Differential microRNAs expression profiles in liver from three different lifestyle modification mice models.

BACKGROUND: MicroRNAs play an important role in many fundamental biological and pathological processes. Defining the microRNAs profile underlying the processes by beneficial and detrimental lifestyles, including caloric restriction (CR), exercise and high-fat diet (HF), is necessary for understanding both normal physiology and the pathogenesis of metabolic disease. We used the microarray to detect microRNAs expression in livers from CR, EX and HF mice models. After predicted potential target genes of differentially expressed microRNAs with four algorithms, we applied GO and KEGG to analyze the function of predicted microRNA targets. RESULTS: We describe the overall microRNAs expression pattern, and identified 84 differentially expressed microRNAs changed by one or two or even all the three lifestyle modifications. The common and different enriched categories of gene function and main biochemical and signal transduction pathways were presented. CONCLUSIONS: We provided for the first time a comprehensive and thorough comparison of microRNAs expression profiles in liver among these lifestyle modifications. With this knowledge, our findings provide us with an overall vision of microRNAs in the molecular impact of lifestyle on health as well as useful clues for future and thorough research of the role of microRNAs.

Dexrazoxane ameliorates doxorubicin-induced cardiotoxicity by inhibiting both apoptosis and necroptosis in cardiomyocytes.

Doxorubicin, as a first line chemotherapeutic agent, its usage is limited owing to cardiotoxicity. Necroptosis is a new form of programmed cell death, and recent investigations indicated that necroptosis is vitally involved in serious cardiac pathological conditions. Dexrazoxane is the only cardiac protective drug approved by FDA for anthracycline. We aimed to explore whether and how dexrazoxane regulates doxorubicin-induced cardiomyocyte necroptosis. First, doxorubicin could cause heart failure and reduce cardiomyocyte viability by promoting cell apoptosis and necroptosis in vivo and in vitro. Second, necroptosis plays an important role in doxorubicin induced cardiomyocyte injury, which could be inhibited by Nec-1. Third, dexrazoxane increased cell viability and protect heart function by decreasing both cardiomyocyte apoptosis and necroptosis after doxorubicin treatment. Forth, dexrazoxane attenuated doxorubicin-induced inflammation and necroptosis by the inhibition of p38MAPK/NF-κB pathways. These results indicated that dexrazoxane ameliorates cardiotoxicity and protects heart function by attenuating both apoptosis and necroptosis in doxorubicin induced cardiomyocyte injury.

Wnt1 inhibits vascular smooth muscle cell calcification by promoting ANKH expression.

AIMS: Wnt signaling plays a critical role in vascular calcification (VC). Wnt factors induce different physiological and pathological effects on cardiovascular functions. Wnt1, a ligand of Wnt/ß-catenin signaling, promotes pro-angiogenesis and reduces myocardial infarction. The role of Wnt1 on VC in chronic kidney disease (CKD) is not fully understood. METHODS AND RESULTS: We used human vascular smooth muscle cells (VSMCs) and a rat model of chronic renal failure (CRF), and observed a native protective mechanism by which VC is reduced via the activation of Wnt1 and its transcriptional target ANKH inorganic pyrophosphate transport regulator (ANKH) gene. ANKH is an essential calcification inhibitor that effluxes inorganic pyrophosphate (PPi) from VSMCs to play an inhibitory role in VC. Vascular ANKH and plasma PPi were significantly downregulated in the rat model of CRF. The knockdown or inhibition of ANKH reversed the effect of Wnt1 on VC in VSMCs. Clinical analysis revealed low plasma levels of Wnt1 and PPi were associated with CKD in patients. Applying a Wnt/ß-catenin signaling agonist can alleviate the progression of VC. CONCLUSION: This work reveals the ANKH regulation of Wnt1 in VSMCs is essential for blocking VC. Our findings may contribute to the development of medications that target Wnt signaling and/or ANKH to inhibit VC.

Changes of upright body posture in the sagittal plane of men and women occurring with aging - a cross sectional study.

