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Calcific aortic valve disease (CAVD) primarily involves osteogenic differentiation in human aortic valve interstitial cells (hVICs). Schisandrol B (SolB), a natural bioactive constituent, has known therapeutic effects on inflammatory and fibrotic disorders. However, its impact on valve calcification has not been reported. We investigated the effect of SolB on osteogenic differentiation of hVICs. Transcriptome sequencing was used to analyze potential molecular pathways affected by SolB treatment. The study also included an in vivo murine model using aortic valve wire injury surgery to observe SolB's effect on valve calcification. SolB inhibited the osteogenic differentiation of hVICs, reversing the increase in calcified nodule formation and osteogenic proteins. In the murine model, SolB significantly decreased the peak velocity of the aortic valve post-injury and reduced valve fibrosis and calcification. Transcriptome sequencing identified the p53 signaling pathway as a key molecular target of SolB, demonstrating its role as a molecular glue in the mouse double minute 2 (MDM2)-p53 interaction, thereby promoting p53 ubiquitination and degradation, which further inhibited p53-related inflammatory and senescence response. These results highlighted therapeutic potential of SolB for CAVD via inhibiting p53 signaling pathway and revealed a new molecular mechanism of SolB which provided a new insight of theraputic mechanism for CAVD.
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Estenosis de la Válvula Aórtica , Válvula Aórtica , Calcinosis , Ciclooctanos , Lignanos , Ratones Endogámicos C57BL , Proteína p53 Supresora de Tumor , Animales , Lignanos/farmacología , Válvula Aórtica/patología , Válvula Aórtica/efectos de los fármacos , Válvula Aórtica/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Humanos , Calcinosis/tratamiento farmacológico , Calcinosis/patología , Calcinosis/metabolismo , Ciclooctanos/farmacología , Ratones , Estenosis de la Válvula Aórtica/tratamiento farmacológico , Estenosis de la Válvula Aórtica/patología , Osteogénesis/efectos de los fármacos , Masculino , Transducción de Señal/efectos de los fármacos , Senescencia Celular/efectos de los fármacos , Inflamación/tratamiento farmacológico , Inflamación/patología , Inflamación/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Células Cultivadas , Diferenciación Celular/efectos de los fármacos , Modelos Animales de EnfermedadRESUMEN
BACKGROUND AND AIMS: Valve interstitial cells (VICs) undergo a transition to intermediate state cells before ultimately transforming into the osteogenic cell population, which is a pivotal cellular process in calcific aortic valve disease (CAVD). Herein, this study successfully delineated the stages of VIC osteogenic transformation and elucidated a novel key regulatory role of lumican (LUM) in this process. METHODS: Single-cell RNA-sequencing (scRNA-seq) from nine human aortic valves was used to characterize the pathological switch process and identify key regulatory factors. The in vitro, ex vivo, in vivo, and double knockout mice were constructed to further unravel the calcification-promoting effect of LUM. Moreover, the multi-omic approaches were employed to analyse the molecular mechanism of LUM in CAVD. RESULTS: ScRNA-seq successfully delineated the process of VIC pathological transformation and highlighted the significance of LUM as a novel molecule in this process. The pro-calcification role of LUM is confirmed on the in vitro, ex vivo, in vivo level, and ApoE-/-//LUM-/- double knockout mice. The LUM induces osteogenesis in VICs via activation of inflammatory pathways and augmentation of cellular glycolysis, resulting in the accumulation of lactate. Subsequent investigation has unveiled a novel LUM driving histone modification, lactylation, which plays a role in facilitating valve calcification. More importantly, this study has identified two specific sites of histone lactylation, namely, H3K14la and H3K9la, which have been found to facilitate the process of calcification. The confirmation of these modification sites' association with the expression of calcific genes Runx2 and BMP2 has been achieved through ChIP-PCR analysis. CONCLUSIONS: The study presents novel findings, being the first to establish the involvement of lumican in mediating H3 histone lactylation, thus facilitating the development of aortic valve calcification. Consequently, lumican would be a promising therapeutic target for intervention in the treatment of CAVD.
