Visfatin Promotes Monocyte Adhesion by Upregulating ICAM-1 and VCAM-1 Expression in Endothelial Cells via Activation of p38-PI3K-Akt Signaling and Subsequent ROS Production and IKK/NF-κB Activation.

BACKGROUND/AIMS: Visfatin is known to act as a mediator in several metabolic disorders, such as obesity, diabetes, and cardiovascular diseases. This study aimed to investigate the effect of visfatin on the adhesion of THP-1 monocytes to human vascular endothelial cells and the underlying mechanism. METHODS: Monocytes adhesion to endothelial cells was determined by using fluorescence-labeled monocytes. ICAM-1 and VCAM-1 expression in endothelial cells were measured by western blotting. Production of reactive oxygen species (ROS) was measured by using a fluorescent dye. The amounts of nuclear factor-kappa B (NF-κB) and phosphorylation of inhibitory factor of NF-κB (IκB) were determined by using western blot analysis. The translocation of NF-κB from the cytoplasm to the nucleus was determined by using immunofluorescence. RESULTS: Here we showed that visfatin significantly caused the upregulation of intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in endothelial cells, as well as enhanced monocyte adhesion to endothelial cells. Moreover, we found that inhibition of PI3K, Akt, and p38 MAPK activation significantly prevented visfatin-enhanced expression of ICAM-1 and VCAM-1 and monocyte adhesion to endothelial cells. Visfatin enhanced ROS production and IKK/NF-кB activation and then led to upregulation of ICAM-1 and VCAM-1 and enhanced monocyte adhesion to endothelial cells. These effects were also p38/PI3K/Akt-dependent. CONCLUSION: These results demonstrated that visfatin promoted monocyte-endothelial cell adhesion by increasing ICAM-1 and VCAM-1 expression via the activation of p38/PI3K/Akt signaling and downstream ROS production and IKK/NF-кB activation.

Association of 24 h-systolic blood pressure variability and cardiovascular disease in patients with obstructive sleep apnea.

BACKGROUND: To evaluate association of 24 h-systolic blood pressure (SBP) variability and obstructive sleep apnea (OSA) as defined by the apnea-hypopnea index ≥5/h; and association of 24 h-SBP variability and prevalent cardiovascular disease (CVD) in OSA patients. METHODS: Participants underwent polysomongraphy to evaluate the presence of OSA, and 24 h-ambulatory blood pressure monitoring was applied to evaluate 24 h-SBP variability as indexed by weighted 24 h-standard deviation (SD) of SBP. Between-group differences were evaluated in participants with and without OSA. Participants with OSA were divided into high and low 24 h-SBP variability groups and between-group differences were evaluated. RESULTS: Mean age of 384 participants was 50 years old and 42.2% had OSA. Mean 24 h-systolic/diastolic BP were 130/78 mmHg, with mean weighted 24 h-SD of systolic/diastolic BP were 12.9/7.3 mmHg. Compared to those without OSA, OSA participants had higher clinic-, 24 h-, daytime- and nighttime-SBP, and weighted 24 h, daytime- and nighttime-SD of SBP. Age, prevalent CVD and OSA, usage of angiotensin converting enzyme inhibitor/angiotensin receptor blocker, calcium channel blocker and diuretic were significantly associated with 24 h-SBP variability. In OSA patients, compared to those with low variability, participants with high variability had higher weighted 24 h, daytime- and nighttime-SD of SBP. After adjusted for covariates including clinic-SBP and 24 h-SBP, per 1-SD increment weighted 24 h-SD of SBP was associated with 21% increased prevalent CVD. CONCLUSIONS: Patients with newly-diagnosed OSA have higher 24 h-SBP variability compared to those without OSA; in OSA patients, increased 24 h-SBP variability is associated with increased prevalence of CVD.

High interleukin-12 production from stimulated peripheral blood mononuclear cells of type 2 diabetes patients.

