RESUMO
Macrophages are the major primary immune cells that mediate the inflammatory response. In this process, long non-coding RNAs (lncRNAs) play an important, yet largely unknown role. Therefore, utilizing several publicly available RNA sequencing datasets, we predicted and selected lncRNAs that are differentially expressed in M1 or M2 macrophages and involved in the inflammatory response. We identified SUGCT-AS1, which is a human macrophage-specific lncRNA whose expression is increased upon M1 macrophage stimulation. Conditioned media of SUGCT-AS1-depleted M1 macrophages induced an inflammatory phenotype of vascular smooth muscle cells, which included increased expression of inflammatory genes (IL1B and IL6), decreased contractile marker proteins (ACTA2 and SM22α), and increased cell migration. Depletion of SUGCT-AS1 promoted the expression and secretion of proinflammatory cytokines, such as TNF, IL1B, and IL6, in M1 macrophages, and transcriptomic analysis showed that SUGCT-AS1 has functions related to inflammatory responses and cytokines. Furthermore, we found that SUGCT-AS1 directly binds to hnRNPU and regulates its nuclear-cytoplasmic translocation. This translocation of hnRNPU altered the proportion of the MALT1 isoforms by regulating the alternative splicing of MALT1, a mediator of NF-κB signaling. Overall, our findings suggest that lncRNAs can be used for future studies on macrophage regulation. Moreover, they establish the SUGCT-AS1/hnRNPU/MALT1 axis, which is a novel inflammatory regulatory mechanism in macrophages.
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RNA Longo não Codificante , Humanos , RNA Longo não Codificante/genética , Interleucina-6/genética , Processamento Alternativo , Proteínas Contráteis , Citocinas/genética , MacrófagosRESUMO
The regeneration of myocardium following acute circulatory events remains a challenge, despite numerous efforts. Mesenchymal stem cells (MSCs) present a promising cell therapy option, but their differentiation into cardiomyocytes is a time-consuming process. Although it has been demonstrated that PSME4 degrades acetyl-YAP1, the role of PSME4 in the cardiac commitment of MSCs has not been fully elucidated. Here we reported the novel role of PSME4 in MSCs cardiac commitment. It was found that overnight treatment with apicidin in primary-cultured mouse MSCs led to rapid cardiac commitment, while MSCs from PSME4 knock-out mice did not undergo this process. Cardiac commitment was also observed using lentivirus-mediated PSME4 knockdown in immortalized human MSCs. Immunofluorescence and Western blot experiments revealed that YAP1 persisted in the nucleus of PSME4 knockdown cells even after apicidin treatment. To investigate the importance of YAP1 removal, MSCs were treated with shYAP1 and apicidin simultaneously. This combined treatment resulted in rapid YAP1 elimination and accelerated cardiac commitment. However, overexpression of acetylation-resistant YAP1 in apicidin-treated MSCs impeded cardiac commitment. In addition to apicidin, the universal effect of histone deacetylase (HDAC) inhibition on cardiac commitment was confirmed using tubastatin A and HDAC6 siRNA. Collectively, this study demonstrates that PSME4 is crucial for promoting the cardiac commitment of MSCs. HDAC inhibition acetylates YAP1 and facilitates its translocation to the nucleus, where it is removed by PSME4, promoting cardiac commitment. The failure of YAP1 to translocate or be eliminated from the nucleus results in the MSCs' inability to undergo cardiac commitment.
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The blots of control and docetaxel for caspase-9, caspase-3, caspase-8, Bcl-XL, and tubulin in the Figure 4f were reused from Figure 4 of our previous paper published in Journal of Urology in 2010 ( https://doi.org/10.1016/j.juro.2010.07.035 ).
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Although cell therapy is emerged for cardiac repair, its efficacy is modest by intracoronary infusion. Therefore, we established the intramyocardial delivery technique using a left ventricular (LV) mapping system (NOGA® XP) using 18 pigs. After adipose tissue-derived mesenchymal stem cells (ATSCs) were delivered intramyocardially to porcine infarcted heart, LV ejection fraction (EF) was increased, and LV chamber size was decreased. We proved the therapeutic effect of intramyocardial injection of ATSC through a LV mapping system in the porcine model for the first time in Korea. The adoption of this technique may accelerate the translation into a clinical application in the near future.