BACKGROUND: Body posture is a fundamental indicator for assessing health and quality of life, especially for elderly people. Deciphering the changes in body posture occurring with age is a current topic in the field of geriatrics. The aims of this study were to assess the parameters of standing body posture in the global sagittal plane and to determine the dynamics of changes in standing body posture occurring with age and differences between men and women. METHODS: The measurements were performed on 226 individuals between the ages of 20 to 89 with a new photogrammetry, via which we assessed five postural angles - neck, thorax, waist, hip and knee. The data were analyzed with t-test, one-way ANOVA, linear regression model and generalized additive model. RESULTS: Among these segments studied here, neck changed most, while the middle segments of the body, waist and hip, were relative stable. Significant differences between men and women were found with respect to the angles of neck, thorax and hip. Three of the five postural angles were significantly influenced with aging, including increasing cervical lordosis, thoracic kyphosis and knee flexion, starting from no older than around 50 yrs. showed by fitting curve derived with generalized additive model. These changes were more marked among women. Besides, this study highlights the effects of age and gender on the complex interrelation between adjacent body segments in standing. CONCLUSIONS: The presented results showed changes in the parameters describing body posture throughout consecutive ages and emphasized that for an individualized functional analysis, it is essential to consider age-and gender-specific changes in the neck, thorax and knee. This paper presents useful externally generalizable information not only for clinical purposes but also to inform further research on larger numbers of subjects.

Ginkgolide B inhibits platelet and monocyte adhesion in TNFα-treated HUVECs under laminar shear stress.

BACKGROUND: Endothelial cells are sensitive to changes in both blood components and mechanical stimuli. Endothelial cells may undergo phenotypic changes, such as changes in adhesion protein expression, under different shear stress conditions. Such changes may impact platelet and monocyte adhesion to endothelial cells. This phenomenon is linked to chronic vascular inflammation and the development of atherosclerosis. In the present study, we investigated the effects of ginkgolide B on platelet and monocyte adhesion to human umbilical vein endothelial cells (HUVECs) under different conditions of laminar shear stress. METHODS: Platelet and monocyte adhesion to endothelial cells was determined by the Bioflux 1000. HUVECs were incubated with ginkgolide B or aspirin for 12 h, and then TNFα was added for 2 h to induce the inflammatory response under conditions of 1 and 9 dyn/cm2 laminar shear stress. The protein expression was analyzed by Western blot. RESULTS: The number of platelets that adhered was greater under conditions of 1 dyn/cm2 than under conditions of 9 dyn/cm2 of laminar shear stress (74.8 ± 19.2 and 59.5 ± 15.1, respectively). Ginkgolide B reduced the tumor necrosis factor α (TNFα)-induced increase in platelet and monocyte adhesion to HUVECs at 1 and 9 dyn/cm2 of laminar shear stress. In TNFα-treated HUVECs, the number of monocytes that adhered was greater under conditions of 1 dyn/cm2 of laminar shear stress compared with 9 dyn/cm2 (29.1 ± 4.9 and 22.7 ± 3.7, respectively). Ginkgolide B inhibited the TNFα-induced expression of vascular cell adhesion molecule-1(VCAM-1), VE-cadherin, and Cx43 in HUVECs at 1 and 9 dyn/cm2. The expression of these proteins was not different between 1 and 9 dyn/cm2. CONCLUSIONS: Ginkgolide B suppressed platelet and monocyte adhesion under different conditions of laminar shear stress. Moreover, ginkgolide B reduced VCAM-1, VE-cadherin and Cx43 expression in TNFα-treated HUVECs under laminar shear stress. This suggested that ginkgolide B might shed light on the treatment of inflammation in atherosclerosis.

Thioredoxin attenuates oxidized low-density lipoprotein induced oxidative stress in human umbilical vein endothelial cells by reducing NADPH oxidase activity.