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BACKGROUND: Macrophages play a crucial role in the development of cardiac fibrosis (CF). Although our previous studies have shown that glycogen metabolism plays an important role in macrophage inflammatory phenotype, the role and mechanism of modifying macrophage phenotype by regulating glycogen metabolism and thereby improving CF have not been reported. METHODS: Here, we took glycogen synthetase kinase 3ß (GSK3ß) as the target and used its inhibitor NaW to enhance macrophage glycogen metabolism, transform M2 phenotype into anti-fibrotic M1 phenotype, inhibit fibroblast activation into myofibroblasts, and ultimately achieve the purpose of CF treatment. RESULTS: NaW increases the pH of macrophage lysosome through transmembrane protein 175 (TMEM175) and caused the release of Ca2+ through the lysosomal Ca2+ channel mucolipin-2 (Mcoln2). At the same time, the released Ca2+ activates TFEB, which promotes glucose uptake by M2 and further enhances glycogen metabolism. NaW transforms the M2 phenotype into the anti-fibrotic M1 phenotype, inhibits fibroblasts from activating myofibroblasts, and ultimately achieves the purpose of treating CF. CONCLUSION: Our data indicate the possibility of modifying macrophage phenotype by regulating macrophage glycogen metabolism, suggesting a potential macrophage-based immunotherapy against CF.
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Fibrosis , Macrófagos , Macrófagos/inmunología , Macrófagos/metabolismo , Animales , Ratones , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Miofibroblastos/metabolismo , Glucógeno/metabolismo , Calcio/metabolismo , Lisosomas/metabolismo , Fibroblastos/metabolismo , Humanos , Proteínas de la Membrana/metabolismo , Masculino , Ratones Endogámicos C57BLRESUMEN
Background: Hearing loss and tinnitus have been linked to mild cognitive impairment (MCI); however, the evidence is constrained by ethical and temporal constraints, and few prospective studies have definitively established causation. This study aims to utilize Mendelian randomization (MR) and cross-sectional studies to validate and analyze this association. Methods: This study employs a two-step approach. Initially, the genetic data of the European population from the Genome-wide association studies (GWAS) database is utilized to establish the causal relationship between hearing loss and cognitive impairment through Mendelian randomization using the inverse variance weighted (IVW) method. This is achieved by identifying strongly correlated single nucleotide polymorphisms (SNPs), eliminating linkage disequilibrium, and excluding weak instrumental variables. In the second step, 363 elderly individuals from 10 communities in Qingdao, China are assessed and examined using methods questionnaire survey and pure tone audiology (PTA). Logistic regression and multiple linear regression were used to analyze the risk factors of MCI in the elderly and to calculate the cutoff values. Results: Mendelian randomization studies have shown that hearing loss is a risk factor for MCI in European populations, with a risk ratio of hearing loss to MCI loss of 1. 23. The findings of this cross-sectional study indicate that age, tinnitus, and hearing loss emerged as significant risk factors for MCI in univariate logistic regression analysis. Furthermore, multivariate logistic regression analysis identified hearing loss and tinnitus as potential risk factors for MCI. Consistent results were observed in multiple linear regression analysis, revealing that hearing loss and age significantly influenced the development of MCI. Additionally, a notable finding was that the likelihood of MCI occurrence increased by 9% when the hearing threshold exceeded 20 decibels. Conclusion: This study provides evidence from genomic and epidemiological investigations indicating that hearing loss may serve as a risk factor for cognitive impairment. While our epidemiological study has found both hearing loss and tinnitus as potential risk factors for cognitive decline, additional research is required to establish a causal relationship, particularly given that tinnitus can manifest as a symptom of various underlying medical conditions.
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The prevalence of calcific aortic valve disease (CAVD) remains substantial while there is currently no medical therapy available. Forkhead box O1 (FOXO1) is known to be involved in the pathogenesis of cardiovascular diseases, including vascular calcification and atherosclerosis; however, its specific role in calcific aortic valve disease remains to be elucidated. In this study, we identified FOXO1 significantly down-regulated in the aortic valve interstitial cells (VICs) of calcified aortic valves by investigating clinical specimens and GEO database analysis. FOXO1 silencing or inhibition promoted VICs osteogenic differentiation in vitro and aortic valve calcification in Apoe-/- mice, respectively. We identified that FOXO1 facilitated the ubiquitination and degradation of RUNX2, which process was mainly mediated by SMAD-specific E3 ubiquitin ligase 2 (SMURF2). Our discoveries unveil a heretofore unacknowledged mechanism involving the FOXO1/SMURF2/RUNX2 axis in CAVD, thereby proposing the potential therapeutic utility of FOXO1 or SMURF2 as viable strategies to impede the progression of CAVD.