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Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Infarto do Miocárdio/terapia , Animais , Modelos Animais de Doenças , Vias de Administração de Medicamentos , Ecocardiografia , Coração/diagnóstico por imagem , Infarto do Miocárdio/diagnóstico por imagem , República da Coreia , Suínos , Função Ventricular Esquerda/fisiologiaRESUMO
RATIONALE: Small heterodimer partner (SHP; NR0B2) is an atypical orphan nuclear receptor that lacks a conventional DNA-binding domain. Through interactions with other transcription factors, SHP regulates diverse biological events, including glucose metabolism in liver. However, the role of SHP in adult heart diseases has not yet been demonstrated. OBJECTIVE: We aimed to investigate the role of SHP in adult heart in association with cardiac hypertrophy. METHODS AND RESULTS: The roles of SHP in cardiac hypertrophy were tested in primary cultured cardiomyocytes and in animal models. SHP-null mice showed a hypertrophic phenotype. Hypertrophic stresses repressed the expression of SHP, whereas forced expression of SHP blocked the development of hypertrophy in cardiomyocytes. SHP reduced the protein amount of Gata6 and, by direct physical interaction with Gata6, interfered with the binding of Gata6 to GATA-binding elements in the promoter regions of natriuretic peptide precursor type A. Metformin, an antidiabetic agent, induced SHP and suppressed cardiac hypertrophy. The metformin-induced antihypertrophic effect was attenuated either by SHP small interfering RNA in cardiomyocytes or in SHP-null mice. CONCLUSIONS: These results establish SHP as a novel antihypertrophic regulator that acts by interfering with GATA6 signaling. SHP may participate in the metformin-induced antihypertrophic response.
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Cardiomegalia/prevenção & controle , Fator de Transcrição GATA6/metabolismo , Miócitos Cardíacos/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Transdução de Sinais , Animais , Fator Natriurético Atrial/genética , Fator Natriurético Atrial/metabolismo , Sítios de Ligação , Cardiomegalia/genética , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Modelos Animais de Doenças , Fator de Transcrição GATA6/genética , Regulação da Expressão Gênica , Genótipo , Células HEK293 , Humanos , Masculino , Metformina/farmacologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Fenótipo , Regiões Promotoras Genéticas , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Receptores Citoplasmáticos e Nucleares/deficiência , Receptores Citoplasmáticos e Nucleares/genética , Transdução de Sinais/efeitos dos fármacos , TransfecçãoRESUMO
We examined whether a shift in macrophage phenotype could be therapeutic for myocardial infarction (MI). The mouse macrophage cell line RAW264.7 was stimulated with peptidoglycan (PGN), with or without 5-azacytidine (5AZ) treatment. MI was induced by ligation of the left anterior descending coronary artery in rats, and the rats were divided into two groups; a saline-injection group and a 5AZ-injection group (2.5 mg/kg/day, intraperitoneal injection). LV function was evaluated and immunohistochemical analyses were performed 2 weeks after MI. Cardiac fibrosis was induced by angiotensin II (AngII) infusion with or without 5AZ (5 mg/kg/day) in mice. Nitric oxide was produced by PGN, which was reduced by 77.87% after 5AZ treatment. Both induction of inducible nitric oxide synthase (iNOS) and iNOS promoter activity by PGN were inhibited by 5AZ. Ejection fraction (59.00 ± 8.03% versus 42.52 ± 2.58%), contractility (LV dP/dt-max, 8299.76 ± 411.56 mmHg versus 6610.36 ± 282.37 mmHg) and relaxation indices (LV dP/dt-min, -4661.37 ± 210.73 mmHg versus -4219.50 ± 162.98 mmHg) were improved after 5AZ administration. Cardiac fibrosis in the MI+5AZ was 8.14 ± 1.00%, compared with 14.93 ± 2.98% in the MI group (P < 0.05). Arginase-1(+)CD68(+) macrophages with anti-inflammatory phenotype were predominant in the infarct border zone of the MI+5AZ group, in comparison with the MI group. AngII-induced cardiac fibrosis was also attenuated after 5AZ administration. In cardiac fibroblasts, pro-fibrotic mediators and cell proliferation were increased by AngII, and these increases were attenuated after 5AZ treatment. 5AZ exerts its cardiac protective role through modulation of macrophages and cardiac fibroblasts.
