Viability and oxidative response of human colorectal HCT-116 cancer cells treated with visfatin/eNampt in vitro.

Visfatin/eNampt is a novel adipokine, secreted by visceral and subcutaneous fat, which could be involved in the development of obesity-associated cancer. Only few studies revealed reactive oxygen species (ROS)-dependent action of visfatin in endothelial cells, myotubes and melanoma cells. The potential pro-apoptotic properties of visfatin/eNampt in human colorectal HCT-116 cells remain unknown. The aim of the study was to examine the effects of visfatin/eNampt on cell viability along with the determination of apoptosis/necrosis extent and ROS level in HCT-116 cells. Additionally antioxidant enzymes' activities (i.e catalase (CAT), gluthatione peroxidase (GSH-Px)), and lipid peroxidation intensity in HCT-116 cells line was evaluated. Viability of HCT-116 cells was decreased after visfatin/eNampt treatment for 24 hours. The number of apoptotic cells in tested cells treated with increasing visfatin/eNampt concentrations (10, 100, 250 ng/ml) was elevated compared to untreated cells (6.4%, 9.7%, 16% vs. 3.2%; respectively). After 24 hours in the visfatin/eNampt treated group (10 - 100 ng/ml) CAT and GSH-Px activities significantly increased and this observation was accompanied by the decrease of ROS level when compared to the control group. Interestingly ROS level (using DCF detection technique) and lipid peroxidation ratio were increased in cells stimulated by visfatin/eNampt in concentration of 250 ng/ml along with the decreased activity of selected antioxidant enzymes when compared to remaining study groups, including control. We concluded that visfatin/eNampt induces decrease of cell viability and apoptosis boost in human colorectal cancer HCT-116 cells line. Visfatin/eNampt affected the level of ROS as well as antioxidant capacity, however the association of ROS level and apoptosis rate was not linear. The role for visfatin/eNampt in cancer redox status in vitro may provide a greater insight into the association between fat derived visfatin/eNampt and its endocrine action in colorectal carcinoma cells.

Eptifibatide infusion versus placebo in high risk patients with non-st segment elevation acute coronary syndromes managed with urgent coronary artery bypass graft surgery. a prospective multicenter randomized placebo-controlled clinical trial.

AIM: This randomized prospective clinical trial aimed to evaluate safety and efficacy of preoperative use of eptifibatide in high risk patients with non--ST--segment elevation acute coronary syndrome (NSTE--ACS), requiring urgent coronary artery bypass graft surgery (CABG). METHODS: A total of 140 patients with NSTE--ACS eligible for urgent surgical revascularization received either eptifibatide (bolus plus infusion) 12--48 hours prior to surgery (n=72 patients) or placebo (normal saline; n=68 patients) followed by routinely administered enoxaparin and aspirin. Patients were regarded as unsuitable for percutaneous coronary intervention by the heart team. CABG was performed 4 hours after discontinuation of eptifibatide or placebo infusion. The primary end point was major adverse cardiac and cerebrovascular events (MACCE) defined as death, nonfatal myocardial infarction (MI), stroke and the need for re--hospitalization due to recurrent ischaemia at 12 months follow--up. Secondary endpoints included MACCE rate at 1 month, bleeding complications, platelet inhibition efficacy and correlation of platelet activity with MACCE rate. RESULTS: Cumulative one year MACCE rate was 35% vs 14% in the control and treated group respectively (p=0.012). Mortality rate at 30 days follow--up was 10% vs 3% (p=0.021) and was not changed at 12 months follow--up. There was a significant difference between both groups regarding perioperative MI (22% vs. 8%, p=0.03). The rates of stroke, blood loss and blood transfusion were similar in both groups. CONCLUSION: Preoperative use of eptifibatide vs. placebo is linked to significantly reduced 12--month MACCE rate in patients with NSTE--ACS requiring urgent CABG, while it simultaneously seems not to confer a greater risk of postoperative bleeding.

Changes in subcellular localization of visfatin in human colorectal HCT-116 carcinoma cell line after cytochalasin B treatment.