Oxidative stress is recognized as one of the most important contributing factors to the development of atherosclerosis. Oxidized low-density lipoprotein (ox-LDL) can induce vascular reactive oxygen species (ROS) production, trigger endothelial dysfunction and initiate the progression of atherosclerosis. Previous studies have demonstrated that thioredoxin-1 (Trx) is one of the key regulators of intracellular redox, which is pivotal in atherogenesis. However, the regulation mechanism is still unclear. In this study, we investigated the effects of Trx1 on NADPH oxidase in human umbilical vein endothelial cells (HUVECs), whose ROS level is mainly produced by NADPH oxidase, especially Nox4 isoform. Our data demonstrated that Trx decreased NADPH oxidase activity, ROS production and ICAM-1 expression in ox-LDL treated HUVECs. Genetic gain-of-function and loss-of-function studies showed that Trx1 suppressed ox-LDL-induced Nox4 and p22phox expression. A co-immunoprecipitation assay indicated that Trx1 decreased Nox4-p22phox complex level during ox-LDL stimulation. Transient transfection of Nox4 and p22phox significantly increased intracellular ROS generation, which could be blocked by Trx overexpression. In addition, Trx overexpression also prevented ox-LDL-induced Nox2 and Rac1 protein levels. These results suggest that Trx suppresses NADPH oxidase activity in vascular endothelia under pathological conditions and may prevent the initiation of atherosclerosis by attenuating exceeding ROS production.

T-box20 suppresses oxidized low-density lipoprotein-induced human vascular endothelial cell injury by upregulation of PPAR-γ.

BACKGROUND: Atherosclerosis is a chronic inflammation disease which is initiated by endothelial cell injury Oxidized low-density lipoprotein (ox-LDL) is directly associated with chronic vascular inflammation. Many transcription factors take part in the initiation and progression of atherosclerosis. As a transcription factor mainly expressed in cardiovascular system, T-box20 (Tbx20) plays an important role in embryonic cardiovascular system development and homeostasis. However, the role of Tbx20 in endothelial cell injury and atherosclerosis is still not clear. We showed that Tbx20 might affect ox-LDL-induced inflammatory responses in human umbilical vein endothelial cells (HUVECs). METHODS AND RESULTS: First, Tbx20 expression was down regulated in the C57BL/6 mice with high-fat diet-induced artery injury, which was accompanied by elevated reactive oxygen species (ROS) generation and cell adhesion molecule expression. Second, ox-LDL led to concurrent decreased Tbx20 expression and increased levels of ROS and adhesion molecules in the HUVECs. Third, over-expression of Tbx20 by adenovirus reduced ox-LDL-induced HUVEC injury via attenuation of ROS generation and cell adhesion molecule expression. Fourth, knock down of Tbx20 by siRNA significantly increased adhesion molecule expression and decreased cell viability. Moreover, Tbx20 could directly regulate PPAR-γ expression, as shown by Tbx20 knock down and PPAR-γ inhibition, which significantly reversed Tbx20's HUVEC protection effect. CONCLUSIONS: These results indicate that misregulation of Tbx20 could reduce HUVEC tolerance of ox-LDL-induced cell injury, suggesting that Tbx20 might be a crucial regulator and potential therapeutic target for atherosclerosis.

Astragalus polysaccharides suppress ICAM-1 and VCAM-1 expression in TNF-α-treated human vascular endothelial cells by blocking NF-κB activation.

AIM: To investigate the effects Astragalus polysaccharides (APS) on tumor necrosis factor (TNF)-α-induced inflammatory reactions in human umbilical vein endothelial cells (HUVECs) and to elucidate the underlying mechanisms. METHODS: HUVECs were treated with TNF-α for 24 h. The amounts of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were determined with Western blotting. HUVEC viability and apoptosis were detected using cell viability assay and Hoechst staining, respectively. Reactive oxygen species (ROS) production was measured by DHE staining. Monocyte and HUVEC adhesion assay was used to detect endothelial cell adhesive function. NF-κB activation was detected with immunofluorescence. RESULTS: TNF-α (1-80 ng/mL) caused dose- and time-dependent increases of ICAM-1 and VCAM-1 expression in HUVECs, accompanied by significant augmentation of IκB phosphorylation and NF-κB translocation into the nuclei. Pretreatment with APS (10 and 50 µg/mL) significantly attenuated TNFα-induced upregulation of ICAM-1 VCAM-1 and NF-κB translocation. Moreover, APS significantly reduced apoptosis, ROS generation and adhesion function damage in TNF-α-treated HUVECs. CONCLUSION: APS suppresses TNFα-induced adhesion molecule expression by blocking NF-κB signaling and inhibiting ROS generation in HUVECs. The results suggest that APS may be used to treat and prevent endothelial cell injury-related diseases.

