Effects of 17ß-estradioland raloxifene on endothelial OPG and RANKL secretion.

OBJECTIVES: This study aims to asses the effects of estradiol vs. raloxifene on the levels of osteoprotegerin and soluble Receptor Activator of Nuclear Factor kB Ligand (sRANKL) in Human Umbilical Vein Endothelial Cells (HUVEC) culture in standard and calcifying medium. MATERIAL AND METHODS: Human Umbilical Vein Endothelial Cells were isolated from human umbilical vein by standard method. The supernatant concentrations of osteoprotegerin (OPG) and sRANKL (ELISA) were determined after incubation with glicerophosphate, estradiol , raloxifene, glicerophoshate and estradiol, glicerophosphate and raloxifene in comparison with control group at four designated time points (0, 1, 2 and 4 days of incubation). RESULTS: Incubation of estradiol with HUVEC colony lowered the OPG level significantly after day 2 and 4. Meantime, the level of sRANKL was stable. Raloxifene added to standard growth medium also significantly lowered OPG concentration after day 4 only, with no impact on sRANKL concentration. When added to calcifying medium, both estradiol and raloxifene significantly changed OPG level during the experiment. In all treated groups OPG levels were lower than in groups exposed to calcifying medium only. Neither estradiol, nor raloxifene changed sRANKL levels during the experiment. CONCLUSIONS: Estradiol and raloxifene affect OPG secretion from endothelial cells in vitro which may suggest their modifying role in pathogenesis of vascular calcification in postmenopausal women.

Expression of MUC1 mucin in human umbilical vein endothelial cells (HUVEC).

Mucin 1 (MUC1) is a membrane-bound glycoprotein that is expressed by various epithelial cell types. MUC1 functions include modulation of cell adhesion, signal transduction, lubrication and hydration of epithelial surfaces, and their protection from infection. In this study we demonstrated that MUC1 is expressed in human umbilical vein endothelial cells (HUVECs) and could be released/shed from cellular membrane. MUC1 presence in these cells was verified using three methods: Western blotting, flow cytometry and metabolic labeling. We also showed that mucin expression is stimulated by proinflammatory cytokines: about a 2-fold increase was observed after TNF-α treatment and lower after IFN-γ alone and in combination with TNF-α treatment. It can be assumed that the presence of MUC1 in endothelial cells may have an important role in the interactions with different cell types in physiological and pathological processes.

Polyphenols action against oxidative stress formation in endothelial cells.

The aim of this study was to investigate the influence of epigallocatechin-3-gallate (EGCG), theaflavins (TFs) and black tea extract (BTE) on oxidative stress formation as well as on antioxidant system of human vein endothelial cells (HUVEC). HUVEC were incubated for 0,5 h with 100 mM tert-butyl hydroperoxide (t-BHP) for oxidative stress formation. The influence of EGCG, TFs, and BTE on oxidative stress and antioxidant system parameters was investigated by the pre-incubation for 2 h with 50 mg/mL of each compound. Half hour exposure to t-BHP caused statistically significant decrease in GSH-Px activity and in the content of GSH, vitamin A, vitamin E as well as tryptophan. Moreover, pretreatment of cells with t-BHP caused statistically significant increase in activities of Cu,Zn-SOD, GSSG-R and in the level of MDA and dityrosine. Pretreated with t-BHP endothelial cells, subjected to EGCG, TFs and black tea extract, are partially protected against oxidative activity of t-BHP causing statistically significant increase in GSH-Px activity, GSH and tryptophan level and decrease in MDA and dityrosine level in comparison with HUVEC pretreated with t-BHP group. These results indicate the beneficial effect of tea polyphenolic compounds on HUVEC antioxidant abilities and, in consequence, their protective effect in cell components.

[Influence of insulin-like growth factor 1 on PECAM-1 and ICAM-1 adhesion molecules expression in tumor necrosis factor alpha and hyperglycemia induced umbilical vein endothelial cells]. / Wplyw insulinopodobnego czynnika wzrostu 1 na ekspresji czasteczek adhezyjnych PECAM-1 i ICAM-1 na powierzchni komórek sródblonka naczyn zylnych pepowiny indukowanych czynnikiem martwicy nowotworu alpha oraz hiperglikemia.