UNLABELLED: Sugar control is important in patients with sepsis. Interleukin (IL)-12 induces the polarization of CD4(+) T cells to the T helper 1 (Th1) phenotype. Regulatory T (T(reg)) cells are important in immunity and disease. The aim of this work is to determine whether hyperglycemia or insulin alters IL-12 response in peripheral mononuclear cells (PBMCs). METHODS: The PBMCs from 15 type 2 diabetes mellitus (DM) patients and 13 healthy controls were used for cell analysis and culture with or without treatment by glucose and insulin or stimulation by lipopolysaccharide (LPS) for 1, 2, and 3 days. RESULTS: The IL-12 level in the supernatant of LPS-stimulated PBMCs in the DM patients was significantly higher than that of healthy controls from day 1 to day 3. Kinetic IL-12 responses of LPS-stimulated PBMCs in the DM patients from day 1 to day 3 were significantly higher than that in healthy controls. The LPS-stimulated PBMCs under glucose treatment produced more IL-12 in DM patients but this did not happen in healthy controls. In DM patients, insulin could suppress IL-12 production from stimulated PBMCs but not with additional glucose treatment. CONCLUSION: The PBMCs of LPS-treated DM patients produced more IL-12 than that of LPS-treated healthy controls did. Hyperglycemia influenced IL-12 response from PBMCs in DM patients to some degree during infection.

An injectable chitosan/dextran/ß -glycerophosphate hydrogel as cell delivery carrier for therapy of myocardial infarction.

Acute myocardial infarction (MI) is a common cardiovascular disease with high mortality. In this study, an injectable thermosensitive hydrogel of chitosan (CS)/dextran (DEX)/ß-glycerophosphate (ß-GP) loaded with umbilical cord mesenchymal stem cells (UCMSCs) was prepared for MI treatment. The good biocompatibility of hydrogels was confirmed by 3T3 cells and HUVECs culture study in vitro. HUVECs encapsulated in the hydrogels showed a cell delivery ability. Furthermore, the results indicated that hydrogel could encapsulate most of UCMSCs and the cells exhibited good viability in CS/ 1.0DEX/ß-GP hydrogels. The expression of cardiac markers of cTnI and Cx43 and signaling pathways of p-Akt and p-ERK1/2 were studied, and it showed that UCMSCs differentiate towards myocardium and has a great potential for therapeutic use of cardiac repair. In conclusion, the thermosensitive hydrogel of CS/1.0 DEX/ß-GP loaded UCMSCs was a promising candidate as cell delivery vehicle for cardiac repair to reconstitute damaged myocardium.

Arterial stiffness is associated with target organ damage in subjects with pre-hypertension.

INTRODUCTION: Present study was to evaluate whether increased arterial stiffness was associated with target organ damage in pre-hypertensive subjects. MATERIAL AND METHODS: Pre-hypertensive subjects enrolled and echocardiography was used to evaluate left ventricular hypertrophy (LVH) and the first morning urine was collected to evaluate albumin and creatinine ratio (ALB/Cr ratio). Carotid-femoral pulse wave velocity (cf-PWV) was measured. RESULTS: A total of 420 subjects were recruited and mean age was 42.6 years. Mean systolic/diastolic blood pressure (SBP/DBP) were 130 ±9 mm Hg and 85 ±4 mm Hg. The prevalence of albuminuria and left ventricular hypertrophy was 8.6 % and 11.7 %. Mean cf-PWV was 9.2 ±1.0 m/s, with arterial stiffness prevalence was 8.8%. Subjects with arterial stiffness had higher cf-PWV value (10.6 ±0.4 m/s vs. 8.7 ±0.6 m/s, p < 0.05), and ALB/Cr ratio (28.3 ±13.2 µg/mg vs. 23.1 ± 11.4 µg/mg, p < 0.05). Overall, with multivariate regression analysis, aging and arterial stiffness were significantly associated with pre-hypertension. With stepwise adjusted for potential covariates including age, male gender, fasting plasma glucose, presence of current cigarette smoking, dyslipidemia, statins and SBP, increased cf-PWV remained independently associated with left ventricular hypertrophy and albuminuria, with an increased odds of 41 % and 24 % (p < 0.05), respectively. CONCLUSIONS: In pre-hypertensive subjects, arterial stiffness is independently associated with LVH and albuminuria and cf-PWV may be a useful marker to identify target organ damage in pre-hypertensive subjects.

Apelin/APJ axis improves angiotensin II-induced endothelial cell senescence through AMPK/SIRT1 signaling pathway.