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Azacitidina/farmacologia , Fibrose/prevenção & controle , Macrófagos/patologia , Infarto do Miocárdio/prevenção & controle , Óxido Nítrico/metabolismo , Disfunção Ventricular Esquerda/tratamento farmacológico , Animais , Antimetabólitos Antineoplásicos/farmacologia , Western Blotting , Células Cultivadas , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose/metabolismo , Fibrose/patologia , Técnicas Imunoenzimáticas , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Óxido Nítrico Sintase/metabolismo , Peptidoglicano/farmacologia , Fenótipo , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Disfunção Ventricular Esquerda/metabolismo , Disfunção Ventricular Esquerda/patologia , Remodelação VentricularRESUMO
Addressing age-related immunological defects through therapeutic interventions is essential for healthy aging, as the immune system plays a crucial role in controlling infections, malignancies, and in supporting tissue homeostasis and repair. In our study, we show that stimulating toll-like receptor 5 (TLR5) via mucosal delivery of a flagellin-containing fusion protein effectively extends the lifespan and enhances the healthspan of mice of both sexes. This enhancement in healthspan is evidenced by diminished hair loss and ocular lens opacity, increased bone mineral density, improved stem cell activity, delayed thymic involution, heightened cognitive capacity, and the prevention of pulmonary lung fibrosis. Additionally, this fusion protein boosts intestinal mucosal integrity by augmenting the surface expression of TLR5 in a certain subset of dendritic cells and increasing interleukin-22 (IL-22) secretion. In this work, we present observations that underscore the benefits of TLR5-dependent stimulation in the mucosal compartment, suggesting a viable strategy for enhancing longevity and healthspan.
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Longevidade , Receptor 5 Toll-Like , Animais , Camundongos , Flagelina/metabolismo , Mucosa Intestinal/metabolismo , Longevidade/genética , Pulmão/metabolismoRESUMO
BACKGROUND: Angiogenesis is the main therapeutic mechanism of cell therapy for cardiovascular diseases, but diabetes is reported to reduce the function and number of progenitor cells. Therefore, we studied the effect of streptozotocin-induced diabetes on the bone marrow-mesenchymal stem cell (MSC) function, and examined whether diabetes-impaired MSC could be rescued by pretreatment with oxytocin. RESULTS: MSCs were isolated and cultured from diabetic (DM) or non-diabetic (non-DM) rat, and proliferation rate was compared. DM-MSC was pretreated with oxytocin and compared with non-DM-MSC. Angiogenic capacity was estimated by tube formation and Matrigel plug assay, and therapeutic efficacy was studied in rat myocardial infarction (MI) model.The proliferation and angiogenic activity of DM-MSC were severely impaired but significantly improved by pretreatment with oxytocin. Krüppel-like factor 2 (KLF2), a critical angiogenic factor, was dramatically reduced in DM-MSC and significantly restored by oxytocin. In the Matrigel plug assay, vessel formation of DM-BMSCs was attenuated but was recovered by oxytocin. In rat MI model, DM-MSC injection did not ameliorate cardiac injury, whereas oxytocin-pretreated DM-MSC improved cardiac function and reduced fibrosis. CONCLUSIONS: Our results show that diabetes influenced MSC by reducing angiogenic capacity and therapeutic potential. We demonstrate the striking effect of oxytocin on stem cell dysfunction and suggest the use of oxytocin as a priming reagent in autologous stem cell therapy.
