ICAM-1 and VCAM-1 expression by endothelial cells grown on fibronectin-coated TCPS and PS

Small-diameter vascular grafts rapidly fail as a result of blood coagulation and platelet deposition. Endothelial cells lining the inner side of blood vessels can provide the graft lumen with an antithrombogenic surface. One of the remaining problems is cell detachment after restoration of blood flow, because of infiltration of leukocytes that respond to an inflammatory-like activation of the endothelial cells. This endothelial activation is possibly caused by the surface characteristics of the underlying polymer. To get more insight into the effects of the polymer surface on endothelial cell activation, we seeded human umbilical vein endothelial cells (HUVECs) in various densities and subsequently grew them on tissue culture polystyrene (TCPS; hydrophilic) and polystyrene (PS; hydrophobic) surfaces. To improve cell adhesion, surfaces were coated with purified fibronectin prior to cell seeding. During proliferation, the expressions of the leukocyte adhesion molecules ICAM-1 and VCAM-1 were determined. Results indicate that ICAM-1 expression is not influenced by the character of the polymer surface, and that VCAM-1 expression is slightly higher on the TCPS surface. Expressions of both adhesion molecules are influenced by the seeding density and time of proliferation. At low seeding densities (< or = 10,000 cells/cm(2)), a relatively low percentage of nonexogenously activated cells expressed ICAM-1 during the first 3 days of proliferation compared to higher seeding densities. Although less pronounced, this was also observed for the percentage of cells expressing VCAM-1. During proliferation, the amount of ICAM-1 per endothelial cell increased, whereas the expression of VCAM-1 remained low. The absence of large differences in leukocyte adhesion molecule expression by endothelial cells grown on TCPS or PS is possibly caused by coating of the surfaces with fibronectin. It is known that surface hydrophilicity influences protein adsorption. Although this had no or little effect on leukocyte adhesion molecule expression, endothelial cell growth was affected, because proliferation was slower on the hydrophobic PS.     

IL-10对脑缺氧复氧后干扰素调节因子-1表达的抑制作用

目的:探讨白介素-10(IL-10)对缺氧复氧后干扰素调节因子-1(IRF-1)表达的作用。方法:采用缺氧8h复氧2h法建立脑微血管内皮细胞和中性粒细胞共培养缺氧复氧模型。实验分组:正常对照组,缺氧复氧组、IL-10干预组,其中IL-10干预组根据浓度不同分为1μg/L组、10μg/L组和30μg/L组。RT-PCR和Western blot检测IRF-1表达的变化。结果:缺氧复氧组IRF-1表达较正常组显著升高(P0.05)。IL-10干预组IRF-1表达较缺氧复氧组显著下降,并与IL-10剂量有关,呈剂量依赖性,在30μg/L表达最低(P0.05)。结论:IL-10可显著抑制脑缺氧复氧后IRF-1的表达。     

Endothelial cells upregulate eosinophil superoxide generation via VCAM-1 expression

BACKGROUND: In vitro eosinophil (EOS) adhesion to recombinant human (rh)-vascular cell adhesion molecule (VCAM)-1 stimulates superoxide anion (O2-) generation and enhances formyl-methionyl-leucyl phenylalanine (FMLP)-activated O2- generation. Therefore, EOS adhesion via VLA-4 to VCAM-1 expressed on endothelium may be instrumental in the selective recruitment and function of EOS in airway inflammation. OBJECTIVE: We hypothesized that EOS interaction with endothelial cells expressing VCAM-1 will undergo an enhancement in inflammatory function. METHODS: To determine this possibility, human umbilical vein endothelial cells (HUVEC) were stimulated with either a combination of interleukin (IL)-4 and tumour necrosis factor (TNF)-alpha (100 pM) or medium alone for 24 h; the expression of adhesion proteins on HUVEC and their effect on EOS O2- generation was subsequently determined. RESULTS: As determined by both enzyme-linked immunosorbent assay and flow cytometry, IL-4 and TNFalpha acted synergistically to induce VCAM-1 expression on HUVEC. Treating HUVEC with IL-4/TNFalpha also increased EOS adhesion and primed subsequent FMLP (0.1 microM) activated EOS O2- generation. Although EOS adhesion was partially inhibited by both antialpha4 and antibeta2 monoclonal antibodies (MoAbs), O2- generation was completely inhibited by either antialpha4 integrin MoAb (HP1/2) or anti-VCAM MoAb (BBIG-V1). Furthermore, enhanced O2- generation, but not adhesion, associated with IL-4 + TNFalpha-treatment of HUVEC was inhibited when EOS were treated with the platelet activating factor (PAF)-antagonist WEB 2086 (20 microM), thus suggesting an involvement of PAF in priming EOS. However, paraformaldehyde fixation of IL-4/TFN-alpha treated HUVEC did not significantly alter EOS function. CONCLUSIONS: These results suggest EOS adhesion to endothelial cells via an VLA-4/VCAM-1 interaction may be important in the development of the function of this cell. Furthermore, our results suggest that modulation of EOS function involves two priming factors: EOS adhesion to HUVEC expressing VCAM-1 and PAF.     