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Estenosis de la Válvula Aórtica , Válvula Aórtica , Calcinosis , Subunidad alfa 1 del Factor de Unión al Sitio Principal , Proteína Forkhead Box O1 , Ubiquitina-Proteína Ligasas , Ubiquitinación , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Animales , Válvula Aórtica/metabolismo , Válvula Aórtica/patología , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Ratones , Humanos , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Calcinosis/metabolismo , Calcinosis/patología , Calcinosis/genética , Estenosis de la Válvula Aórtica/metabolismo , Estenosis de la Válvula Aórtica/patología , Estenosis de la Válvula Aórtica/genética , Masculino , Osteogénesis/genética , Modelos Animales de Enfermedad , Diferenciación CelularRESUMEN
Background: The crucial role of inflammation in aortic aneurysm (AA) is gaining prominence, while there is still a lack of key cytokines or targets for effective clinical translation. Methods: Mendelian randomization (MR) analysis was performed to identify the causal relationship between 91 circulating inflammatory proteins and AA and between 731 immune traits and AA. Bulk RNA sequencing data was utilized to demonstrate the expression profile of the paired ligand-receptor. Gene enrichment analysis, Immune infiltration, and correlation analysis were employed to deduce the potential role of CX3CR1. We used single-cell RNA sequencing data to pinpoint the localization of CX3CL1 and CX3CR1, which was further validated by multiplex immunofluorescence staining. Cellchat analysis was utilized to infer the CX3C signaling pathway. Trajectory analysis and the Cytosig database were exploited to determine the downstream effect of CX3CL1-CX3CR1. Results: We identified 4 candidates (FGF5, CX3CL1, IL20RA, and SCF) in multiple two-sample MR analyses. Subsequent analysis of the expression profile of the paired receptor revealed the significant upregulation of CX3CR1 in AA and its positive correlation with pro-inflammatory macrophages. Two sample MR between immune cell traits and AA demonstrated the potential causality between intermediate monocytes and AA. We finally deciphered in single-cell sequencing data that CX3CL1 sent by endothelial cells (ECs) acted on CX3CR1 of intermediated monocytes, leading to its recruitment and pro-inflammatory responses. Conclusion: Our study presented a genetic insight into the pathogenetic role of CX3CL1-CX3CR1 in AA, and further deciphered the CX3C signaling pathway between ECs and intermediate monocytes.
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Aneurisma de la Aorta , Receptor 1 de Quimiocinas CX3C , Quimiocina CX3CL1 , Análisis de la Aleatorización Mendeliana , Receptor 1 de Quimiocinas CX3C/genética , Receptor 1 de Quimiocinas CX3C/metabolismo , Humanos , Quimiocina CX3CL1/genética , Quimiocina CX3CL1/metabolismo , Aneurisma de la Aorta/genética , Aneurisma de la Aorta/metabolismo , Perfilación de la Expresión Génica , Transcriptoma , Transducción de Señal , Predisposición Genética a la EnfermedadRESUMEN
The senescence of aortic valve interstitial cells (VICs) plays a critical role in the progression of calcific aortic valve disease (CAVD). However, the precise mechanisms underlying the senescence of VICs remain unclear, demanding the identification of a novel target to mitigate this process. Previous studies have highlighted the anti-aging potential of morusin. Thus, this study aimed to explore the therapeutic potential of morusin in CAVD. Cellular experiments reveal that morusin effectively suppresses cellular senescence and cause a shift toward osteogenic differentiation of VICs in vitro. Mechanistically, morusin activate the Nrf2-mediated antiaging signaling pathway by downregulating CCND1 expression and aiding Keap1 degradation through Trim 25. This activation lead to the upregulated expression of antioxidant genes, thus reducing reactive oxygen species production and thereby preventing VIC osteogenic differentiation. In vivo experiments in ApoE-/- mice on a high-fat Western diet demonstrate the positive effect of morusin in mitigating aortic valve calcification. These findings emphasize the antiaging properties of morusin and its potential as a therapeutic agent for CAVD.
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Estenosis de la Válvula Aórtica , Calcinosis , Senescencia Celular , Flavonoides , Transducción de Señal , Animales , Humanos , Masculino , Ratones , Válvula Aórtica/metabolismo , Válvula Aórtica/patología , Estenosis de la Válvula Aórtica/metabolismo , Estenosis de la Válvula Aórtica/genética , Estenosis de la Válvula Aórtica/patología , Calcinosis/metabolismo , Calcinosis/genética , Senescencia Celular/efectos de los fármacos , Ciclina D1/metabolismo , Ciclina D1/genética , Modelos Animales de Enfermedad , Ratones Endogámicos C57BL , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Osteogénesis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Flavonoides/administración & dosificaciónRESUMEN
BACKGROUND: It is well-established that CD8+ T-cells play a critical role in graft rejection. The basic leucine zipper ATF-like transcription factor (BATF) and BATF3 are transcriptional factors expressed in T lymphocytes. Herein, we investigated the functions of BATF and BATF3 in the differentiation and exhaustion of CD8+ T cells following alloantigen activation. METHODS: Wild-type CD8+ T cells, BATF-deficient (Batf-/-) CD8+ T cells, and CD8+ T cells deficient in both BATF and BATF3 (Batf-/-Batf3-/-) were transferred to B6.Rag1-/- mice, which received skin allografts from BALB/c mice. Flow cytometry was conducted to investigate the number of CD8+ T cells and the percentage of effector subsets. RESULTS: BATF expression positively correlated with effector CD8+ T cell differentiation. BATF and BATF3 deficiency promoted skin allograft long-term survival and attenuated the CD8+ T cell allo-response and cytokine secretion. Finally, BATF and BATF3 deficiency prompted the generation of exhausted CD8+ T cells. CONCLUSIONS: Overall, our findings provide preliminary evidence that both BATF and BATF3 deficiency influences the differentiation of effector CD8+ T cells and mediates the exhaustion of CD8+ T cells, prolonging transplant survival. Targeting BATF and BATF3 to inhibit CD8+ T cell function has huge prospects for application as a therapeutic approach to prevent transplant rejection.