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Diabetes Mellitus Experimental/tratamento farmacológico , Células-Tronco Mesenquimais/efeitos dos fármacos , Infarto do Miocárdio/tratamento farmacológico , Neovascularização Fisiológica/efeitos dos fármacos , Ocitócicos/farmacologia , Ocitocina/farmacologia , Pele/irrigação sanguínea , Animais , Biomarcadores/metabolismo , Glicemia/metabolismo , Células Cultivadas , Colágeno/química , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Combinação de Medicamentos , Expressão Gênica , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Laminina/química , Masculino , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Camundongos Nus , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Proteoglicanas/química , Ratos , Ratos Sprague-Dawley , Pele/efeitos dos fármacos , Pele/patologia , EstreptozocinaRESUMO
Myocardial infarction (MI) is a major cause of death worldwide. After the occurrence of MI, the heart frequently undergoes serious pathological remodeling, leading to excessive dilation, electrical disconnection between cardiac cells, and fatal functional damage. Hence, extensive efforts have been made to suppress pathological remodeling and promote the repair of the infarcted heart. In this study, we developed a hydrogel cardiac patch that can provide mechanical support, electrical conduction, and tissue adhesiveness to aid in the recovery of an infarcted heart function. Specifically, we developed a conductive and adhesive hydrogel (CAH) by combining the two-dimensional titanium carbide (Ti3C2Tx) MXene with natural biocompatible polymers [i.e., gelatin and dextran aldehyde (dex-ald)]. The CAH was formed within 250 s of mixing the precursor solution and could be painted. The hydrogel containing 3.0 mg/mL MXene, 10% gelatin, and 5% dex-ald exhibited appropriate material characteristics for cardiac patch applications, including a uniform distribution of MXene, a high electrical conductivity (18.3 mS/cm), cardiac tissue-like elasticity (30.4 kPa), strong tissue adhesion (6.8 kPa), and resistance to various mechanical deformations. The CAH was cytocompatible and induced cardiomyocyte (CM) maturation in vitro, as indicated by the upregulation of connexin 43 expression and a faster beating rate. Furthermore, CAH could be painted onto the heart tissue and remained stably adhered to the beating epicardium. In vivo animal studies revealed that CAH cardiac patch treatment significantly improved cardiac function and alleviated the pathological remodeling of an infarcted heart. Thus, we believe that our MXene-based CAH can potentially serve as a promising platform for the effective repair of various electroactive tissues including the heart, muscle, and nerve tissues.
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Hidrogéis , Infarto do Miocárdio , Animais , Hidrogéis/farmacologia , Gelatina/metabolismo , Adesivos/farmacologia , Infarto do Miocárdio/patologia , Miócitos Cardíacos , Polímeros/farmacologia , Condutividade ElétricaRESUMO
Atherosclerosis, the leading cause of death, is a vascular disease of chronic inflammation. We recently showed that angiopoietin-like 4 (ANGPTL4) promotes cardiac repair by suppressing pathological inflammation. Given the fundamental contribution of inflammation to atherosclerosis, we assessed the role of ANGPTL4 in the development of atherosclerosis and determined whether ANGPTL4 regulates atherosclerotic plaque stability. We injected ANGPTL4 protein twice a week into atherosclerotic Apoe-/- mice and analyzed the atherosclerotic lesion size, inflammation, and plaque stability. In atherosclerotic mice, ANGPTL4 reduced atherosclerotic plaque size and vascular inflammation. In the atherosclerotic lesions and fibrous caps, the number of α-SMA(+), SM22α(+), and SM-MHC(+) cells was higher, while the number of CD68(+) and Mac2(+) cells was lower in the ANGPTL4 group. Most importantly, the fibrous cap was significantly thicker in the ANGPTL4 group than in the control group. Smooth muscle cells (SMCs) isolated from atherosclerotic aortas showed significantly increased expression of CD68 and Krüppel-like factor 4 (KLF4), a modulator of the vascular SMC phenotype, along with downregulation of α-SMA, and these changes were attenuated by ANGPTL4 treatment. Furthermore, ANGPTL4 reduced TNFα-induced NADPH oxidase 1 (NOX1), a major source of reactive oxygen species, resulting in the attenuation of KLF4-mediated SMC phenotypic changes. We showed that acute myocardial infarction (AMI) patients with higher levels of ANGPTL4 had fewer vascular events than AMI patients with lower levels of ANGPTL4 (p < 0.05). Our results reveal that ANGPTL4 treatment inhibits atherogenesis and suggest that targeting vascular stability and inflammation may serve as a novel therapeutic strategy to prevent and treat atherosclerosis. Even more importantly, ANGPTL4 treatment inhibited the phenotypic changes of SMCs into macrophage-like cells by downregulating NOX1 activation of KLF4, leading to the formation of more stable plaques.
