3 receptor regulates the expression and function of D4 receptor in RPT cells via PLC /PKC signaling pathway, the loss of this interaction might be involved in the pathogenesis of hypertension.


Assuntos
Túbulos Renais Proximais/citologia , Receptores de Dopamina D3/metabolismo , Receptores de Dopamina D4/metabolismo , Animais , Benzopiranos/farmacologia , Células Cultivadas , Agonistas de Dopamina/farmacologia , Estrenos/farmacologia , Túbulos Renais Proximais/química , Túbulos Renais Proximais/metabolismo , Oxazinas/farmacologia , Fragmentos de Peptídeos/antagonistas & inibidores , Inibidores de Fosfodiesterase/farmacologia , Proteína Quinase C/antagonistas & inibidores , Pirrolidinonas/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptores de Dopamina D3/análise , Receptores de Dopamina D3/antagonistas & inibidores , Receptores de Dopamina D3/genética , Receptores de Dopamina D4/análise , Receptores de Dopamina D4/genética , Transdução de Sinais , ATPase Trocadora de Sódio-Potássio/metabolismo , Regulação para Cima
18.
Biochem Biophys Res Commun ; 474(3): 599-605, 2016 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-27018249

RESUMO

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 & dosagem
19.
J Am Heart Assoc ; 4(2)2015 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-25687731

RESUMO

BACKGROUND: Oxidative stress plays an important role in the pathogenesis of hypertension, especially in obesity-related hypertension. The natriuretic and antinatriuretic components of the renal renin angiotensin system (RAS) maintain sodium homeostasis and blood pressure. Here, we test the hypothesis that increased oxidative stress leads to the imbalance of RAS components and hypertension in obese Zucker rats. METHODS AND RESULTS: Lean and obese rats received vehicle or tempol, a superoxide dismutase mimetic in the drinking water for 4 weeks. Compared with vehicle-treated lean rats, vehicle-treated obese rats exhibited higher blood pressure and increased renal oxidative stress, accompanied by increased diuretic and natriuretic responses to AT1R antagonist (Candesartan) and AT2R agonist (CGP-42112A) and reduced diuretic and natriuretic response to MasR agonist (Ang-[1 to 7]). Moreover, obese rats had higher ACE, AT1R and AT2R, lower ACE2 and MasR expressions in the kidney. All of the above-mentioned abnormalities were reversed to some degree by tempol treatment. In primary cultures of renal proximal tubular (RPT) cells from lean and obese rats, tempol treatment also increased AT2R, ACE2, and MasR expressions but decreased AT1R and ACE expressions in obese rats. CONCLUSIONS: Taken together, our study indicated that the imbalance of renal RAS components was associated with increased oxidative stress in obese rats. Furthermore, antioxidant treatment with tempol reversed the imbalance of renal RAS components and led to diuresis and natriuresis, which, at least in part, explains the blood pressure-lowering effect of antioxidant supplementation in obesity-related hypertension.


Assuntos
Antioxidantes/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Óxidos N-Cíclicos/farmacologia , Hipertensão/etiologia , Estresse Oxidativo , Sistema Renina-Angiotensina/efeitos dos fármacos , Animais , Benzimidazóis/farmacologia , Compostos de Bifenilo , Óxidos N-Cíclicos/administração & dosagem , Modelos Animais de Doenças , Ingestão de Líquidos , Hipertensão/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Masculino , Obesidade/metabolismo , Oligopeptídeos/farmacologia , Peptidil Dipeptidase A/metabolismo , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas/metabolismo , Ratos , Ratos Zucker , Receptor Tipo 2 de Angiotensina/agonistas , Receptor Tipo 2 de Angiotensina/metabolismo , Receptores de Angiotensina/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Marcadores de Spin , Tetrazóis/farmacologia
20.
PLoS One ; 9(8): e105200, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25133686

RESUMO

Our previous study showed that besides mRNAs and microRNAs, there are DNA fragments within extracellular vesicles (EVs). The BCR/ABL hybrid gene, involved in the pathogenesis of chronic myeloid leukemia (CML), could be transferred from K562 EVs to neutrophils and decrease their phagocytic activity in vitro. Our present study provides evidence that BCR/ABL DNAs transferred from EVs have pathophysiological significance in vivo. Two months after injection of K562 EVs into the tail vein of Sprague-Dawley (SD) rats, they showed some characteristics of CML, e.g., feeble, febrile, and thin, with splenomegaly and neutrophilia but with reduced neutrophil phagocytic activity. These findings were also observed in immunodeficient NOD/SCID mice treated with K562 EVs; BCR/ABL mRNA and protein were found in their neutrophils. The administration of actinomycin D, an inhibitor of de novo mRNA synthesis, prevented the abnormalities caused by K562 EVs in NOD/SCID mice related to CML, including neutrophilia and bone marrow hyperplasia. As a specific inhibitor of tyrosine kinases, imatinib blocked the activity of tyrosine kinases and the expression of phospho-Crkl, induced by the de novo BCR/ABL protein caused by K562 EVs bearing BCR/ABL DNA. Our current study shows the pathophysiological significance of transferred tumor gene from EVs in vivo, which may represent an important mechanism for tumorigenesis, tumor progression, and metastasis.


Assuntos
DNA/metabolismo , Proteínas de Fusão bcr-abl/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/enzimologia , Animais , Linhagem Celular , Ensaio de Imunoadsorção Enzimática , Células HEK293 , Humanos , Immunoblotting , Células K562 , Camundongos , Camundongos SCID , Neutrófilos/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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