Chronic unpredictable mild stress promotes atherosclerosis via adipose tissue dysfunction in ApoE-/- mice.

Background: Chronic unpredictable mild stress (CUMS) has been shown to exacerbate atherosclerosis, but the underlying mechanism remains unknown. Adipose tissue is an energy storage organ and the largest endocrine organ in the human body, playing a key role in the development of cardiovascular disease. In this research, it was hypothesized that CUMS may exacerbate the development of atherosclerosis by inducing the hypertrophy and dysfunction of white adipocytes. Methods: The CUMS-induced atherosclerosis model was developed in Western diet-fed apolipoprotein E (ApoE)-/- mice. White adipose tissue (WAT), serum, aortic root, and the brachiocephalic trunk were collected and tested after 12 weeks of CUMS development. The mouse model of CUMS was evaluated for depression-like behavior using the open field test (OFT) and the elevated plus maze (EPM) test. Enzyme-linked immunosorbent assay (ELISA) was conducted to detect serum noradrenaline and urine adrenaline protein levels. Serological assays were used to detect serum low-density lipoprotein (LDL), high-density lipoprotein (HDL), total cholesterol (TC), and free fatty acid (FFA) concentrations. Hematoxylin and eosin (H&E) staining and oil red O were used to detect atherosclerotic plaque area, lipid deposition, and adipocyte size. The mRNA levels of genes related to aberrant adipose tissue function were determined using real-time PCR. Immunofluorescence assay and western blotting were conducted to examine the expression of proteins in the adipose tissue samples. Results: CUMS aggravated vascular atherosclerotic lesions in ApoE-/- mice. It decreased body weight while increasing the percentage of WAT. The serological results indicated that the concentration of HDL decreased in CUMS mice. Notably, adipocyte hypertrophy increased, whereas the mRNA levels of Pparg and its target genes (Slc2a4 (encodes for GLUT4), Adipoq, and Plin1) decreased. Further investigation revealed that CUMS increased subcutaneous inguinal WAT (iWAT) lipid synthesis and adipocyte inflammation while decreasing lipid hydrolysis and the expression of HDL-associated protein ApoA-I. Moreover, CUMS aggravated insulin resistance in mice and inhibited the insulin pathway in iWAT. Conclusions: These findings indicated that CUMS induces adipose tissue dysfunction via a mechanism that leads to dyslipidemia, increased inflammation, and insulin resistance in the body, thereby exacerbating atherosclerosis. Notably, CUMS that is involved in decreasing the expression of HDL-associated proteins in adipose tissue may be a crucial link between adipose hypertrophy and advanced atherosclerosis. This study reveals a novel mechanism via which CUMS exacerbates atherosclerosis from the novel perspective of abnormal adipose function and identifies a novel potential therapeutic target for this disease.

Delphinidin induces autophagic flux blockage and apoptosis by inhibiting both multidrug resistance gene 1 and DEAD-box helicase 17 expressions in liver cancer cells.