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Linfocitos T CD8-positivos , Trasplante de Piel , Ratones , Animales , Linfocitos T CD8-positivos/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Factores de Transcripción/metabolismo , Regulación de la Expresión Génica , Ratones Endogámicos C57BLRESUMEN
BACKGROUND AND AIMS: Although calcific aortic valve disease (CAVD) is a common valvular disease among elderly populations and its incidence has markedly increased in recent decades, the pathogenesis of CAVD remains unclear. In this study, we explored the potential role of interleukin (IL)-22 and the underlying molecular mechanism in CAVD. METHODS AND RESULTS: Our results showed that IL-22 was upregulated in calcific aortic valves from CAVD patients, and its main sources were CD3+ T cells and CD68+ macrophages. Human aortic valve interstitial cells (VICs) expressed the IL-22-specific receptor IL-22R1, and IL-22R1 expression also was elevated in calcified valves. Treatment of cultured human VICs with recombinant human IL-22 resulted in markedly increased expression of osteogenic proteins Runt-related transcription factor 2 (RUNX2) and alkaline phosphatase (ALP), as well as increased matrix calcium deposition. Moreover, siRNA silencing of IL-22R1 blocked the pro-osteogenic effect of IL-22 in VICs. In IL-22-treated VICs, we also observed increased phosphorylation of JAK3 and STAT3 and nuclear translocation of STAT3. Pretreatment with a specific JAK3 inhibitor, WHIP-154, or siRNA knockout of STAT3 effectively mitigated the IL-22-induced osteoblastic trans-differentiation of human VICs. CONCLUSIONS: Together, these data indicate that IL-22 promotes osteogenic differentiation of VICs by activating JAK3/STAT3 signaling. Based on our results demonstrating a pro-osteogenic role of IL-22 in human aortic valves, pharmacological inhibition of IL-22 signaling may represent a potential strategy for alleviating CAVD.
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Estenosis de la Válvula Aórtica , Válvula Aórtica , Calcinosis , Interleucina-22 , Anciano , Humanos , Válvula Aórtica/patología , Estenosis de la Válvula Aórtica/metabolismo , Células Cultivadas , Osteogénesis , ARN Interferente Pequeño/metabolismoRESUMEN
Resveratrol (RES) is a functional polyphenol that suffers from low water solubility and poor bioavailability. A novel RES-loaded soy protein isolate-dipotassium glycyrrhizinate (SPI-DG) nanocomplex (RES@SPI-DG) was designed and evaluated in this study. RES@SPI-DG was prepared using a simple but novel self-assembly ultrasonic-assisted pH-driven method. The interactions between RES and SPI-DG were non-covalent bonds, including hydrophobic interactions, hydrogen bonds, and van der Waals interactions. RES@SPI-DG exhibited high encapsulation efficiency (97.60 ± 0.38 %) and loading capacity (8.74 ± 0.03 %) of RES with a uniform small size (68.39 ± 1.10 nm). RES in RES@SPI-DG was in an amorphous state and demonstrated a 24-h apparent solubility 482.53-fold higher than bare RES. RES@SPI-DG also showed strong in vitro antioxidant properties. The pH-responsive hydrogel character of SPI-DG makes it an effective intestine-targeted delivery system that could retard the release of RES in a simulated stomach and accelerate it in a simulated intestine. In animal experiments, the bioavailability of RES@SPI-DG was 5.17 times higher than that of bare RES, and the biodistribution was also significantly improved. RES@SPI-DG demonstrated a strong hepatoprotective effect against overdose acetaminophen-induced liver injury. The SPI-DG complex might be a promising nano-platform for enhancing the bioavailability and efficacy of hydrophobic polyphenols such as RES.