Multitasking of interferon regulatory factor-4 in early B cells

Immunoglobulin heavy- and light-chain genes are rearranged in a temporally ordered manner. In this issue, Johnson et al. (2008) show that interferon regulatory factor-4 regulates light-chain gene rearrangement by activating enhancers and attenuating interleukin-7 signaling.     

The role of interferon regulatory factor-1 and interferon regulatory factor-2 in IFN-gamma growth inhibition of human breast carcinoma cell lines.

Interferon (IFN) regulatory factor-1 (IRF-1) and IRF-2 play opposing roles in the regulation of many IFN-gamma-inducible genes. To investigate the signal transduction pathway in response to IFN-gamma in light of differences in growth effects, we selected four human breast carcinoma cell lines based on a spectrum of growth inhibition by IFN-gamma. MDA468 growth was markedly inhibited by IFN-gamma, and it showed substantial induction of IRF-1 mRNA but little IRF-2 induction. SKBR3 showed little growth inhibition and little induction of IRF-1 mRNA but significant induction of IRF-2 mRNA. HS578T and MDA436 growth inhibition and IRF-1/IRF-2 induction were intermediate. All four cell lines showed intact receptor at the cell surface and Stat1 translocation to the nucleus by immunostaining. By EMSA, there were marked differences in the induced ratio of IRF-1 and IRF-2 binding activity between the cell lines that correlated with growth inhibition. Finally, antisense oligonucleotides specific for IRF-1 attenuated IFN-gamma growth inhibition in MDA436 and MDA468, confirming the direct role of IRF-1 in IFN-gamma growth inhibition. Induction of IRF-1 causes growth inhibition in human breast cancer cell lines, and induction of IRF-2 can oppose this. The relative induction of IRF-1 to IRF-2 is a critical control point in IFN-gamma response.     

锌指蛋白基因抑制内皮细胞粘附分子表达及单核细胞粘附

目的单核细胞粘附是动脉粥样硬化发生的重要阶段,研究锌指蛋白A20基因在氧化型低密度脂蛋白诱导内皮细胞粘附分子表达及单核细胞粘附中的作用。方法脂质体DOTAP转染pEGFPEHA20-N1于原代培养的脐静脉内皮细胞,经G418筛选,A20表达经免疫荧光鉴定。激光共聚焦显微镜及微管吸吮系统分别检测转染前后氧化型低密度脂蛋白诱导单核细胞粘附情况及内皮细胞粘附分子VCAM-1表达。结果氧化型低密度脂蛋白能显著提高单核细胞粘附力及内皮细胞VCAM-1表达,A20基因能显著抑制氧化型低密度脂蛋白诱导单核细胞粘附及内皮细胞VCAM-1表达。结论A20基因对动脉粥样硬化的防治可能有一定作用。     

Simvastatin reduces VCAM-1 expression in human umbilical vein endothelial cells exposed to lipopolysaccharide

Objective  

Reducing the expression of endothelial cell adhesion molecules (ECAMs) is known to decrease inflammation-induced vascular complications. In this paper we looked at whether statins can reduce inflammation-induced ECAM expression after lipopolysaccharide (LPS) treatment in endothelial cells.     

Experimental autoimmune thyroiditis in nonobese diabetic mice lacking interferon regulatory factor-1

Interferon regulatory factor-1 (IRF-1) is pivotal in the regulation of interferon (IFN)-mediated immune reactions, and studies suggest that IRF-1 is involved in the development of autoimmune diseases. IRF-1+/+, +/-, and -/- nonobese diabetic (NOD) mice were immunized with mouse thyroglobulin (mTg) to determine whether IRF-1 is required in experimental autoimmune thyroiditis (EAT), a murine model for Hashimoto's thyroiditis (HT). IRF-1-deficient mice developed EAT and anti-mTg antibodies comparable to IRF-1+/+ and +/- mice. Whereas both CD4+ and CD8+ T cells were found in thyroids of IRF-1+/+ mice, the latter was not in IRF-1-/- mice. Major histocompatibility complex class II antigen was comparably expressed in thyroids of IRF-1+/+ and -/- mice. Lack of IRF-1 resulted in decreased CD8+ T cell number in the spleen and reduced IFNgamma production by splenocytes. Our results suggest that IRF-1 is not pivotal in EAT in NOD mice.     