UNLABELLED: Activation, proliferation and regeneration of endothelium form developmental background of many pathological states. In those processes adhesion molecules and growth factors are crucial. Insulin-like growth factor 1 (IGF-1) is an important regulator of vascular function, capable of expressing pleiotropic actions. Interactions between endothelial cells and growth factors are complex and environment dependent. Cytokines, including tumor necrosis factor alpha (TNF-alpha) activate endothelium cells during inflammatory process. Endothelial dysfunction, with enhanced adhesion molecules expression, can be caused also by hyperglycemia. Aim of the study was to assess the influence of IGF-1 on platelet/endothelial cell adhesion molecule 1 (PECAM-1; CD31) and intercellular adhesion molecule 1 (ICAM-1; CD54) expression on human umbilical vein endothelial cells (HUVECs) stimulated by hyperglycemia and TNF-alpha. MATERIAL AND METHODS: Material for the study was obtained from fresh umbilical cord. Cell cultures were incubated with IGF-1 concentrations of 50 ng/ml and 200 ng/ml, glucose concentrations of 11.1 mmol/l and 22.2 mmol/l, TNF-alpha concentrations of 10 ng/ml and 20 ng/ml, as well as combinations of above. Ratio of PECAM-1 and ICAM-1 expression revealing cells was obtained using flow cytometry method. In statistic analysis ANOVA test and Duncan's new multiple range test were used. RESULTS: It was demonstrated, that IGF-1 concentrations of 50 ng/ml, 200 ng/ml enhance PECAM-1 and ICAM-1 endothelial cells expression (respectively to 57.2 +/- 4.8%, 76.1 +/- 1.5% and 91.7 +/- 5.0%, 86.8 +/- 4.7%) compared to control group (respectively: 20.9 +/- 0.5% and 26.6 +/- 0.5%). Expression under influence of 10 ng/ml and 20 ng/ml TNF-alpha increases up to respectively: 78.4 +/- 0.9% and 86.5 +/- 0.7% for CD31 and respectively: 65.1 +/- 3.8% and 70.8 +/- 1.4% for CD54. Glucose concentration of 22.2 mmol/l increases CD31 expression to 44.6 +/- 5.2% and CD54 expression to 47.3 +/- 4.8%. Combination of influence of IGF-1 and hyperglycemia enhances CD31 expression from 82.2 +/- 0.6% up to 85.3 +/- 0.7%, whereas CD54 expression from 89.3 +/- 3.5% up to 94.4 +/- 0.5%, depending of concentrations. Combination of 20 ng/ml TNF-alpha with 50 ng/ml IGF-1 enhances expression of CD54 to 96.8 +/- 1.2%, with 200 ng/ml IGF-1 enhances CD31 and CD54 expression respectively to 90.5 +/- 2.3% and 96.6 +/- 0.6%. CONCLUSIONS: 1. It was concluded, that IGF-1, hyperglycemia and TNF-alpha enhance PECAM-1 and ICAM-1 adhesion molecules expression on HUVEC cells, activating the endothelium. 2. Combination of IGF-1, TNF-alpha and hyperglycemia influence synergistically enhances PECAM-1 and ICAM-1 expression on endothelial cells surface.

[Evaluation of insulin-like growth factor-1 influence on PECAM-1 and ICAM-1 adhesion molecules expression on human umbilical endothelial cells surface]. / Ocena wplywu insulinopodobnego czynnika wzrostu-1 na ekspresje czasteczek adhezyjnych PECAM-1 i ICAM-1 na powierzchni komórek sródblonka zyly pepowinowej.

UNLABELLED: Activation, proliferation and regeneration of endothelium form developmental background of many pathological states. In those processes adhesion molecules and growth factors are crucial. Insulin-like growth factor-1 (IGF-1) is important regulator of vascular function, expressing pleiotropic actions. Interactions between endothelial cells and growth factors are complex and environment dependent. THE AIM OF THE STUDY was to assess IGF-1 influence on PECAM-1 and ICAM-1 adhesion molecules expression on human umbilical endothelial cells (HUVEC). MATERIAL AND METHODS: Material for study was obtained from fresh umbilical cord. Cell lines were incubated with IGF-1 concentrations of 50 ng/ml and 200 ng/ml. Ratio of PECAM-1 and ICAM-1 expression revealing cells was obtained using the flow cytometry method. In statistic analysis ANOVA test and Duncan's new multiple range test were used. RESULTS: It was demonstrated, that IGF-1 concentrations of 50 ng/ml and 200 ng/ml enhance PECAM-1 and ICAM-1 endothelial cells expression (respectively to 57.2+/-4.8%, 76.1+/-1.5% and 91.7+/-5.0%, 86.8+/-4.7%) compared to control group (respectively: 20.9+/-0.5% and 26.6+/-0.5%). CONCLUSION: Insulin-like growth factor-1 enhances PECAM-1 and ICAM-1 adhesion molecules expression on HUVEC cells, activating the endothelium.

[Endothelium--characteristics and functions]. / Sródblonek--charakterystyka i funkcje.

Endothelium is more and more likely to be considered as the biggest endocrine gland of human body. According to number, mass, surface of endothelial cells in adolescent, ability of synthesis and release of substances with multidirectional biological functions, endothelium should be treated as integral organ. Endothelial cells form barrier between blood and smooth muscle cells, playing crucial role in regulation of vasomotorics, hemostasis, angiogenesis, inflammatory processes and immunology. Strategic localization places endothelial cells in very first line of contact with cells and substances migrating from blood to tissue. Endothelium, via large number of yet not in-depth known mechanisms, can react to changes in pressure, blood flow and gas concentration. It is an easy stimulated and fast reacting structure, capable of quick activation and change in its own function. Mechanical damage, or loss of functional integrity, disturbs homeostasis of microenvironment, leading to development of pathological states, like hypertension, atherogenesis, thrombotic lesions, disturbances in perfusion of tissue and organs. The aim of this study was rewiev of the role of endothelial cells in vasomotorics, hemostasis, tumorgenesis and angiogenesis.

