Katarzyna Wojdan

IL-33 and IL-4 impair barrier functions of human vascular endothelium via different mechanisms

The vascular endothelium forms a barrier that controls flow of solutes and proteins and the entry of leukocytes into tissue. Injured tissue releases IL-33, which then alarms the immune system and attracts Th2 cells, thus increasing local concentration of IL-4. The aim of the study was to assess the influence of IL-33 and IL-4 on barrier functions of the human endothelium, expression of tight and adherent junction proteins, apoptosis and adhesive molecule surface expression in human endothelium in order to describe the mechanism of this effect. IL-33 and IL-4 decreased endothelial integrity and increased permeability. When added together, both cytokines lowered the endothelial integrity twice as much as used alone. This effect was accompanied by the down-regulation of occludin and VE-cadherin mRNA expression. Additionally, IL-4, but not IL-33, induced cell apoptosis. Both IL-33 and IL-4 showed the additive potency to down-regulate VE-cadherin mRNA expression. IL-33, unlike IL-4, increased the surface expression of ICAM-1, but not PECAM-1 in endothelial cells. Our results indicate that IL-33 may reversibly destabilize the endothelial barrier, thus accelerating the supply with immunomodulators and assisting leukocytes to reach wounded tissue. However, extended and less-controlled down-regulation of endothelial barrier, which may be a consequence of IL-33-initiated, but in fact IL-4-induced apoptosis of endothelial cells, may be deleterious and may eventually lead to the aggravation of inflammatory processes and the prolongation of tissue dysfunction.

The effect of melatonin on circadian blood pressure in patients with type 2 diabetes and essential hypertension

Introduction The aim of this study was to evaluate the effect of melatonin on blood pressure in patients with essential hypertension receiving medical treatment and with type 2 diabetes in good metabolic control. Material and methods The study lasted 8 weeks. Patients were equipped with a 24-hour ambulatory blood pressure monitor and took melatonin (3 mg a day in the evening) for 4 weeks. The patients were divided into four groups: group 1 (n = 32) including dippers, group 2 (n = 34) non-dippers treated with melatonin; and two control groups: group 3 (n = 28) including dippers and group 4 (n = 30) non-dippers treated without melatonin. After 4 weeks patients took melatonin for the next 4 weeks (5 mg a day). In each visit were analyzed: systolic, diastolic and mean blood pressure in both day and night time. Results We observed that 29.5% non-dippers (n = 10) treated with melatonin in a dose of 3 mg/day achieved features of dippers compared to control group (p < 0.05). Five mg of melatonin per day restored normal diurnal blood pressure rhythm in 32.4% non-dippers (n = 11, p < 0.05). In non-dippers treated with melatonin significant decreases of diastolic, systolic and mean night blood pressure values (p < 0.05) were observed. Conclusions More than 30% of non-dippers with type 2 diabetes treated with melatonin were restored to the normal circadian rhythm of blood pressure. The effect of melatonin in both doses (3 mg and 5 mg) was significant for non-dippers only and included nocturnal systolic, diastolic and mean arterial pressure.

The effect of 7-ketocholesterol and 25-hydroxycholesterol on the integrity of the human aortic endothelial and intestinal epithelial barriers

Objective and designThe damage of barrtier tissues, such as the vascular endothelium and intestinal epithelium, may lead to disturbances of local immune homeostasis. The aim of the study was to assess and compare the effect of oxidized cholesterols (7-ketocholesterol and 25-hydroxycholesterol) on the barrier properties of human primary aortic endothelium (HAEC) and intestinal epithelium Caco-2 cells using a real-time cell electric impedance sensing system (RTCA-DP).Materials and methodsHAEC and Caco-2 cells were stimulated with 7-ketocholesterol and 25-hydroxycholesterol by the RTCA-DP system. Apoptosis was assessed by flow cytometry and cell monolayer morphology was assessed under a light microscope.Results7-ketocholesterol decreased impedance (nCI) in both the endothelium and epithelium. However, the decrease was more profound in the endothelium. Similarly, although 25-hydroxycholesterol decreased nCI in both the endothelium and epithelium, the effect was weaker than that of 7-ketocholesterol, which caused extensive damage to the endothelial monolayer, while 25-hydroxycholesterol caused partial damage and did not affect the epithelial monolayer. 7-ketocholesterol, but not 25-hydroxycholesterol, increased endothelial cell apoptosis and decreased the viability of endothelial cells. However, 7-ketocholesterol and 25-hydroxycholesterol decreased epithelial cell apoptosis and increased viability.ConclusionOxidized cholesterols destroy the HAEC, but not the Caco-2 epithelial barrier, via cell apoptosis dependent on the site of oxidation. Damage to the endothelium by oxidized cholesterol may disrupt local homeostasis and provide open access to inner parts of the vascular wall for lipids, other peripheral blood-derived agents, and immune cells, leading to inflammation and atherogenesis.