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Aterosclerose , Placa Aterosclerótica , Camundongos , Animais , Placa Aterosclerótica/patologia , Fator 4 Semelhante a Kruppel , Músculo Liso Vascular , Regulação para Baixo , Camundongos Knockout para ApoE , Aterosclerose/patologia , Fenótipo , Miócitos de Músculo Liso/metabolismo , Inflamação/metabolismo , Camundongos Endogâmicos C57BL , Células CultivadasRESUMO
BACKGROUND: Thin-cap fibroatheroma is a rupture-prone vulnerable plaque that leads to acute coronary syndrome (ACS). However, its underlying mechanisms are not fully understood. Several studies have investigated the clinical association between angiopoietin-like protein 4 (ANGPTL4) and coronary artery disease. Therefore, this study aimed to investigate the correlation of plasma ANGPTL4 in culprit lesion of ACS patients using intravascular ultrasound (IVUS) and virtual-histology IVUS (VH-IVUS). METHODS: Fifty patients newly diagnosed with ACS between March to September 2021 were selected. Blood samples for baseline laboratory tests, including ANGPTL4, were collected before percutaneous coronary intervention (PCI), and all pre- and post-PCI IVUS examinations were performed of the culprit lesions. RESULTS: Linear regression analysis between plasma ANGPTL4 and grayscale IVUS/VH-IVUS parameters revealed that plasma ANGPTL4 was strongly correlated with the necrotic core (NC) of the minimal lumen site (r = -0.666, p = 0.003) and largest NC site (r = -0.687, p < 0.001), and patients with lower plasma ANGPTL4 levels showed a significantly higher proportion of TFCA. CONCLUSION: The present study further demonstrated the protective role of ANGPTL4 in the spectrum of atherosclerotic development in patients with ACS by culprit lesion morphology analysis using IVUS and VH-IVUS.
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Síndrome Coronariana Aguda , Doença da Artéria Coronariana , Intervenção Coronária Percutânea , Placa Aterosclerótica , Humanos , Síndrome Coronariana Aguda/diagnóstico por imagem , Síndrome Coronariana Aguda/patologia , Angiopoietinas , Angiografia Coronária , Doença da Artéria Coronariana/patologia , Vasos Coronários/diagnóstico por imagem , Necrose/patologia , Placa Aterosclerótica/diagnóstico por imagem , Placa Aterosclerótica/patologia , Ultrassonografia de IntervençãoRESUMO
Heart failure is a leading cause of death and is often accompanied by activation of quiescent cardiac myofibroblasts, which results in cardiac fibrosis. In this study, we aimed to identify novel circular RNAs that regulate cardiac fibrosis. We applied transverse aortic constriction (TAC) for 1, 4, and 8 weeks in mice. RNA sequencing datasets were obtained from cardiac fibroblasts isolated by use of a Langendorff apparatus and then further processed by use of selection criteria such as differential expression and conservation in species. CircSMAD4 was upregulated by TAC in mice or by transforming growth factor (TGF)-ß1 in primarily cultured human cardiac fibroblasts. Delivery of si-circSMAD4 attenuated myofibroblast activation and cardiac fibrosis in mice treated with isoproterenol (ISP). si-circSmad4 significantly reduced cardiac fibrosis and remodeling at 8 weeks. Mechanistically, circSMAD4 acted as a sponge against the microRNA miR-671-5p in a sequence-specific manner. miR-671-5p was downregulated during myofibroblast activation and its mimic form attenuated cardiac fibrosis. miR-671-5p mimic destabilized fibroblast growth factor receptor 2 (FGFR2) mRNA in a sequence-specific manner and interfered with the fibrotic action of FGFR2. The circSMAD4-miR-671-5p-FGFR2 pathway is involved in the differentiation of cardiac myofibroblasts and thereby the development of cardiac fibrosis.