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Ácido Glicirrínico , Proteínas de Soja , Animales , Resveratrol , Proteínas de Soja/química , Hidrogeles , Disponibilidad Biológica , Distribución Tisular , Tamaño de la Partícula , Concentración de Iones de HidrógenoRESUMEN
A novel functional food for hyperlipidemia named icariin (ICA) pro-glycymicelles (ICA-PGs) using glycyrrhizin as a phytonanomaterial was easily prepared with improved storage, pH, and salt stabilities. ICA-PGs can easily dissolve in water to self-assemble into a clear glycymicelle solution with high ICA encapsulation efficiency. The ICA in ICA-PGs exhibits significantly increased aqueous solubility, faster in vitro release, and higher bioaccessibility than bare ICA. The ICA-PGs exhibited improved in vitro activities including antioxidant, anti-α-glucosidase, anti-lipase, and anti-cholesterol esterase activities. The ICA-PG also demonstrated improved antioxidant activity in cells. In vivo evaluation confirmed that the ICA-PG demonstrated a significant protective effect against experimental hyperlipidemia in mice, exhibiting decreasing levels of triglycerides (TGs), total cholesterol (TC), and low-density lipoprotein-cholesterol (LDL-C) in the serum, and restoring the hepatic morphology to the normal state. These results indicated that the ICA-PG could improve in vitro/in vivo profiles of ICA, providing a new concept and a promising functional food for hyperlipidemia.
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Hiperlipidemias , Ratones , Animales , Hiperlipidemias/tratamiento farmacológico , Alimentos Funcionales , Flavonoides/farmacología , LDL-Colesterol , Antioxidantes/farmacologíaRESUMEN
BACKGROUND: The effects of the glycoprotein IIb/IIIa receptor inhibitor tirofiban in patients with acute ischemic stroke but who have no evidence of complete occlusion of large or medium-sized vessels have not been extensively studied. METHODS: In a multicenter trial in China, we enrolled patients with ischemic stroke without occlusion of large or medium-sized vessels and with a National Institutes of Health Stroke Scale score of 5 or more and at least one moderately to severely weak limb. Eligible patients had any of four clinical presentations: ineligible for thrombolysis or thrombectomy and within 24 hours after the patient was last known to be well; progression of stroke symptoms 24 to 96 hours after onset; early neurologic deterioration after thrombolysis; or thrombolysis with no improvement at 4 to 24 hours. Patients were assigned to receive intravenous tirofiban (plus oral placebo) or oral aspirin (100 mg per day, plus intravenous placebo) for 2 days; all patients then received oral aspirin until day 90. The primary efficacy end point was an excellent outcome, defined as a score of 0 or 1 on the modified Rankin scale (range, 0 [no symptoms] to 6 [death]) at 90 days. Secondary end points included functional independence at 90 days and a quality-of-life score. The primary safety end points were death and symptomatic intracranial hemorrhage. RESULTS: A total of 606 patients were assigned to the tirofiban group and 571 to the aspirin group. Most patients had small infarctions that were presumed to be atherosclerotic. The percentage of patients with a score of 0 or 1 on the modified Rankin scale at 90 days was 29.1% with tirofiban and 22.2% with aspirin (adjusted risk ratio, 1.26; 95% confidence interval, 1.04 to 1.53, P = 0.02). Results for secondary end points were generally not consistent with the results of the primary analysis. Mortality was similar in the two groups. The incidence of symptomatic intracranial hemorrhage was 1.0% in the tirofiban group and 0% in the aspirin group. CONCLUSIONS: In this trial involving heterogeneous groups of patients with stroke of recent onset or progression of stroke symptoms and nonoccluded large and medium-sized cerebral vessels, intravenous tirofiban was associated with a greater likelihood of an excellent outcome than low-dose aspirin. Incidences of intracranial hemorrhages were low but slightly higher with tirofiban. (Funded by the National Natural Science Foundation of China; RESCUE BT2 Chinese Clinical Trial Registry number, ChiCTR2000029502.).