OBJECTIVES: To investigate the function and regulatory mechanisms of delphinidin in the treatment of hepatocellular carcinoma. METHODS: HepG2 and HuH-7 cells were treated with different concentrations of delphinidin. Cell viability was analysed by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. The cell autophagy and autophagic flux were analysed by LC3b-green fluorescent protein (GFP)-Adv and LC3b-GFP-monomeric red fluorescent protein-Adv transfected HepG2 and HuH-7 cells, respectively. Cell apoptosis was analysed by Hoechst33342 staining, terminal deoxynucleotidyl transferase dUTP nick end labeling staining and DNA laddering. Cell autophagy, apoptosis and survival related protein expressions were detected by Western blotting. KEY FINDINGS: After treatment with different concentrations of delphinidin, the cell survival rate was significantly decreased. Delphinidin could block the autophagic flux, resulting in a significant increase in autophagosomes, and led to an increase in cell apoptosis. The combined application of delphinidin and cisplatin could promote the antitumour effect and reduce the dose of cisplatin in tumour cells. Further mechanism studies reveal that delphinidin could inhibit the multidrug resistance gene 1 (MDR1) and the tumour-promoting transcription cofactor DEAD-box helicase 17 (DDX17) expression in tumour cells. Overexpression of DDX17 could reverse delphinidin's antitumor function in tumour cells. CONCLUSIONS: Delphinidin has a strong anti-tumour effect by inducing tumour cell autophagic flux blockage and apoptosis by inhibiting of both MDR1 and DDX17 expression.

Corrigendum: DDX5 and DDX17-multifaceted proteins in the regulation of tumorigenesis and tumor progression.

[This corrects the article DOI: 10.3389/fonc.2022.943032.].

Aspirin inhibits the production of reactive oxygen species by downregulating Nox4 and inducible nitric oxide synthase in human endothelial cells exposed to oxidized low-density lipoprotein.

Aspirin has antithrombotic activity and is commonly used to protect patients from cardiovascular disease attacks. The present study investigated whether aspirin reduces reactive oxygen species and proinflammatory proteins in oxidized low-density lipoprotein (ox-LDL)-stimulated human umbilical vein endothelial cells. The results showed that aspirin attenuated reactive oxygen species generation induced by ox-LDL and downregulated Nox4 and inducible nitric oxide synthase expression. Redox-sensitive transcription factor nuclear factor kappa B was inactivated by aspirin, significantly preventing nuclear factor kappa B p65 subunit translocation into the nucleus. The expression of the monocyte/macrophage chemotactic protein 1 also decreased, but endothelial nitric oxide synthase expression increased in aspirin-treated cells. Aspirin ameliorated oxidative stress by downregulating Nox4 and inducible nitric oxide synthase and improved endothelial cell function by increasing endothelial nitric oxide synthase expression. Thus, aspirin may possess protective effects against ox-LDL-induced endothelial cell injury.

Design of functionalized magnetic silica multi-core composite nanoparticles for synergistic magnetic hyperthermia/radiotherapy in cancer cells.

Nanomaterials in particular the magnetic nanoparticles (MNPs) offer tremendous potential for cancer treatment due to their unique intrinsic properties. Combining materials with a variety of functional groups, and forming a multifunctional nanosystem to overcome the limitations of monotherapy for cancer treatment has always been a research focus with notable difficulties. Considering the many challenges faced by radiotherapy and hyperthermia, in this study, we designed a rational strategy for magnetic hyperthermia using Fe3O4@SiO2@Sec2@FA nanoparticles as a novel nano-radiosensitizer to simultaneously enhance the therapeutic effects of radiotherapy in the future. Fe3O4@SiO2 core-shell structured nanoparticles were synthesized with an appropriate silica layer thickness to maintain good saturation magnetization. The as-prepared Fe3O4@SiO2@Sec2@FA nanoparticles had the specific absorption rate (SAR)value of 57 W/g, which was below the clinically acceptable alternating magnetic field value of 4.9 × 109 Am-1s-1, indicating good heat generation efficiency (the temperature level ΔT=6-10 °C). Moreover, Folate-modified nanoparticles exhibited approximately 6-fold higher cellular internalization of Hela cells with no obvious cytotoxicity for the Hela and MDA-MB-231 cells, and lower cytotoxicity for the HUVECs in a concentration range of 0-150 µg/mL. In addition, these nanoparticles were modified on the silica surface by L-selenocystine, which could enhance the elimination of tumor cells by producing reactive oxygen species under X-rays, resulting in a novel radiosensitization effect. Therefore, the as-prepared Fe3O4@SiO2@Sec2@FA nanoparticles with good biocompatibility and active targeting would possess synergistic magnetic hyperthermia/radiotherapy effect.

miR-488-3p Protects Cardiomyocytes against Doxorubicin-Induced Cardiotoxicity by Inhibiting CyclinG1.