The aim of the study was to assess the expression and subcellular localization of visfatin in HCT-116 colorectal carcinoma cells after cytokinesis failure using Cytochalasin B (CytB) and the mechanism of apoptosis of cells after CytB. We observed translocation of visfatin's antigen in cytB treated colorectal carcinoma HCT-116 cells from cytosol to nucleus. Statistical and morphometric analysis revealed significantly higher area-related numerical density visfatin-bound nano-golds in the nuclei of cytB-treated HCT-116 cells compared to cytosol. Reverse relation to visfatin subcellular localization was observed in un-treated HCT-116 cells. The total amount of visfatin protein and visfatin mRNA level in HCT-116 cells was also decreased after CytB treatment. Additionally, CytB significantly decreased cell survival, increased levels of G2/M fractions, induced bi-nuclei formation as well as increased reactive oxygen species (ROS) level in HCT-116 cells. CytB treatment showed cytotoxic effect that stem from oxidative stress and is connected with the changes in the cytoplasmic/nuclear amount of visfatin in HCT-116 cells.

Exogenous administration of visfatin affects cytokine secretion and increases oxidative stress in human malignant melanoma Me45 cells.

Visfatin has recently been established as a novel adipokine that is predominantly expressed in visceral fat. Recombinant visfatin has immunomodulating properties, which can activate human leukocytes in vitro to induce cytokine production (IL-1ß, TNF-α, and IL-6). Only few studies have investigated the effect of visfatin on prostate, breast, ovarian cancer as well as astrocytoma cell biology. There have been no studies on the cytokine secretion in human melanoma cells in response to visfatin stimulation along with intracellular protein kinases inhibitors. ELISA assay was performed in supernatants of Me45 cells stimulated with visfatin in the presence or the absence of specific pharmacological inhibitors of the indicated protein kinases (p38, MEK 1, PI3k and JAK kinase) and nuclear factor kappa B (NK-κB) inhibitor. Intracellular reactive oxygen species level was measured in 2', 7'-dichlorodihydrofluorescein diacetate (H2DCF-DA)-loaded cells using a fluorescent measurement system. For determination of NF-κB activation, activated NF-κB p65 subunit was determined using an EZ-TFA-detect chemiluminescent transcription factor assay. We report that visfatin led to the significant increase in IL-6 and IL-8 level in culture supernatants of human malignant melanoma Me45 cells. Additionally visfatin resulted in the increase of the intracellular reactive oxygen species level. PI3k and NF-κB pathways were activated upon visfatin stimulation. The results may reflect the fact that PI3k pathway stimulation by visfatin may further lead to NF-κB activation and pro-inflammatory response.

The effects of 1 month antihypertensive treatment with perindopril, bisoprolol or both on the ex vivo ability of monocytes to secrete inflammatory cytokines.

INTRODUCTION: Monocytes are key elements in pathogenesis of atherosclerosis and inflammation. The data regarding associations between antihypertensive treatment and monocytes' function are still lacking. The aim of this study was to evaluate the influence of antihypertensive drugs (bisoprolol, perindopril or both) in patients suffering from mild to moderate hypertension. PATIENTS AND METHODS: The study population consisted of 67 patients divided into 3 groups (2 consisted of patients with Grade I essential hypertension and one consisted of patients with Grade II essential hypertension). At baseline and 1 month after treatment we performed 24-h ambulatory noninvasive blood pressure monitoring and measured IL-1beta, IL-6, IL-10, MCP-1 and TNF-alpha in a medium derived from LPS-stimulated monocytes' culture. RESULTS: Both monotherapies with bisoprolol or perindopril were equally effective in lowering blood pressure (reduction in mean 24-h systolic blood pressure 12.07 vs. 15.91 mmHg, p = 0.678). Antihypertensive treatment led to significant decrease in IL-1b, IL-6, MCP-1 and TNF-alpha concentration and significant rise in IL-10 level compared to the baseline levels and the decrease was associated with reduction in blood pressure. CONCLUSIONS: Bisoprolol and perindopril effectively reduced elevated blood pressure. As a result, an alteration in cytokine net was observed at the end of the study. These results support the concept of possible anti-inflammatory effects of antihypertensive drugs (e.g., perindopril and bisoprolol).