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Fibrinolíticos , Accidente Cerebrovascular Isquémico , Tirofibán , Humanos , Aspirina/efectos adversos , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/etiología , Fibrinolíticos/efectos adversos , Fibrinolíticos/uso terapéutico , Hemorragias Intracraneales/inducido químicamente , Accidente Cerebrovascular Isquémico/diagnóstico , Accidente Cerebrovascular Isquémico/tratamiento farmacológico , Accidente Cerebrovascular Isquémico/etiología , Inhibidores de Agregación Plaquetaria/efectos adversos , Inhibidores de Agregación Plaquetaria/uso terapéutico , Tirofibán/efectos adversos , Tirofibán/uso terapéutico , Resultado del Tratamiento , Enfermedades Arteriales Cerebrales/tratamiento farmacológico , Enfermedades Arteriales Cerebrales/etiologíaRESUMEN
The ability of carrier selective contact is mainly determined by the surface passivation and work function for dopant-free materials applied in crystalline silicon (c-Si) solar cells, which have received considerable attention in recent years. In this contribution, a novel electron-selective material, lanthanide terbium trifluoride (TbFx ), with an ultra-low work function of 2.4 eV characteristic, is presented, allowing a low contact resistivity (ρc ) of ≈3 mΩ cm2 . Additionally, the insertion of ultrathin passivated SiOx layer deposited by PECVD between TbFx and n-Si resulted in ρc only increase slightly. SiOx /TbFx stack eliminated fermi pinning between aluminum and n-type c-Si (n-Si), which further enhanced the electron selectivity of TbFx on full-area contacts to n-Si. Last, SiOx /TbFx /Al electron-selective contacts significantly improves the open circuit voltage (Voc ) for silicon solar cells, but rarely impacts the short circuit current (Jsc ) and fill factor (FF), thus champion efficiency cell achieved approaching 22% power conversion efficiency (PCE). This study indicates a great potential for using lanthanide fluorides as electron-selective material in photovoltaic devices.
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BACKGROUND: IRF4 is the pioneer factor for effector T cell maturation. Here we investigated the function of IRF4 in maintaining OX40-related T cell responses following alloantigen activation in a mouse heart transplantation model. METHODS: Irf4flox/flox mice were bred with Ox40cre/+ mice to generate Irf4flox/floxOx40cre/+ mice. Wild type C57BL/6, Irf4flox/floxOx40cre/+ mice were transplanted with BALB/c heart allografts, with or without BALB/c skin-sensitization. CD4+ TEa T cells co-transfer experiments and flow cytometric analysis were conducted to investigate the amount of CD4+ T cells and the percentage of the T effector subset. RESULTS: Irf4flox/floxOx40cre/+ and Irf4flox/floxOx40cre/+ TEa mice were constructed successfully. IRF4 ablation in activated OX40-mediated alloantigen specific CD4+ TEa T cells reduced effector T cell differentiation (CD44hiCD62Llo, Ki67, IFN-γ), which caused long-term allograft survival (> 100 d) in the chronic rejection model. In the donor skin-sensitized heart transplantation model, the formation and function of alloantigen-specific memory CD4+ TEa cells were also impaired in Irf4flox/floxOx40cre/+ mice. Additionally, deletion of IRF4 after T cell activation in Irf4flox/floxOx40cre/+ mice reduced T cell reactivation in vitro. CONCLUSIONS: IRF4 ablation after OX40-related T cell activation could reduce effector and memory T cell formation and inhibit their function in response to alloantigen stimulation. These findings could have significant implications in targeting activated T cells to induce transplant tolerance.
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Trasplante de Corazón , Células T de Memoria , Animales , Ratones , Memoria Inmunológica , Isoantígenos , Células T de Memoria/inmunología , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
BACKGROUND AND AIMS: Inflammatory molecules play important roles in atherosclerosis. We aimed to illustrate the roles of serum amyloid A (SAA), and interleukin (IL)-1ß in low density lipoproteins (LDL) transcytosis and atherosclerosis. METHODS: Effects of SAA and IL-1ß on transcytosis of LDL were measured by an in vitro LDL transcytosis model. NF-κB/caveolin-1/cavin-1 pathway activation was investigated by Western blots and ELISA. Effects of SAA and IL-1ß on the retention of LDL in aorta of C57BL/6J mice were detected by IVIS spectrum. Effects of SAA and IL-1ß on atherosclerosis in Apoe-/- mice were examined by Oil Red O staining. RESULTS: SAA and IL-1ß stimulated LDL transcytosis across endothelial cells (ECs), which was accompanied by an increase in LDL uptake by ECs. SAA and IL-1ß enhanced the activity of nuclear factor (NF)-κB, consequently facilitating an up-regulation of proteins involved in caveolae formation, including caveolin-1 and cavin-1, along with an assembly of NLRP3 inflammasome. Furthermore, SAA- and IL-1ß-induced effects were blocked by NF-κB subunit p65 siRNA. Meanwhile, SAA- and IL-1ß-induced LDL transcytosis were effectively blocked by caveolin-1 siRNA or cavin-1 siRNA. Interestingly, SAA and IL-1ß facilitated LDL entering into the aorta of C57BL/6J mice. In Apoe-/- mice, SAA and IL-1ß increased the areas of lipid-rich atherosclerotic lesions in the both ascending and root of aorta. Furthermore, a significant increase in the NLRP3 inflammasome, accompanied by accumulation of cavin-1 and caveolin-1, was observed in the aortic endothelium of Apoe-/- mice. CONCLUSIONS: SAA and IL-1ß accelerated LDL transcytosis via the NF-κB/caveolin-1/cavin-1 axis.