INTRODUCTION: Previous studies have shown that endothelial cell senescence is involved in cardiovascular diseases such as cardiac fibrosis, atherosclerosis and heart failure. Accumulating evidence indicates that apelin exerts protective effects on ageing-related endothelial dysfunction. In this study, we aim to investigate the role of the apelin/APJ axis in angiotensin II (AngII)-induced endothelium senescence and its associated mechanisms. MATERIAL AND METHODS: Senescence-related ß-gal activity assay and western blot were used to evaluate human umbilical vein endothelial cell (HUVEC) senescence. In addition, DCFH-DA staining was carried out to detect the generation of reactive oxygen species (ROS). A validated, high-sensitivity real-time quantitative telomeric repeat amplification protocol (RQ-TRAP) was applied to determine telomerase activity in HUVECs, and a CCK-8 assay was employed to measure cellular viability. RESULTS: AngII induced an increase in SA-ß-Gal-positive cells and upregulation on expression of P21 and PAI-1 compared to the control group (p < 0.05), while apelin against this process (p < 0.05). The protective effects were attenuated when APJ, AMPK and SIRT1 expression was knocked down (p < 0.05). Furthermore, apelin reduced AngII-induced ROS generation and enhanced telomerase activity in HUVECs (p < 0.05), which contributed to increased HUVEC viability as assessed by the CCK-8 assay (p < 0.05). CONCLUSIONS: The apelin/APJ axis improved AngII-induced HUVEC senescence via the AMPK/SIRT1 signaling pathway, and the underlying mechanisms might be associated with reduced ROS production and enhanced telomerase activity.

Additive effects of atorvastatin combined with sitagliptin on rats with myocardial infarction: a pilot study.

INTRODUCTION: Atorvastatin and sitagliptin are able to exert cardio-protective effects. However, whether atorvastatin plus sitagliptin could confer additive benefits for rats with myocardial infarction (MI) is unknown. MATERIAL AND METHODS: Forty rats with MI were produced and 37 surviving rats were randomly divided into atorvastatin (10 mg/kg daily, n = 9), sitagliptin (10 mg/kg daily, n = 9), combined (10 mg/kg daily atorvastatin plus 10 mg/kg daily sitagliptin, n = 9), and control groups (3 ml normal saline daily, n = 10). Fourteen days later, cardiac function was detected and fasting venous blood was sampled for lipid profiles and glucose evaluation. Cardiac tissues were used for hematoxylin-eosin staining, for interleukin-6 (IL-6) and tumor necrotic factor-α (TNF-α) evaluation, and for rho-associated kinase 2 (ROCK2) assessment. RESULTS: Fourteen days after MI, the inflammatory reaction regarding the degree of leukocyte infiltration and IL-6 and TNF-α expression in cardiac tissues was ameliorated in atorvastatin and sitagliptin groups compared to the control group (p < 0.05). In addition, ROCK2 was attenuated by either atorvastatin or sitagliptin (p < 0.05). Echocardiography showed that cardiac function was significantly improved with atorvastatin and sitagliptin therapy (p < 0.05). Overall, all these benefits were further enhanced by combined therapy, suggesting that atorvastatin combined with sitagliptin therapy has additive effects on reducing cardiac inflammation and improving cardiac function. No significant changes in lipid profiles or glucose were observed, suggesting that the benefits derived from atorvastatin and sitagliptin therapy might not depend on cholesterol and glucose modulation. CONCLUSIONS: In rats with MI, atorvastatin plus sitagliptin therapy provides additive effects for cardio-protection, and mechanisms operating in these processes may be due to ROCK2 diminishment.

Human Endothelial Progenitor Cell-Derived Exosomes Increase Proliferation and Angiogenesis in Cardiac Fibroblasts by Promoting the Mesenchymal-Endothelial Transition and Reducing High Mobility Group Box 1 Protein B1 Expression.

Myocardial fibrosis is a characteristic feature of cardiomyopathies. However, no effective strategies to attenuate cardiac fibrosis are currently available. Late-stage endothelial progenitor cells (EPCs) are precursors of endothelial cells (ECs) that repair the heart through a paracrine mechanism. In the present study, we tested whether EPC-derived exosomes regulate the differentiation of fibroblasts into ECs. We isolated late-stage EPCs from human peripheral blood (PB) and used immunofluorescence and flow cytometry to confirm their identity. Next, we isolated exosomes from the EPCs and characterized their morphology using electron microscopy and confirmed the expression of exosome-specific marker proteins using Western blots. We then investigated the in vitro effects of exosomes on the proliferation and angiogenesis of cardiac fibroblasts (CFs) and on the expression of the mesenchymal-endothelial transition (MEndT)-related genes and the myocardial fibrosis-regulated protein, high mobility group box 1 protein B1 (HMGB1). We found that human PB-EPC-derived exosomes enhanced the proliferation and angiogenesis of CFs in vitro. Furthermore, CFs stimulated with these exosomes showed increased expression of the EC-specific markers, like cluster of differentiation 31 and vascular endothelial growth factor receptor 2, and decreased expression of proteins involved in fibrosis, like alpha-smooth muscle actin, vimentin, collagen I, transforming growth factor-beta, and tumor necrosis factor-alpha. In addition, CFs stimulated with human PB-EPC-derived exosomes, inhibited the expression of HMGB1. Taken together, our study demonstrated that EPC-derived exosomes promote the proliferation and angiogenesis of CFs by inhibiting MEndT and decreasing the expression of HMGB1.