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We synthesized a novel hydroxamate-based pan-histone deacetylase inhibitor (HDACI), CG200745 {(E)-2-(Naphthalen-1-yloxymethyl)-oct-2-enedioic acid 1-[(3-dimethylamino-propyl)-amide] 8-hydroxyamide]}. Like other inhibitors, for example vorinostat and belinostat, CG200745 has the hydroxamic acid moiety to bind zinc at the bottom of catalytic pocket. Firstly, we analyzed its inhibitory activity against histone deacetylase (HDAC) in hormone-dependent LNCaP cells and hormone-independent DU145 and PC3 cells. CG200745 inhibited deacetylation of histone H3 and tubulin as much as vorinostat and belinostat did. CG200745 also inhibited growth of prostate cancer cells, increased sub-G1 population, and activated caspase-9, -3 and -8 in LNCaP, DU145 and PC3 cells. These results indicate that CG200745 induces apoptosis. Next, we examined the effect of CG200745 on cell death induced by docetaxel in DU145 cells in vitro and in vivo. Compared to mono-treatment with each drug, pre-treatment of DU145 cells with docetaxel followed by CG200745 showed synergistic cytotoxicity, and increased the apoptotic sub-G1 population, caspase activation, and tubulin acetylation. Moreover, the combination treatment decreased Mcl-1 and Bcl-(XL). Docetaxel and CG200745 combination reduced tumor size in the DU145 xenograft model. These preclinical results show that combination treatment with docetaxel and new HDACI, CG200745, potentiated anti-tumor effect in hormone-refractory prostate cancer (HRPC) cells via activation of apoptosis.
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Antineoplásicos/uso terapêutico , Inibidores de Histona Desacetilases/uso terapêutico , Ácidos Hidroxâmicos/uso terapêutico , Naftalenos/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Taxoides/uso terapêutico , Proteína bcl-X/metabolismo , Acetilação/efeitos dos fármacos , Animais , Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Docetaxel , Sinergismo Farmacológico , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/farmacologia , Histonas/metabolismo , Humanos , Ácidos Hidroxâmicos/química , Ácidos Hidroxâmicos/farmacologia , Concentração Inibidora 50 , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteína de Sequência 1 de Leucemia de Células Mieloides , Naftalenos/química , Naftalenos/farmacologia , Neoplasias da Próstata/patologia , Taxoides/farmacologia , Tubulina (Proteína)/metabolismoRESUMO
Oxytocin stimulates the cardiomyogenesis of embryonic stem cells and adult cardiac stem cells. We previously reported that oxytocin has a promigratory effect on umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs). In this study, UCB-MSCs were cultured with oxytocin and examined for their therapeutic effect in an infarcted heart. UCB-MSCs were pretreated with 100 nM oxytocin and cardiac markers were assessed by immunofluorescence staining. Next, oxytocin-supplemented USC-MSCs (OT-USCs) were cocultured with hypoxia/reoxygenated neonatal rat cardiomyocytes and cardiac markers and dye transfer were then examined. For the in vivo study, ischemia/reperfusion was induced in rats, and phosphate-buffered saline (group 1), 1-day OT-USCs (group 2), or 7-day OT-USCs (group 3) were injected into the infarcted myocardium. Two weeks after injection, histological changes and cardiac function were examined. UCB-MSCs expressed connexin 43 (Cnx43), cardiac troponin I (cTnI), and α-sarcomeric actin (α-SA) after oxytocin supplementation and coculture with cardiomyocytes. Functional gap junction formation was greater in group 3 than in groups 1 and 2. Cardiac fibrosis and macrophage infiltration were lower in group 3 than in group 2. Restoration of Cnx43 expression was greater in group 3 than in group 2. Cnx43- and cTnI-positive OT-USCs in the peri-infarct zone were observed in group 2 and more frequently in group 3. The ejection fraction (EF) was increased in groups 2 and 3 in 2 weeks. The improved EF was sustained for 4 weeks only in group 3. Our findings suggest that the supplementation of UCB-MSCs with oxytocin can contribute to the cardiogenic potential for cardiac repair.