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Aterosclerosis , FN-kappa B , Proteína Amiloide A Sérica , Animales , Ratones , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerosis/genética , Aterosclerosis/metabolismo , Caveolina 1/genética , Caveolina 1/metabolismo , Caveolina 1/farmacología , Células Endoteliales/metabolismo , Inflamasomas/metabolismo , Interleucinas/metabolismo , Ratones Endogámicos C57BL , FN-kappa B/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , ARN Interferente Pequeño/metabolismo , Proteína Amiloide A Sérica/metabolismo , TranscitosisRESUMEN
Calcified aortic valve disease (CAVD) is a major non-rheumatic heart valve disease in the world, with a high mortality rate and without suitable pharmaceutical therapy due to its complex mechanisms. Src-associated in mitosis 68-KD (Sam68), an RNA binding protein, has been reported as a signaling adaptor in numerous signaling pathways (Huot in Mol Cell Biol, 29(7), 1933-1943, 2009), particularly in inflammatory signaling pathways. The effects of Sam68 on the osteogenic differentiation process of hVICs and its regulation on signal transducer and activator of transcription 3 (STAT3) signaling pathway have been investigated in this study. Human aortic valve samples detection found that Sam68 expression was up-regulated in human calcific aortic valves. We used tumor necrosis factor α (TNF-α) as an activator for osteogenic differentiation in vitro and the result indicated that Sam68 was highly expressed after TNF-α stimulation. Overexpression of Sam68 promoted osteogenic differentiation of hVICs while Sam68 knockdown reversed this effect. Sam68 interaction with STAT3 was predicted by using String database and was verified in this study. Sam68 knockdown reduced phosphorylation of STAT3 activated by TNF-α and the downstream gene expression, which further influenced autophagy flux in hVICs. STAT3 knockdown alleviated the osteogenic differentiation and calcium deposition promoted by Sam68 overexpression. In conclusion, Sam68 interacts with STAT3 and participates in its phosphorylation to promote osteogenic differentiation of hVICs to induce valve calcification. Thus, Sam68 may be a new therapeutic target for CAVD. Regulatory of Sam68 in TNF-α/STAT3/Autophagy Axis in promoting osteogenesis of hVICs.
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Objective: Chronic rejection remains the main factor that influence long-term survival of patients after heart transplantation. Interleukin-10 (IL-10) play critical role in macrophages-mediated transplant immune responses. We investigated the mechanism of IL-10 in macrophage related chronic rejection after mouse heart transplantation. Methods: Mouse heart transplant chronic rejection model was established to evaluate pathological changes in the allograft. Myocardial interstitial fibrosis, apoptosis, and inflammatory factor levels were detected in ad-IL-10-treated mice. The positive iNOS+ and Arg-1+ expressions, macrophage subset changes, and the proportion of regulatory T-cells (Tregs) and TIGIT+ Tregs were quantified by flow. In in vitro experiments, ad-IL-10 was transfected into macrophages followed by detection of apoptosis, phagocytosis, and CD163, CD16/32, and CD206 expression. The expression and relationships between IL-10, miR-155, and SOCS5 were also detected and verified. A rescue experiment was performed to evaluate macrophage function through the combined treatment of ad-IL-10 and overexpression of miR-155. Results: Significantly decreased IL-10 expression in chronic rejection during mouse heart transplantation was observed. Ad-IL-10-treated mice showed decreased pathological injury, perivascular fibrosis, apoptosis, inflammation, and iNOS+ and CD16/32+ expression, and increased Treg/TIGIT+ Treg cell, Arg-1+ and CD206+ cell proportion. Ad-IL-10-treated macrophages in vitro showed reduced apoptosis, improved phagocytosis, and M2 polarization. Mechanically, IL-10 negatively regulated miR-155 to activate SOCS5. Overexpression of miR-155 reversed IL-10 mediated-positive regulation of macrophage function. Conclusion: IL-10 downregulated miR-155 and activated SOCS5, thereby promoting macrophage M2 polarization to relieve chronic rejection after heart transplantation.