ATP-sensitive K⁺ channels contribute to the protective effects of exogenous hydrogen sulfide against high glucose-induced injury in H9c2 cardiac cells.

Hyperglycemia, as well as diabetes mellitus, has been shown to impair ATP-sensitive K+ (KATP) channels in human vascular smooth muscle cells. Hydrogen sulfide (H2S) is also known to be an opener of KATP channels. We previously demonstrated the cardioprotective effects exerted by H2S against high-glucose (HG, 35 mM glucose)-induced injury in H9c2 cardiac cells. As such, we hypothesized that KATP channels play a role in the cardioprotective effects of H2S against HG-induced injury. In this study, to examine this hypothesis, H9c2 cardiac cells were treated with HG for 24 h to establish a model of HG-induced insults. Our findings revealed that treatment of the cells with HG markedly decreased the expression level of KATP channels. However, the decreased expression of KATP channels was reversed by the treatment of the cells with 400 µM sodium hydrogen sulfide (NaHS, a donor of H2S) for 30 min prior to exposure to HG. Additionally, the HG-induced cardiomyocyte injuries, including cytotoxicity, apoptosis, oxidative stress and mitochondrial damage, were ameliorated by treatment with NaHS or 100 µM diazoxide (a mitochondrial KATP channel opener) or 50 µM pinacidil (a non-selective KATP channel opener) for 30 min prior to exposure to HG, as indicated by an increase in cell viability, as well as a decrease in the number of apoptotic cells, the expression of cleaved caspase-3, the generation of reactive oxygen species (ROS) and the dissipation of mitochondrial membrane potential (MMP). Notably, treatment of the H9c2 cardiac cells with 100 µM 5-hydroxydecanoic acid (5-HD, a mitochondrial KATP channel blocker) or 1 mM glibenclamide (Gli, a non-selective KATP channel blocker) for 30 min prior to treatment with NaHS and exposure to HG significantly attenuated the above-mentioned cardioprotective effects exerted by NaHS. Notably, treatment of the cells with 500 µM N-acetyl­L­cysteine (NAC, a scavenger of ROS) for 60 min prior to exposure to HG markedly reduced the HG-induced inhibitory effect on the expression of KATP channels. Taken together, our results suggest that KATP channels play an important role in the cardioprotective effects of exogenous H2S against HG-induced injury. This study also provides novel data demonstraring that there is an antagonistic interaction between ROS and KATP channels in HG-exposed H9c2 cardiac cells.

RASSF8 downregulation promotes lymphangiogenesis and metastasis in esophageal squamous cell carcinoma.

Lymphatic vessels are the major routes of human esophageal squamous cell carcinoma (ESCC) metastasis. Tumor cells secrete pro-lymphangiogenic factors to induce new lymphatic vessels, promoting lymph node metastasis. In this study, we show that RAS association domain family 8 (RASSF8) expression in ESCC clinical samples was inversely correlated with lymph node metastasis and patients survival. Tumor cells with low RASSF8 expression had higher apparent migratory ability, and promoted and lymphangiogenesis both in vitro and in vivo. RASSF8 downregulation enhanced VEGF-C expression and caused subcellular redistribution of p65 in ESCC. Our results show that RASSF8 acts as a tumor suppressor in ESCC and is a potential therapeutic target for preventing lymph node metastasis.

Glucose increases interleukin-12 gene expression and production in stimulated peripheral blood mononuclear cells of type 2 diabetes patients.

BACKGROUND: Lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs) of type 2 diabetes patients produce more interleukin (IL)-12 under glucose treatment. The aim of this study was to determine whether increased IL-12 response in hyperglycemic LPS-stimulated PBMCs is due to increased gene expression or osmolarity. METHODS: LPS-stimulated PBMCs of 13 type 2 diabetes patients and 8 healthy controls were used for culture in the presence or absence of glucose or mannitol for 24 h. The IL-12 gene expressions of PBMCs and IL-12 protein levels in supernatants were evaluated. RESULTS: After 24 h, the stimulated PBMCs of diabetes patients expressed more IL-12 mRNA and produced more IL-12 protein following glucose treatment than those without glucose treatment and with mannitol treatment. Stimulated PBMCs of controls did not express more IL-12 mRNA and produce more IL-12 protein following glucose treatment than those without glucose treatment and with mannitol treatment. CONCLUSIONS: Glucose increases the IL-12 production in stimulated PBMCs of diabetes patients through increased IL-12 gene expression.