OBJECTIVE: To investigate the protective effects and regulatory mechanism of miR-488-3p on doxorubicin-induced cardiotoxicity. METHODS: The C57BL/6 mice and primary cardiomyocytes were used to construct doxorubicin-induced cardiomyocyte injury models in vivo and in vitro. The levels of miR-488-3p and its downstream target genes were analyzed by quantitative real-time PCR. Mouse cardiac function, cell survival, cellular injury-related proteins, and the apoptosis level of cardiomyocytes were analyzed by echocardiography, MTT analysis, Western blotting, and DNA laddering separately. RESULTS: Cardiomyocyte injury caused by a variety of stimuli can lead to the reduction of miR-488-3p level, especially when stimulated with doxorubicin. Doxorubicin led to significant decrease in cardiac function, cell autophagic flux blockage, and apoptosis in vivo and in vitro. The expression of miR-488-3p's target gene, CyclinG1, increased remarkably in the doxorubicin-treated neonatal mouse cardiomyocytes. Overexpression of miR-488-3p inhibited CyclinG1 expression, increased cardiomyocyte viability, and attenuated doxorubicin-induced cardiomyocyte autophagic flux blockage and apoptosis. CONCLUSIONS: miR-488-3p is one of the important protective miRNAs in doxorubicin-induced cardiotoxicity by inhibiting the expression of CyclinG1, which provides insight into the possible clinical application of miR-488-3p/CyclinG1 as therapeutic targets in doxorubicin-induced cardiovascular diseases.

DDX5 and DDX17-multifaceted proteins in the regulation of tumorigenesis and tumor progression.

DEAD-box (DDX)5 and DDX17, which belong to the DEAD-box RNA helicase family, are nuclear and cytoplasmic shuttle proteins. These proteins are expressed in most tissues and cells and participate in the regulation of normal physiological functions; their abnormal expression is closely related to tumorigenesis and tumor progression. DDX5/DDX17 participate in almost all processes of RNA metabolism, such as the alternative splicing of mRNA, biogenesis of microRNAs (miRNAs) and ribosomes, degradation of mRNA, interaction with long noncoding RNAs (lncRNAs) and coregulation of transcriptional activity. Moreover, different posttranslational modifications, such as phosphorylation, acetylation, ubiquitination, and sumoylation, endow DDX5/DDX17 with different functions in tumorigenesis and tumor progression. Indeed, DDX5 and DDX17 also interact with multiple key tumor-promoting molecules and participate in tumorigenesis and tumor progression signaling pathways. When DDX5/DDX17 expression or their posttranslational modification is dysregulated, the normal cellular signaling network collapses, leading to many pathological states, including tumorigenesis and tumor development. This review mainly discusses the molecular structure features and biological functions of DDX5/DDX17 and their effects on tumorigenesis and tumor progression, as well as their potential clinical application for tumor treatment.

Ginkgolide B reduces inflammatory protein expression in oxidized low-density lipoprotein-stimulated human vascular endothelial cells.

Ginkgolide B is a herbal constituent extracted from leaves of the ginkgo biloba tree. Previous studies have shown that ginkgolide B is a specific platelet activating factor (PAF) receptor antagonist, and it suppresses PAF-mediated platelet activation via competitive binding. In this study, the effect of ginkgolide B on nicotinamide adenine dinucleotide phosphate oxidase and other inflammatory proteins in ox-LDL (low-density lipoprotein)-stimulated human vascular endothelial cells was investigated. Another PAF receptor antagonist CV3988 was employed to compare with ginkgolide B in this study. Our results show that the enhancement of Nox4 expression and reactive oxygen species generation was attenuated by ginkgolide B in cells treated with ox-LDL but not with CV3988. Increases in monocyte chemoattractant protein-1 and intercellular adhesion molecule 1 expression induced by ox-LDL, however, were inhibited by both ginkgolide B and CV3988. The translocation of NF-kappaB p65 (NF-κB) into the nucleus was inhibited by both ginkgolide B and CV3988. In conclusion, both ginkgolide B and CV3988 can inhibit the expression of inflammatory proteins by blocking NF-κB translocation. It seems that ginkgolide B possesses some pharmacological action on intracellular oxidative stress in association with the downregulation of Nox4 expression.