Comprehensive insight into immune regulatory mechanisms and vascular wall determinants of atherogenesis – emerging perspectives of immunomodulation

For many years atherosclerosis was believed to be the passive accumulation of cholesterol in vessel walls. Today the picture is more complex, as immune processes occur in atherogenesis. Considerable attention is focused on the particular role of adaptive immune responses orchestrated by T cell subsets. Since the role of Th1/Th2 balance and Th1 cell domination in atherogenesis is already known, the involvement of regulatory T lymphocytes and recently described Th17 cells raises new concerns. On one hand, each of these cells may specifically drive responses of vascular wall tissues and immune cells; however, they are subject to the control of a plethora of tissue- and pathogen-derived agents. Due to ineffective tissue regeneration, remodeling of the vascular wall occurs. The understanding of the immune regulatory network gives perspectives of innovative immunomodulatory therapies of atherosclerosis and the prevention of its complications, such as coronary artery disease.

The effect of oxidized cholesterol on barrier functions and IL-10 mRNA expression in human intestinal epithelium co-cultured with dendritic cells in the transwell system.

The intestinal epithelium is exposed to oxygenated cholesterol products present in foodstuffs. In vitro studies demonstrate the effect of oxysterols on cytokine release by intestinal cells cultured alone. However, physiologically, the response of the intestinal epithelium to external agents occurs in the presence of dendritic cells (DCs). The aim of the study was to analyze the effect of 7-ketocholesterol on the barrier functions and IL-10 mRNA expression of Caco-2 cells in the presence of DCs, and secondly, on IL-10 mRNA expression in DCs. Caco-2 cells were co-cultured with monocyte-derived dendritic cells and induced with 7-ketocholesterol in a transwell system. DCs did not affect the transepithelial electrical resistance (TER) of the Caco-2 cell monolayer, but increased IL-10 mRNA expression in Caco-2 cells. 7-ketocholesterol decreased the TER of Caco-2 cells co-cultured with DCs and diminished IL-10 mRNA expression in Caco-2 cells induced by the presence of DCs. IL-10 mRNA expression fell in DCs co-cultured with Caco-2 cells after treatment with 7-ketocholesterol. Oxidized cholesterols present in gut mucosa may contribute to the decrease of epithelial barrier functions and the inappropriate development of an inflammatory response to food compounds.

Innate lymphoid cells type 2 – emerging immune regulators of obesity and atherosclerosis

The low-grade inflammation present in obese visceral adipose tissue impairs glucose metabolism, and contributes to the development of insulin resistance and weight gain. Immune processes occurring in response to the deposition of cholesterol within the vascular walls support atherosclerotic plaque growth and contribute to the cardiovascular complications. In both the obese adipose tissue and the atherosclerotic plaque, the Th1-type immune environment dominates over the Th2/Treg-type due to the overproduction of pro-inflammatory cytokines (IFN-γ, IL-6, TNF-α) and the deficiency of Th2-type processes and interleukins (IL-4, IL-5, IL-10, IL-13). So far, Th2 cells and eosinophils have been considered as the main providers of Th2-type mediators and the basis of Th2-type immunity in tissues. However, recently discovered innate lymphoid type 2 cells (ILC2s), which infiltrate lean visceral adipose tissue and the vascular wall, are believed to orchestrate local Th2-like immune responses. Upon activation by tissue-derived IL-33 and IL-25, ILCs2 secrete mostly IL-4, IL-5, IL-9 and IL-13: cytokines responsible for the accumulation of eosinophils and polarization of alternatively-activated macrophages, which altogether create the beneficial anti-inflammatory and metabo-regulatory environment in the adipose tissue and the vascular wall. Consequently, ILC2s-orchestrated immune environment seems to prevent obesity and atherosclerosis. Thus, ILCs2 appear to be the emerging immune regulators of immune and metabolic homeostasis in both adipose tissue and the vascular wall.

Disorders of erythrocyte structure and function in hypertensive patients

Summary Background The prevalence of hypertension is growing at an alarming rate. Increasing attention is being focussed on the oxidative stress accompanying this disease. In this study we examined the impact of this disease on some parameters of erythrocytes and human blood plasma. Material/Methods We examined the impact of hypertension on some parameters of erythrocytes and human plasma. The study involved 13 patients with hypertension and 19 healthy subjects. We determined lipid peroxidation, SH groups concentration, antioxidants enzymes activity, ATPase activity, total antioxidant capacity, total cholesterol level and erythrocyte membrane fluidity. Results We found an increased level of lipid peroxidation and the concentration of SH groups in membrane proteins in patients with hypertension, and a decrease in the activity of catalase and superoxide dysmutase. No changes were observed in glutathione peroxidase and ATPase activity, level of total antioxidant capacity, total cholesterol level and fluidity of erythrocyte membranes. Conclusions These results suggest the existence of an impaired oxidative balance in hypertensive human erythrocytes.

