RESUMO

BACKGROUND: Many patients with heart failure with preserved ejection fraction have metabolic syndrome and develop exercise-induced pulmonary hypertension (EIPH). Increases in pulmonary vascular resistance in patients with heart failure with preserved ejection fraction portend a poor prognosis; this phenotype is referred to as combined precapillary and postcapillary pulmonary hypertension (CpcPH). Therapeutic trials for EIPH and CpcPH have been disappointing, suggesting the need for strategies that target upstream mechanisms of disease. This work reports novel rat EIPH models and mechanisms of pulmonary vascular dysfunction centered around the transcriptional repression of the soluble guanylate cyclase (sGC) enzyme in pulmonary artery (PA) smooth muscle cells. METHODS: We used obese ZSF-1 leptin-receptor knockout rats (heart failure with preserved ejection fraction model), obese ZSF-1 rats treated with SU5416 to stimulate resting pulmonary hypertension (obese+sugen, CpcPH model), and lean ZSF-1 rats (controls). Right and left ventricular hemodynamics were evaluated using implanted catheters during treadmill exercise. PA function was evaluated with magnetic resonance imaging and myography. Overexpression of nuclear factor Y α subunit (NFYA), a transcriptional enhancer of sGC ß1 subunit (sGCß1), was performed by PA delivery of adeno-associated virus 6. Treatment groups received the SGLT2 inhibitor empagliflozin in drinking water. PA smooth muscle cells from rats and humans were cultured with palmitic acid, glucose, and insulin to induce metabolic stress. RESULTS: Obese rats showed normal resting right ventricular systolic pressures, which significantly increased during exercise, modeling EIPH. Obese+sugen rats showed anatomic PA remodeling and developed elevated right ventricular systolic pressure at rest, which was exacerbated with exercise, modeling CpcPH. Myography and magnetic resonance imaging during dobutamine challenge revealed PA functional impairment of both obese groups. PAs of obese rats produced reactive oxygen species and decreased sGCß1 expression. Mechanistically, cultured PA smooth muscle cells from obese rats and humans with diabetes or treated with palmitic acid, glucose, and insulin showed increased mitochondrial reactive oxygen species, which enhanced miR-193b-dependent RNA degradation of nuclear factor Y α subunit (NFYA), resulting in decreased sGCß1-cGMP signaling. Forced NYFA expression by adeno-associated virus 6 delivery increased sGCß1 levels and improved exercise pulmonary hypertension in obese+sugen rats. Treatment of obese+sugen rats with empagliflozin improved metabolic syndrome, reduced mitochondrial reactive oxygen species and miR-193b levels, restored NFYA/sGC activity, and prevented EIPH. CONCLUSIONS: In heart failure with preserved ejection fraction and CpcPH models, metabolic syndrome contributes to pulmonary vascular dysfunction and EIPH through enhanced reactive oxygen species and miR-193b expression, which downregulates NFYA-dependent sGCß1 expression. Adeno-associated virus-mediated NFYA overexpression and SGLT2 inhibition restore NFYA-sGCß1-cGMP signaling and ameliorate EIPH.

Assuntos

Fator de Ligação a CCAAT/metabolismo , Insuficiência Cardíaca/etiologia , Hipertensão Pulmonar/complicações , Hipertensão Pulmonar/etiologia , Síndrome Metabólica/genética , Síndrome Metabólica/metabolismo , MicroRNAs/genética , Espécies Reativas de Oxigênio/metabolismo , Guanilil Ciclase Solúvel/genética , Animais , Animais Geneticamente Modificados , Biomarcadores , Modelos Animais de Doenças , Suscetibilidade a Doenças , Exercício Físico , Regulação da Expressão Gênica , Insuficiência Cardíaca/diagnóstico , Humanos , Síndrome Metabólica/complicações , Mitocôndrias Cardíacas , Miócitos de Músculo Liso/metabolismo , Fenótipo , Ratos , Transdução de Sinais , Estresse Fisiológico , Volume Sistólico , Disfunção Ventricular Direita