Water-Soluble Tomato Extract Fruitflow Alters the Phosphoproteomic Profile of Collagen-Stimulated Platelets.

Platelet hyperactivity is a risk factor for cardiovascular disease and thrombosis. Recent studies reported that the tomato extract Fruitflow inhibited platelet function, but the molecular mechanism is still unclear. The present study used proteomics to quantitatively analyze the effect of fruitflow on the inhibition of collagen-stimulated platelets and validated the involvement of several signaling molecules. Fruitflow significantly inhibited human platelet aggregation and P-selectin expression that were induced by collagen. Proteomics analysis revealed that compared fruitflow-treated collagen-stimulated platelets with only collagen-stimulated platelets, 60 proteins were upregulated and 10 proteins were downregulated. Additionally, 66 phosphorylated peptides were upregulated, whereas 37 phosphorylated peptides were downregulated. Gene Ontology analysis indicated that fruitflow treatment downregulated phosphoinositide 3-kinase (PI3K)/protein kinase B and guanosine triphosphatase-mediated signal transduction in collagen-activated platelets. Biological validation indicated that fruitflow decreased Akt, glycogen synthase kinase 3ß, p38 mitogen-activated protein kinase (MAPK), and heat shock protein (Hsp27) phosphorylation in collagen-stimulated platelets. Fruitflow recovered cyclic adenosine monophosphate levels in collagen-activated platelets and reduced protein kinase A substrate phosphorylation that was induced by collagen. These findings suggest that fruitflow is a functional food that can inhibit platelet function, conferring beneficial effects for people who are at risk for platelet hyperactivity-associated thrombosis.

Resveratrol Confers Vascular Protection by Suppressing TLR4/Syk/NLRP3 Signaling in Oxidized Low-Density Lipoprotein-Activated Platelets.

This study investigated the effect of resveratrol on Toll-like receptor 4- (TLR4-) mediated matrix metalloproteinase 3 (MMP3) and MMP9 expression in oxidized low-density lipoprotein- (ox-LDL-) activated platelets and the potential molecule mechanism. Human platelets were used in the present study. The results showed that resveratrol suppressed TLR4, MMP3, and MMP9 expression in ox-LDL-activated platelets. The TLR4 inhibitor CLI-095 also inhibited MMP3 and MMP9 expression and secretion in ox-LDL- and lipopolysaccharide- (LPS-) activated platelets. The combination of resveratrol and CLI-095 synergistically suppressed MMP3 and MMP9 expression in ox-LDL- and LPS-activated platelets. These findings suggest that the resveratrol-induced inhibition of MMP3 and MMP9 expression is linked to the suppression of TLR4 activation. Resveratrol also suppressed spleen tyrosine kinase (Syk) phosphorylation and nucleotide-binding domain leucine-rich repeat containing protein 3 (NLRP3) expression and IL-1ß secretion in ox-LDL- and LPS-treated platelets. The coimmunoprecipitation results showed that resveratrol inhibited the binding of Syk and NLRP3. Finally, resveratrol reduced vascular senescence cells and the expression of TLR4, MMP3, and MMP9 and prevented alterations of vascular structure in 52-week-old mice. Our findings demonstrated that resveratrol decreased inflammatory protein expression and improved vascular structure in aged mice. Resveratrol inhibited the expression of TLR4 and secretion of MMP3, MMP9, and IL-1ß. The mechanism of action of resveratrol appears to be associated with the inhibition of TLR4/Syk/NLRP3 activation in ox-LDL-activated platelets.