[The influence of erythropoietin (Epo) on intercellular adhesion molecule-1 (ICAM-1, CD54) and platelet-endothelial cell adhesion molecule-1 (PECAM-1, CD31) expression on human umbilical vein endothelial cells (HUVEC) induced by tumor necrosis factor-alpha (TNF-alpha)]. / Wplyw erytropoetyny (Epo) na indukowana czynnikiem martwicy nowotworów-alpa (tnf-alpha) ekspresje czasteczek adhezji miedzykomórkowej-1 (ICAM-1, CD54) i czasteczek adhezji komórkowej plytek i sródblonka-1 (PECAM-1, CD31) na powierzchni komórek sródblonka ludzkiej zyly pepowinowej (HUVEC).

UNLABELLED: The latest research proved that erythropoietin (Epo), a sensitive to hypoxia physiological erythropoiesis regulator, used successfully to treat anemia, displays cytoprotective, angiogenic, anti-inflammatory, anti-oxidative and anti-apoptotic properties. The aim of the study was to examine the influence of Epo on intercellular adhesion molecule-1 (ICAM-1) and platelet-endothelial cell adhesion molecule-1 (PECAM--1) expression on human umbilical vein endothelial cells (HUVEC) induced by tumor necrosis factor-a (TNF-alpha). MATERIAL AND METHODS: Human umbilical vein endothelial cells were cultured in a standard medium (M 199). The experiment was performed five times on the forth passage of HUVEC culture. For stimulation TNF-alpha was used in concentrations: 10, 20, 40 ng/ml (6 hours) and Epo in concentrations: 10, 20, 40 IU/ml (24 hours). The expression level of ICAM-1 and PECAM-1 on HUVEC were quantified by flow cytometry. RESULTS: The Incubation of HUVEC with TNF-a significantly increased the ICAM-1 and PECAM-I expression. The cultures pretreated with Epo reduced ICAM-1 and PECAM-I expression induced by TNF-a from 70.0 +/- 3.94% to 59.3 +/- 0.60% and from 83.4 +/- 2.27% to 57.7 +/- 0.66% respectively for ICAM-1 and PECAM-1. CONCLUSIONS: Erythropoietin statistically significantly decreases ICAM-1 and PECAM-1 expression on HUVEC stimulated by TNF-alpha.

[Erythropoietin prevents oxidative stress and lipid peroxidation process in human umbilical vein endothelial cells induced by tumor necrosis factor alpha]. / Erytropoetyna zapobiega stresowi oksydacyjnemu indukowanemu czynnikiem martwicy nowotworów alpha oraz procesowi peroksydacji lipidów w komórkach sródblonka naczyn zylnych ludzkiej pepowiny.

UNLABELLED: According last research erythropoietin (Epo), glycoprotein known as a physiological regulator of erythropoiesis and used successfully to treat anemia, displays cytoprotective properties. It is supposed that the protective effect of EPO is based on its ability to oxidation-reduction stabilization of cells. AIM: The aim of the study was to examine the influence of oxidative stress and lipid peroxidation in human umbilical vein endothelial cells (HUVEC) induced by tumor necrosis factor-alpha (TNF-alpha). MATERIAL AND METHODS: Human umbilical vein endothelial cells were cultured in a standard medium (M 199). The experiment was performed five times on the forth passage of HUVEC culture. For stimulation TNF-alpha was used in concentrations: 10, 20, 40 ng/ml (6 hours) and erythropoietin in concentrations: 10, 20, 40 IU/ml (24 hours). In the HUVEC lysate malonyldialdehyde (MDA), lipid hydroperoxides and reduction gluthation (GSH) concentrations were measured by HPLC method. RESULTS: MDA concentration statistically significantly decreased from 16.48 +/- 1.21 to 14.40 +/- 0.72 nmol/mg protein and LOOH concentration from 73.00 +/- 5.44 do 68.86 +/- 1.89 nmol/mg protein in the cell's lysate due to the preincubation with Epo of HUVEC cells, stimulated with TNF-alpha. MDA and LOOH concentrations in the control culture were 12.91 +/- 1.02 nmol/mg protein and 57.80 +/- 6.16 nmol/mg protein respectively. The application of erythropoietin while the cells were being stimulated with TNF-alpha prevented the decrease in GSH concentration which was 34.77 +/- 0.70 nmol/mg protein in the control culture, 33.11 +/- 1.65 nmol/mg protein in the culture stimulated with TNF-alpha and 34.17 +/- 0.14 nmol/mg protein in culture preincubated with Epo and stimulated by TNF-alpha. CONCLUSIONS: Erythropoietin (Epo) prevents oxidative stress and lipid peroxidation process in human umbilical vein endothelial cells (HUVEC) induced by tumor necrosis factor-alpha.