The effect of interleukin-35 on the integrity, ICAM-1 expression and apoptosis of human aortic smooth muscle cells

BACKGROUND Interleukin-35 (IL-35) is a novel immunomodulatory cytokine produced by CD4+ 25+ foxp3+ regulatory T-cells (T regs). Vascular smooth muscle cells (VSMCs) are involved in local immune homeostasis and certain chronic inflammatory pathologies. The effect of IL-35 on electrical impedance reflecting tissue integrity, the surface expression of ICAM-1 and mRNA expression of IL-32, as well as apoptosis in human primary aortic smooth muscle cells (Ao-SMCs) was investigated. METHODS The influence of IL-35 on Ao-SMC integrity was assessed with the real-time cell electric impedance sensing system (RTCA-DP) based on normalized Cell Index (nCI). Additionally, Ao-SMCs were stimulated with IL-35 in order to assess ICAM-1 surface expression and apoptosis in flow cytometer. IL-32 mRNA expression was measured using real-time PCR. RESULTS We found that the nCI of Ao-SMCs induced with IL-35 was lower after 12, 24 and 48h of incubation than the nCI of unstimulated cells. IL-35 slightly enhanced ICAM-1 surface expression and increased IL-32 mRNA expression in Ao-SMCs after 24h of induction. However, IL-35 did not affect Ao-SMC apoptosis, necrosis or viability. CONCLUSION Our data suggest that IL-35 may be an agent affecting the inflammatory properties of AoSMCs and thus it may regulate immune homeostasis of the vascular wall. Hence, IL-35 may be a novel player affecting Ao-SMC-controlled arterial wall immune homeostasis.

IL-22 modulates inflammatory properties of human primary aortic smooth muscle cells

BACKGROUND IL-22 is expressed at barrier surfaces, which suggests its critical role in the maintenance of normal barrier homeostasis and tissue repair. IL-22 can both promote pathological inflammation and prevent the destruction of tissues. The functional outcomes of IL-22 on vascular smooth muscle cells, which are shown to regulate immune processes within the vascular wall and which are involved in certain pathologies, have not been analyzed. OBJECTIVES The effect of IL-22 on the expression of novel antiand pro-inflammatory and barrier disrupting cytokines, apoptosis and the expression of adhesive molecules in human primary aortic smooth muscle cells (AoSMC) was investigated. MATERIAL AND METHODS Human AoSMC were induced with IL-22 for 24 h in vitro. The influence of IL-22 on IL-35 subunits EBI3 and p35, IL-33, IFN-γ and VEGF mRNA expression in Ao-SMC were assessed using real-time PCR. Additionally, the surface expression of ICAM-1 and apoptosis of AoSMC were analyzed in the flow cytometer. RESULTS IL-22 caused a 2- and 3-fold increase of mRNA expression of the EBI3 and p35 IL-35 subunits, and a 40% decrease of IL-33 mRNA expression in AoSMC. Additionally, IL-22 decreased ICAM-1 expression on the surface of AoSMC by 30%. However, IL-22 affected neither IFN-γ and VEGF mRNA expression in AoSMC nor their apoptosis and viability. CONCLUSIONS Our data suggest that IL-22, which is released by Th22 and NK cells, may be an agent affecting the inflammatory response of AoSMC, and thus it may regulate immune homeostasis of the vascular wall.

Increased plasma concentrations of interleukin 35 in patients with coronary artery disease

Introduction Atherosclerosis leading to coronary artery disease (CAD) is a chronic inflammatory condition. Interleukin 35 (IL-35) released by regulatory T cells (Tregs) has been found to be associated with CAD in the Chinese population. However, nothing is known about the relation between IL-35 concentrations and cholesterol levels. The aim of the study was to assess the levels of IL-35 in CAD patients and healthy subjects from a Caucasian population, and to analyze the relationship between IL-35 and the levels of total cholesterol, low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol, left ventricular ejection fraction (LVEF), sex and postmenopausal status. Material and methods Thirty-one patients with CAD and 30 healthy controls were included in the study. Levels of plasma IL-35 were analyzed by ELISA. The LVEF was assessed by transthoracic echocardiographic examination. Plasma levels of cholesterol fractions and C-reactive protein (CRP) were assessed by immunoenzymatic methods. Results The CAD patients had higher levels of IL-35 as compared to healthy controls (58.1 ±16.6 pg/ml vs. 5.35 ±3.35 pg/ml; p < 0.001). IL-35 levels negatively correlated with total and LDL cholesterol concentrations (R = –0.31, p < 0.01) and positively correlated with HDL cholesterol in men (R = 0.53, p < 0.01). In women, IL-35 levels negatively correlated with LVEF (R = –0.29, p < 0.05) and positively with the duration of postmenopausal status (R = 0.55, p < 0.01). Conclusions These results suggest a possible association between high levels of IL-35 and CAD.