Phase transformation of molecular beam epitaxy-grown nanometer-thick Gd2O3 and Y2O3 on GaN.

High quality nanometer-thick Gd2O3 and Y2O3 (rare-earth oxide, R2O3) films have been epitaxially grown on GaN (0001) substrate by molecular beam epitaxy (MBE). The R2O3 epi-layers exhibit remarkable thermal stability at 1100 °C, uniformity, and highly structural perfection. Structural investigation was carried out by in situ reflection high energy electron diffraction (RHEED) and ex-situ X-ray diffraction (XRD) with synchrotron radiation. In the initial stage of epitaxial growth, the R2O3 layers have a hexagonal phase with the epitaxial relationship of R2O3 (0001)(H)<1120>(H)//GaN(0001)(H)<1120>(H). With the increase in R2O3 film thickness, the structure of the R2O3 films changes from single domain hexagonal phase to monoclinic phase with six different rotational domains, following the R2O3 (201)(M)[020](M)//GaN(0001)(H)<1120>(H) orientational relationship. The structural details and fingerprints of hexagonal and monoclinic phase Gd2O3 films have also been examined by using electron energy loss spectroscopy (EELS). Approximate 3-4 nm is the critical thickness for the structural phase transition depending on the composing rare earth element.

[Effects of rhBMP-7 on osteoblast differentiation of murine bone marrow mesenchymal stem cells in vitro].

AIM: To study the effects of recombinant human bone morphogenetic protein-7 (rhBMP-7) on osteoblast differentiation of murine bone marrow mesenchymal stem cells (BMSCs). METHODS: The BMSCs were collected from murine bone marrow by density gradient centrifugation. Cells were adherent cultured and expanded in vitro. RhBMP-7 was added into the culture medium of the 3rd passage BMSCs. Five days later, we performed alkaline phosphatase (ALP) staining and detected ALP activity and osteocalcin (OC) content. Osteoblast differentiation marker gene including OC and collage I (Col-I) were assayed by RT-PCR. Total protein was isolated and the secretion of Col-I in the cells was measured by Western blotting. RESULTS: The staining intensity of ALP in the group with rhBMP-7 was stronger than that in the negative control group. ALP activity and OC content of rhBMP-7 group were obviously higher than that of the negative control group. The mRNA of OC and Col-I and the protein of Col-I were highly expressed in the group induced by rhBMP-7. CONCLUSION: RhBMP-7 can induce murine BMSCs to differentiate into osteoblast in vitro.

Comparison of plasma interferon-gamma and antigen 60 immunoglobulin G in diagnosing pulmonary Mycobacterium tuberculosis infection.

BACKGROUND: An elevated interferon-gamma (IFN-gamma) level has been reported in the plasma of patients with pulmonary tuberculosis. Serologic diagnosis of tuberculosis using Antigen 60 immunoglobulin G (A60 IgG) is a well-known diagnostic approach. This study evaluated plasma IFN-y compared to A60 IgG in diagnosing Mycobacterium tuberculosis. METHODS: This study recruited 65 patients with tuberculosis and 59 controls. The plasma levels of A60 IgG and IFN-gamma were measured using enzyme-linked immunosorbent assay kits. RESULTS: The cutoff values of IFN-gamma and A60 IgG tests were set at 0.137 pg/ml and 261.2 units. The sensitivity and specificity of the IFN-gamma test were 27.7% and 91.5%. The positive and negative predictive values of the IFN-gamma test were 53.4% and 78.3%. Moreover, the sensitivity and specificity of the A60 IgG test were 53.8% and 67.8%. The positive and negative predictive values of the A60 IgG test were 37.0% and 80.7%. Finally, the sensitivity and specificity of the combined A60 IgG and IFN-gamma tests were 64.6% and 62.7%. The positive and negative predictive values of the combined A60 IgG and IFN-gamma tests were 37.8% and 83.4%. CONCLUSIONS: The combined tests did not achieve significant improvement in disease prediction. The A60 IgG test and IFN-gamma test had similar diagnostic value in pulmonary Mycobacterium tuberculosis infection.