Thioredoxin 1 downregulates MCP-1 secretion and expression in human endothelial cells by suppressing nuclear translocation of activator protein 1 and redox factor-1.

To know whether thioredoxin 1 (Trx1) works for an antioxidant defense mechanism in atherosclerosis, the effect of Trx1 on the release of monocyte chemoattractant protein-1 (MCP-1), a potent chemoattractant for recruitment and accumulation of monocytes/macrophages in the intima of artery vessel, was investigated in human endothelial-like EA.hy 926 cells. It was found that overexpression of Trx1 suppressed, whereas knockdown of endogenous Trx1 enhanced, oxidized low-density lipoprotein (oxLDL)-stimulated MCP-1 release and expression in the cells. It was also observed that overexpression of Trx1 suppressed, whereas depletion of endogenous Trx1 greatly promoted, nuclear translocation of c-Jun and the redox factor-1 (Ref-1). Electrophoretic mobility shift assay showed significantly reduced DNA-binding activity of activator protein-1 (AP-1) in Trx1-overexpressing cells but apparently enhanced DNA binding activity of AP-1 in Trx1-knockdown cells, indicating that nuclear Ref-1 rather than Trx1 itself finally dominates the regulation of AP-1 activity, although Trx1 is considered to upregulate AP-1 activity. It was also observed that Trx1 depressed intracellular generation of reactive oxygen species (ROS). Diphenyleneiodonium (DPI), the inhibitor of NADPH oxidase, suppressed MCP-1 secretion, whereas transient expression of Nox1 enhanced transcription of MCP-1 in endothelial cells. Assays with AP-1 and MCP-1 luciferase reporters further demonstrated that transient expression of Trx1 significantly depressed the transcriptional activity of c-Jun/c-Fos and consequent MCP-1 transcription. This study suggests that Trx1 inherently suppresses MCP-1 expression in vascular endothelium and may prevent atherosclerosis by depressing MCP-1 release. Besides the suppression of intracellular ROS generation, the inhibition of nuclear translocation of AP-1 and Ref-1 are mainly responsible for the downregulation of MCP-1 by Trx1.

Resveratrol Inhibits MMP3 and MMP9 Expression and Secretion by Suppressing TLR4/NF-κB/STAT3 Activation in Ox-LDL-Treated HUVECs.

AIM: Resveratrol is a natural plant polyphenol. The present study investigated the effects of resveratrol on the Toll-like receptor 4- (TLR4-) mediated expression and secretion of matrix metalloproteinases (MMPs) in oxidized low-density lipoprotein- (ox-LDL-) treated human umbilical vein endothelial cells (HUVECs). METHODS: Protein expression was analyzed by immunoblotting. The secretion of MMPs was measured by an enzyme-linked immunosorbent assay. The animal experiments were performed with and without resveratrol treatment in high-fat chow-fed mice. RESULTS: Resveratrol inhibited the expression of TLR4, MMP3, and MMP9 in ox-LDL- and lipopolysaccharide- (LPS-) treated HUVECs. Resveratrol reduced the secretion of MMP3 and MMP9 that was induced by ox-LDL and LPS. The TLR4 inhibitor CLI-095 similarly suppressed the expression and secretion of MMP3 and MMP9 in ox-LDL- and LPS-treated HUVECs. Resveratrol attenuated the phosphorylation of the transcription factors nuclear factor-κB (NF-κB) and signal transducer and activator of transcription 3 (STAT3) that was induced by ox-LDL and LPS. Resveratrol recovered Sirt1 expression. In the animal experiments, resveratrol decreased TLR4 expression in the aorta, MMP9 levels in plasma, and vascular structural changes in high-fat chow-fed mice, with no significant effect on plasma MMP3 levels. CONCLUSION: Resveratrol inhibited the TLR4-mediated expression and secretion of MMP3 and MMP9 in ox-LDL-treated HUVECs. The mechanism of action of resveratrol may be associated with the suppression of NF-κB and STAT3 phosphorylation and restoration of Sirt1 expression. Resveratrol exerts protective effects against vascular structural changes in high-fat chow-fed mice.