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Sangue Fetal/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Miócitos Cardíacos/efeitos dos fármacos , Ocitocina/farmacologia , Cicatrização/efeitos dos fármacos , Animais , Transplante de Células-Tronco de Sangue do Cordão Umbilical , Sangue Fetal/citologia , Humanos , Masculino , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/patologia , Traumatismo por Reperfusão Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/cirurgia , Miócitos Cardíacos/patologia , Ratos , Ratos Sprague-DawleyRESUMO
CRISPR/Cas9-mediated gene-editing technology has gained attention as a new therapeutic method for intractable diseases. However, the use of CRISPR/Cas9 for cardiac conditions such as myocardial infarction remains challenging due to technical and biological barriers, particularly difficulties in delivering the system and targeting genes in the heart. In the present study, we demonstrated the in vivo efficacy of the CRISPR/Cas9 magnetoplexes system for therapeutic genome editing in myocardial infarction. First, we developed CRISPR/Cas9 magnetoplexes that magnetically guided CRISPR/Cas9 system to the heart for efficient in vivo therapeutic gene targeting during heart failures. We then demonstrated that the in vivo gene targeting of miR34a via these CRISPR/Cas9 magnetoplexes in a mouse model of myocardial infarction significantly improved cardiac repair and regeneration to facilitate improvements in cardiac function. These results indicated that CRISPR/Cas9 magnetoplexes represent an effective in vivo therapeutic gene-targeting platform in the myocardial infarction of heart, and that this strategy may be applicable for the treatment of a broad range of cardiac failures.
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Edição de Genes , Infarto do Miocárdio , Animais , Sistemas CRISPR-Cas/genética , Edição de Genes/métodos , Marcação de Genes , Terapia Genética/métodos , Camundongos , Infarto do Miocárdio/genética , Infarto do Miocárdio/terapiaRESUMO
Implantable bioelectrodes enable precise recording or stimulation of electrical signals with living tissues in close contact. However, their performance is frequently compromised owing to inflammatory tissue reactions, which macrophages either induce or resolve by polarizing to an inflammatory (M1) or noninflammatory (M2) phenotype, respectively. Thus, we aimed to fabricate biocompatible and functional implantable conductive polymer bioelectrodes with optimal topography for the modulation of macrophage responses. To this end, we produced heparin-doped polypyrrole (PPy/Hep) electrodes of different surface roughness, with Ra values from 5.5 to 17.6 nm, by varying the charge densities during electrochemical synthesis. In vitro culture revealed that macrophages on rough PPy/Hep electrodes preferentially polarized to noninflammatory phenotypes. In particular, PPy/Hep-900 (Ra = 14 nm) was optimal with respect to electrochemical properties and the suppression of inflammatory M1 polarization. In vivo implantation indicated that PPy/Hep-900 significantly reduced macrophage recruitment, suppressed inflammatory polarization, and mitigated fibrotic tissue formation. In addition, the implanted PPy/Hep-900 electrodes could successfully record electrocardiographic signals for up to 10 days without substantial decreases in sensitivity, while other electrodes substantially lost their signal sensitivity during implantation. Altogether, we demonstrate that modulating the surface features of PPy/Hep can benefit the design and applications of high-performance and high-biocompatibility bioelectrodes.
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Polímeros , Pirróis , Polímeros/química , Pirróis/química , Condutividade Elétrica , Macrófagos , Eletrodos ImplantadosRESUMO
Intensive research has focused on minimizing the infarct area and stimulating endogenous regeneration after myocardial infarction. Our group previously elucidated that apicidin, a histone deacetylase (HDAC) inhibitor, robustly accelerates the cardiac commitment of naïve mesenchymal stem cells (MSCs) through acute loss of YAP1. Here, we propose the novel regulation of YAP1 in MSCs. We found that acute loss of YAP1 after apicidin treatment resulted in the mixed effects of transcriptional arrest and proteasomal degradation. Subcellular fractionation revealed that YAP1 was primarily localized in the cytoplasm. YAP1 was acutely relocalized into the nucleus and underwent proteasomal degradation. Interestingly, phosphor-S127 YAP1 was shuttled into the nucleus, suggesting that a mechanism other than phosphorylation governed the subcellular localization of YAP1. Apicidin successfully induced acetylation and subsequent dissociation of YAP1 from 14-3-3, an essential molecule for cytoplasmic restriction. HDAC6 regulated both acetylation and subcellular localization of YAP1. An acetylation-dead mutant of YAP1 retarded nuclear redistribution upon apicidin treatment. We failed to acquire convincing evidence for polyubiquitination-dependent degradation of YAP1, suggesting that a polyubiquitination-independent regulator determined YAP1 fate. Nuclear PSME4, a subunit of the 26 S proteasome, recognized and degraded acetyl YAP1 in the nucleus. MSCs from PSME4-null mice were injected into infarcted heart, and aberrant sudden death was observed. Injection of immortalized human MSCs after knocking down PSME4 failed to improve either cardiac function or the fibrotic scar area. Our data suggest that acetylation-dependent proteasome subunit PSME4 clears acetyl-YAP1 in response to apicidin treatment in the nucleus of MSCs.