RESUMO

RATIONALE: Unproven theories abound regarding the long-range uptake and endocrine activity of extracellular blood-borne microRNAs into tissue. In pulmonary hypertension (PH), microRNA-210 (miR-210) in pulmonary endothelial cells promotes disease, but its activity as an extracellular molecule is incompletely defined. OBJECTIVE: We investigated whether chronic and endogenous endocrine delivery of extracellular miR-210 to pulmonary vascular endothelial cells promotes PH. METHODS AND RESULTS: Using miR-210 replete (wild-type [WT]) and knockout mice, we tracked blood-borne miR-210 using bone marrow transplantation and parabiosis (conjoining of circulatory systems). With bone marrow transplantation, circulating miR-210 was derived predominantly from bone marrow. Via parabiosis during chronic hypoxia to induce miR-210 production and PH, miR-210 was undetectable in knockout-knockout mice pairs. However, in plasma and lung endothelium, but not smooth muscle or adventitia, miR-210 was observed in knockout mice of WT-knockout pairs. This was accompanied by downregulation of miR-210 targets ISCU (iron-sulfur assembly proteins)1/2 and COX10 (cytochrome c oxidase assembly protein-10), indicating endothelial import of functional miR-210. Via hemodynamic and histological indices, knockout-knockout pairs were protected from PH, whereas knockout mice in WT-knockout pairs developed PH. In particular, pulmonary vascular engraftment of miR-210-positive interstitial lung macrophages was observed in knockout mice of WT-knockout pairs. To address whether engrafted miR-210-positive myeloid or lymphoid cells contribute to paracrine miR-210 delivery, we studied miR-210 knockout mice parabiosed with miR-210 WT; Cx3cr1 knockout mice (deficient in myeloid recruitment) or miR-210 WT; Rag1 knockout mice (deficient in lymphocytes). In both pairs, miR-210 knockout mice still displayed miR-210 delivery and PH, thus demonstrating a pathogenic endocrine delivery of extracellular miR-210. CONCLUSIONS: Endogenous blood-borne transport of miR-210 into pulmonary vascular endothelial cells promotes PH, offering fundamental insight into the systemic physiology of microRNA activity. These results also describe a platform for RNA-mediated crosstalk in PH, providing an impetus for developing blood-based miR-210 technologies for diagnosis and therapy in this disease.

Assuntos

Endotélio Vascular/metabolismo , Hipertensão Pulmonar/metabolismo , Pulmão/irrigação sanguínea , MicroRNAs/metabolismo , Animais , Transplante de Medula Óssea , Receptor 1 de Quimiocina CX3C/genética , Receptor 1 de Quimiocina CX3C/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Endotélio Vascular/fisiopatologia , Feminino , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/fisiopatologia , Hipóxia/complicações , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/sangue , MicroRNAs/genética , Parabiose , Transdução de Sinais

RESUMO

There is no therapy currently available for fully repairing articular cartilage lesions. Our laboratory has recently developed a visible light-activatable methacrylated gelatin (mGL) hydrogel, with the potential for cartilage regeneration. In this study, we further optimized mGL scaffolds by supplementing methacrylated hyaluronic acid (mHA), which has been shown to stimulate chondrogenesis via activation of critical cellular signalling pathways. We hypothesized that the introduction of an optimal ratio of mHA would enhance the biological properties of mGL scaffolds and augment chondrogenesis of human bone marrow-derived mesenchymal stem cells (hBMSCs). To test this hypothesis, hybrid scaffolds consisting of mGL and mHA at different weight ratios were fabricated with hBMSCs encapsulated at 20 × 106  cells/ml and maintained in a chondrogenesis-promoting medium. The chondrogenenic differentiation of hBMSCs, within different scaffolds, was estimated after 8 weeks of culture. Our results showed that mGL/mHA at a 9:1 (%, w/v) ratio resulted in the lowest hBMSC hypertrophy and highest glycosaminoglycan production, with a slightly increased volume of the entire construct. The applicability of this optimally designed mGL/mHA hybrid scaffold for cartilage repair was then examined in vivo. A full-thickness cylindrical osteochondral defect was surgically created in the rabbit femoral condyle, and a three-dimensional cell-biomaterial construct was fabricated by in situ photocrosslinking to fully fill the lesion site. The results showed that implantation of the mGL/mHA (9:1) construct resulted in both cartilage and subchondral bone regeneration after 12 weeks, supporting its use as a promising scaffold for repair and resurfacing of articular cartilage defects, in the clinical setting.

Assuntos

Cartilagem Articular/patologia , Reagentes de Ligações Cruzadas/química , Gelatina/química , Ácido Hialurônico/química , Luz , Alicerces Teciduais/química , Cicatrização , Animais , Contagem de Células , Sobrevivência Celular , Condrogênese , Regulação da Expressão Gênica , Glicosaminoglicanos/metabolismo , Humanos , Hidrogéis/química , Hipertrofia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Metacrilatos/química , Coelhos

RESUMO

ABSTRACT Celosia argentea L., Amaranthaceae, is widely used as traditional medicine with a long history in China. It is a unique source of Semen Celosiae whose contributions include purging the hepatic pathogenic fire, improving eyesight, and treating other eye diseases. Over 79 compounds from this plant were isolated and identified, mainly including saponins, peptides, phenols, fatty acids, and amino acids, of which saponins have been considered as the characteristic and active constituents of Celosia argentea. Experimental evidences manifested that Celosia argentea, with its active compounds, possesses wide-reaching biological activities such as hepatoprotection, tumor treatment, anti-diarrhea, anti-diabetes, anti-oxidant, anti-hypertension, and for treatment of a number of eye diseases. The objective of the study was to provide an overview of the ethno-pharmacology, chemical constituents, pharmacology, and related clinical applications of Celosia argentea, and to reveal their therapeutic potentials, and secure an evidence base for further research works on Celosia argentea.