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Rechazo de Injerto , Trasplante de Corazón , MicroARNs , Animales , Ratones , Adenoviridae/metabolismo , Trasplante de Corazón/efectos adversos , Interleucina-10/genética , Interleucina-10/metabolismo , Macrófagos/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Rechazo de Injerto/prevención & controlRESUMEN
INTRODUCTION: Calcific aortic valve disease (CAVD) is an active and cellular-driven fibrocalcific process characterised by differentiation of valve interstitial cells (VICs) towards an osteogenic-like phenotype. A recently identified lncRNA, lncTSI, has been reported to inhibit fibrogenesis through transforming growth factor (TGF)-ß/Smad3 pathway. Here, the present study aimed to investigate the role of lncTSI in CAVD. METHODS: The effect of TGF-ß1 on lncTSI of VICs was measured. TGF-ß1, RUNX2 and collagen I expression between calcified aortic valve tissue and normal samples by immunohistochemistry and western blotting. Human VICs were cultured and treated with TGF-ß1. SiRNA and pcDNA3.1-lncTSI plasmid transfection were used to silence and overexpress lncTSI in VICs for 48 h, Smads phosphorylation, RUNX2 and collagen I expression were then verified by western blotting. In ApoE-/- mice fed with 0.25 % high-cholesterol diet, AAV2-lncTSI were injected intravenously to observe their effect on the formation of aortic valve calcification. RESULTS: lncTSI was highly expressed in VICs treated with TGF-ß1. lncTSI was transcriptionally regulated by Smad3 and reversely inhibited TGF-ß1-induced Smad3 phosphorylation and downregulated profibrotic gene expression. Silencing lncTSI increased TGF-ß1-induced Smad3 phosphorylation, and subsequently, upregulated RUNX2 and collagen I expressions in VICs. While overexpression of lncTSI reversed the production of RUNX2 and collagen I in VICs. In a mouse CAVD model of 24 week 0.25 % high-cholesterol diet feeding, overexpression of lncTSI significantly reduced calcium deposition, RUNX2, pSmad3, and collagen I expression in aortic valve leaflets, with less aortic valve stenosis. CONCLUSIONS: The novel findings of present study suggested that lncTSI alleviated aortic valve calcification through negative regulation of the TGF-ß/Smad3 pathway. The results may help elucidate new diagnostic and therapeutic targets to prevent CAVD progression.
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Estenosis de la Válvula Aórtica , Válvula Aórtica , ARN Largo no Codificante , Animales , Humanos , Masculino , Ratones , Válvula Aórtica/metabolismo , Estenosis de la Válvula Aórtica/genética , Estenosis de la Válvula Aórtica/metabolismo , Células Cultivadas , Colesterol/metabolismo , Colágeno/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Osteogénesis , Factor de Crecimiento Transformador beta1/farmacología , Factor de Crecimiento Transformador beta1/metabolismo , ARN Largo no Codificante/genéticaRESUMEN
Dysregulation of gene amplification, cell-signaling-pathway transduction, epigenetic and transcriptional regulation, and protein interactions drives tumor-cell proliferation and invasion, while ion channels also play an important role in the generation and development of tumor cells. Overexpression of Ca2+-activated Cl- channel anoctamin 1 (ANO1) is shown in numerous cancer types and correlates with poor prognosis. However, the mechanisms involved in ANO1-mediated malignant cellular transformation and the role of ANO1 in tumor immunity remain unknown. In this review, we discuss recent studies to determine the role of ANO1 in tumorigenesis and provide novel insights into the role of ANO1 in the context of tumor immunity. Furthermore, we analyze the roles and potential mechanisms of ANO1 in different types of cancers, and provide novel notions for the role of ANO1 in the tumor microenvironment and for potential use of ANO1 in clinical applications. Our review shows that ANO1 is involved in tumor immunity and microenvironment, and may, therefore, be an effective biomarker and therapeutic drug target.
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Canales de Cloruro , Neoplasias , Anoctamina-1/genética , Anoctamina-1/metabolismo , Carcinogénesis , Proliferación Celular , Canales de Cloruro/genética , Canales de Cloruro/metabolismo , Canales de Cloruro/farmacología , Humanos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Neoplasias/patología , Microambiente TumoralRESUMEN
The Ca2+-activated Cl- channel ANO1 is widely expressed in epithelial cells, and ANO1 upregulation is implicated in the oncogenesis of many epithelium-originated cancers. However, whether ANO1 plays a causal role in the tumorigenesis of colorectal cancer remains largely unknown. Here, we show that ANO1 channel protein is upregulated in human colorectal cancer tissue samples and its upregulation is correlated with the TNM staging, histological type, pathological differentiation and poor prognosis. Knockdown or pharmacological inhibition of ANO1 suppresses colorectal cancer cell proliferation and induces cell apoptosis. Furthermore, ANO1 knockdown inhibits the growth of subcutaneous xenograft tumors implanted with colorectal cancer HT-29 cells in nude mice. Mechanically, knockdown of endogenous ANO1 inactivates the Wnt/ß-catenin signaling through downregulating critical components, such as Frizzled protein 1, ß-catenin and upregulating GSK3ß. Taken together, our results demonstrate that ANO1 upregulation is involved in the tumorigenesis of colorectal cancer, and inhibition of ANO1 upregulation or inactivating downstream Wnt/ß-catenin signaling may have therapeutic potential for colorectal cancer.