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Mast cells are multifunctional cells containing various mediators, such as cytokines, tryptase, and histamine, and they have been identified in infarct myocardium. Here, we elucidated the roles of mast cells in a myocardial infarction (MI) rat model. We studied the physiological and functional roles of mast cell granules (MCGs), isolated from rat peritoneal fluid, on endothelial cells, neonatal cardiomyocytes, and infarct heart (1-hour occlusion of left coronary artery followed by reperfusion). The number of mast cells had two peak time points of appearance in the infarct region at 1day and 21days after MI induction in rats (p<0.05 in each compared with sham-operated heart). Simultaneous injection of an optimal dose of MCGs modulated the microenvironment and resulted in the increased infiltration of macrophages and decreased apoptosis of cardiomyocytes without change in the mast cell number in infarct myocardium. Moreover, MCG injection attenuated the progression of MI through angiogenesis and preserved left ventricular function after MI. MCG-treated cardiomyocytes were more resistant to hypoxic injury through phosphorylation of Akt, and MCG-treated endothelial cells showed enhanced migration and tube formation. We have shown that MCGs have novel cardioprotective roles in MI via the prolonged survival of cardiomyocytes and the induction of angiogenesis.
Assuntos
Grânulos Citoplasmáticos/metabolismo , Mastócitos/metabolismo , Infarto do Miocárdio/metabolismo , Animais , Apoptose/fisiologia , Western Blotting , Hipóxia Celular/fisiologia , Células Cultivadas , Hemodinâmica , Humanos , Masculino , Mastócitos/fisiologia , Infarto do Miocárdio/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
A Gram-negative, red-pigment-producing marine bacterial strain, designated S1-1, was isolated from the tidal flat sediment of the Yellow Sea, Korea. On the basis of phenotypic, phylogenetic, and genetic data, strain S1-1 (KCTC 11448BP) represented a new species of the genus Zooshikella. Thus, we propose the name Zooshikella rubidus sp. nov. Liquid chromatography and mass spectrometry of the red pigments produced by strain S1-1 revealed that the major metabolic compounds were prodigiosin and cycloprodigiosin. In addition, this organism produced six minor prodigiosin analogues, including two new structures that were previously unknown. To our knowledge, this is the first description of a microorganism that simultaneously produces prodigiosin and cycloprodigiosin as two major metabolites. Both prodigiosin and cycloprodigiosin showed antimicrobial activity against several microbial species. These bacteria were approximately 1.5-fold more sensitive to cycloprodigiosin than to prodigiosin. The metabolites also showed anticancer activity against human melanoma cells, which showed significantly more sensitivity to prodigiosin than to cycloprodigiosin. The secondary metabolite profiles of strain S1-1 and two reference bacterial strains were compared by liquid chromatography-mass spectrometry. Multivariate statistical analyses based on secondary metabolite profiles by liquid chromatography-mass spectrometry indicated that the metabolite profile of strain S1-1 could clearly be distinguished from those of two phylogenetically related, prodigiosin-producing bacterial strains.
Assuntos
Gammaproteobacteria/metabolismo , Indóis/metabolismo , Pigmentos Biológicos/biossíntese , Prodigiosina/análogos & derivados , Prodigiosina/biossíntese , Pirróis/metabolismo , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Linhagem Celular Tumoral/efeitos dos fármacos , Cromatografia Líquida , DNA Bacteriano/análise , DNA Ribossômico/química , DNA Ribossômico/genética , Gammaproteobacteria/genética , Genótipo , Humanos , Espectrometria de Massas , Melanoma , Dados de Sequência Molecular , Fenótipo , Filogenia , Pigmentos Biológicos/genética , RNA Bacteriano/análise , RNA Ribossômico 16S/genética , República da Coreia , Água do Mar/microbiologiaRESUMO
Superparamagnetic iron oxide nanoparticles (SPION) were used to transfer gene into umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) in this study. This novel transfection method using SPION is safe and effective to UCB-MSCs, and would be a tool for genetic optimization with a significant potential for cell tracing.