流体的不同流动方式对血管内皮细胞血管细胞粘附分子-1、核因子κB表达的影响

目的:探讨在低切应力时不同流态对粘附分子表达的影响。方法:置人脐静脉内皮细胞单层于平板流动腔内,经切应力0.2Pa层流或振荡流作用4h、18h,用组织免疫化学方法测其血管细胞粘附因子-1(VCAM-1)表达及核因子(NF-κB)核转位活性,原位杂交法检测VCAM-1mRNA表达。结果:在相同的低切应力作用下,层流组未见VCAM-1mRNA、VCAM-1有明显变化,而振荡流组则明显上调VCAM-1mRNA、VCAM-1表达分别为对照组2倍、2.4倍及NF-κB核转位活性明显高于对照。结论:VEC在低切应力作用下,振荡流明显上调VCAM-1表达,而层流无影响。     

CD4~+CD25~+调节T细胞对内皮细胞炎性因子分泌的影响

目的:探讨CD4+CD25+调节性T细胞(Tregs)对氧化型低密度脂蛋白(ox-LDL)诱导人脐静脉内皮细胞(HU-VECs)VCAM-1、MCP-1、IL-6表达的影响。方法:磁性细胞分离器(MACS)分离CD4+CD25+T细胞及CD4+CD25-T细胞。在ox-LDL作用下,将内皮细胞分别与anti-CD3mAb激活的CD4+CD25+T细胞,CD4+CD25-T细胞共培养24小时。分别应用流式细胞术、ELISA、real-timePCR测定Tregs对ox-LDL诱导损伤HUVECs炎性因子VCAM-1、MCP-1、IL-6表达的影响。应用Transwell小室及中和抗体实验观察Tregs作用于HUVECs的具体机制。结果:与对照组比较,Tregs细胞可显著抑制ox-LDL诱导损伤HU-VECs炎性因子VCAM-1、MCP-1、IL-6的表达;被Transwell隔离或加入中和性抗体anti-IL-10/anti-TGF-β后,Tregs对HUVECs的抑制作用可被部分逆转。结论:Tregs细胞可显著抑制ox-LDL诱导损伤HUVECs炎性因子的表达,其作用机制既依赖于细胞直接接触,又依赖于细胞因子。     

Colonic epithelial cells induce endothelial cell expression of ICAM-1 and VCAM-1 by a NF-kappaB-dependent mechanism.

Epithelial cells are positioned in close proximity to endothelial cells. A non-contact coculture system was used to investigate whether colonic epithelial cells activated with various cytokines are able to provide signals that can modulate ICAM-1 and VCAM-1 expression on endothelial cells. Coculture of human umbilical vein endothelial cells (HUVEC) and human microvascular endothelial cells (HMEC-1) with TNF-alpha/IFN-gamma-stimulated human colon epithelial cell lines led to a significant up-regulation of endothelial ICAM-1 and VCAM-1 expression. Increased ICAM-1 and VCAM-1 expression by endothelial cells was accompanied by an increase in endothelial cell NF-kappaB p65 and NF-kappaB-DNA-binding activity. Inhibition of endothelial NF-kappaB activation using the proteosome inhibitors MG-132 and BAY 11-7082 resulted in a significant decrease of ICAM-1 expression, indicating an important role for NF-kappaB in this response. This cross-talk may represent a biological mechanism for the gut epithelium to control the colonic inflammatory response and the subsequent immune cell recruitment during inflammation.     

HLA antigen expression on cultured human arterial endothelial cells

Flow cytometric analysis and cellular assays were used to determine constitutive and induced expression of class I HLA antigens and class II antigens encoded by the HLA-DR, -DQ and -DP subregions on cultured human arterial endothelial cells (HAEC) derived from transplant donors. Class I but no or minimal quantities of class II HLA antigens were found on HAEC. Prior incubation of HAEC with gamma-IFN increased class I HLA antigen expression and induced class II HLA antigen expression on HAEC. The induced expression of HLA-DQ was lower than that of HLA-DR, but both were significantly reduced in comparison to the frequency of these antigens on EBV transformed lymphoblastoid cell lines derived from the same donor. In addition, supernatants from class I and class II alloreactive clones were shown to induce class II antigen expression on HAEC. By PLT analysis, it was shown that these antigens are functionally capable of generating a lymphocyte response. In this regard, HAEC have proved to be a helpful tool in designing in vitro lymphocyte-endothelial cell studies.